研究者詳細 - 渡部　昌実

論文 - 渡部　昌実

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Circadian Rhythms Fluctuate the Treatment Effects of Intravesical Treatments on Rat Urinary Frequency Models 査読

Tomofumi Watanabe, Takuya Sadahira, Yusuke Tominaga, Yuki Maruyama, Naoya Nagasaki, Takanori Sekito, Kohei Edamura, Toyohiko Watanabe, Motoo Araki, Masami Watanabe

Veterinary Medicine International 2024 1 - 9 2024年5月

掲載種別：研究論文（学術雑誌）出版者・発行元：Hindawi Limited

Objectives. It is still not clear how the intravesical instillation of drugs affects rat urinary frequency. This study aimed to examine the dynamics of intravesical treatments’ treatment effect on rat urinary frequency models by real-time and extended monitoring using a novel continuous urination monitoring system. Methods. Nine eleven-week-old female Wistar rats were divided into three groups to receive intravesical instillation of 0.1% acetic acid (AA), 1.0% AA, or phosphate-buffered saline (PBS). Thirty minutes later, these drugs were voided, and rats were moved to a continuous urination monitoring system, UM-100. UM-100 monitored rat urination quantitatively and continuously for 24 hours. Rats were then euthanized, and histopathologic examinations using a damage score validated the severity of bladder inflammation. We used nine additional rats to determine the treatment effect of various drugs against the urinary frequency. These rats were also treated with 1.0% AA in the same way and divided into three groups (n = 3 each) to receive intravesical instillation of lidocaine, silver nitrate (AgNO₃), or dimethyl sulfoxide (DMSO), respectively. Thirty minutes later, rats were catheterized again and moved to the UM-100, and their voiding was monitored for 24 hours. Results. Intravesical instillation of AA increased the urinary frequency and decreased the mean voided volume (VV) in a concentration-dependent manner, with statistical significance at a concentration of 1.0% (urinary frequency; , mean VV; , respectively) compared with PBS. Histopathological analysis of these models demonstrated a significantly higher damage score of bladder mucosa in both 0.1% AA and 1.0% AA compared with PBS, with the severity in concordance with the clinical severity of urinary frequency (0.1% AA: , 1.0% AA: ). Moreover, intravesical instillation of lidocaine, AgNO₃, and DMSO decreased the urinary frequency. Continuous monitoring with UM-100 also demonstrated that the treatment effect of these intravesically instilled drugs occurred only at night. Conclusions. The extended monitoring of rat urination by UM-100 revealed a significant fluctuation in the treatment effect of intravesically instilled drugs between day and night. These findings may help establish novel therapies for urinary frequency.

DOI： 10.1155/2024/6505595

その他リンク： http://downloads.hindawi.com/journals/vmi/2024/6505595.xml

Trans men can achieve adequate muscular development through low-dose testosterone therapy: A long-term study on body composition changes. 査読国際誌

Yusuke Tominaga, Tomoko Kobayashi, Yuko Matsumoto, Takatoshi Moriwake, Yoshitaka Oshima, Misa Okumura, Satoshi Horii, Takuya Sadahira, Satoshi Katayama, Takehiro Iwata, Shingo Nishimura, Kensuke Bekku, Kohei Edamura, Morito Sugimoto, Yasuyuki Kobayashi, Masami Watanabe, Yuzaburo Namba, Yosuke Matsumoto, Mikiya Nakatsuka, Motoo Araki

Andrology 2024年4月

記述言語：英語掲載種別：研究論文（学術雑誌）

BACKGROUND: Transgender individuals undergo the gender-affirming hormone therapy (GAHT) to achieve physical changes consistent with their gender identity. Few studies are available on the long-term safety and efficacy of GAHT. OBJECTIVES: To investigate the long-term physical effects and the safety of the testosterone therapy for trans men and to assess the impact of differential hormone dose. MATERIALS AND METHODS: Trans men who initiated GAHT between May 2000 and December 2021 were included in this retrospective analysis. Physical findings (body mass index, body fat percentage (BFP), lean body mass (LBM), and grip strength), blood testing results (hemoglobin, hematocrit, uric acid, creatinine, total cholesterol, triglycerides, and total testosterone), and menstrual cessation were recorded. We assessed the effects of testosterone on body composition changes and laboratory parameters, comparing a low-dose group (≤ 62.5 mg/wk) to a high-dose group (> 62.5 mg/wk). RESULTS: Of 291 participants, 188 patients (64.6%) were in the low-dose group and 103 (35.4%) in the high-dose group. Cumulative menstrual cessation rates up to 12 months were not significantly different between groups. Both groups showed a decrease in BFP and an increase in LBM during the first year of therapy, followed by a slight increase in both over the long term. The high-dose group exhibited greater LBM gains during the first year. Higher hormone doses and lower initial LBM values were associated with LBM increases at 3 and 6 months (3 mo, P = 0.006, P < 0.001; 6 mo, P = 0.015, P < 0.001). There were no long-term, dose-dependent side effects such as polycythemia or dyslipidemia. CONCLUSION: Long-term GAHT for trans men is safe and effective. Low-dose testosterone administration is sufficient to increase LBM in trans men. Higher testosterone doses can lead to an earlier increase in muscle mass.

DOI： 10.1111/andr.13640

Establishment of Induced Pancreatic Stem Cells by Yes-Associated Protein 1 査読

Hirofumi Noguchi, Chika Miyagi-Shiohira, Yoshiki Nakashima, Yuka Onishi, Issei Saitoh, Masami Watanabe

Cell Transplantation 2024年1月

Exploration of the protein-dependent mechanism of Lactobacillus crispatus GAI98322 to prevent recurrent cystitis. 査読国際誌

Tomofumi Watanabe, Takuya Sadahira, Hidetada Hirakawa, Peng Huang, Tianyun Zhou, Takehiro Iwata, Takanori Sekito, Ayano Ishii, Masami Watanabe, Motoo Araki

Journal of infection and chemotherapy : official journal of the Japan Society of Chemotherapy 29 ( 10 ) 1001 - 1004 2023年6月

記述言語：英語掲載種別：研究論文（学術雑誌）

OBJECTIVES: To elucidate the mechanism of Lactobacillus crispatus (L. crispatus) suppositories to prevent patients from recurrent cystitis (RC), independent from viable-Lactobacilli-bacteria- and acid-dependent ones such as hydrogen peroxide and lactate. METHODS: We used the GAI98322 strain of L. crispatus in all experiments and pH-matched. cell-free culture supernatant of L. crispatus (CFCS) was collected. The growth inhibitory activity and the biofilm formation inhibitory activity of the CFCS against uropathogenic Escherichia coli (UPEC), Extended Spectrum beta (β) Lactamase producing (ESBL+) UPEC, and Pseudomonas aeruginosa (P. aeruginosa) was assessed by agar-disk diffusion tests and crystal violet assay. Also, CFCS was subjected to mass spectrometry to specify ingredients. RESULTS: The CFCS suppressed the proliferation of E. coli, ESBL + E. coli, and P. aeruginosa. Also, the CFCS at a concentration of 40% significantly impeded the biofilm formation of these three bacteria. The aggregation-promoting factor and Lysin was detected from CFCS. CONCLUSIONS: The cell-free supernatant from the GAI98322 strain of L. crispatus inhibits the growth/biofilm formation of broad pathogens by aggregation promoting factor and lysin, which may prevent hosts from RC regardless of the antimicrobial resistance of the pathogens and even under pH modulation.

DOI： 10.1016/j.jiac.2023.06.013

Pacsin 2-dependent N-cadherin internalization regulates the migration behaviour of malignant cancer cells 査読

Haymar Wint, Jianzhen Li, Tadashi Abe, Hiroshi Yamada, Yasutomo Nasu, Masami Watanabe, Kohji Takei, Tetsuya Takeda

J Cell Sci 136 ( 10 ) 2023年5月

The Genotypic and Phenotypic Characteristics Contributing to Flomoxef Sensitivity in Clinical Isolates of ESBL-Producing E. coli Strains from Urinary Tract Infections 査読

Kazuma Sakaeda, Takuya Sadahira, Yuki Maruyama, Takehiro Iwata, Masami Watanabe, Koichiro Wada, Motoo Araki

Antibiotics 12 ( 3 ) 522 - 522 2023年3月

掲載種別：研究論文（学術雑誌）出版者・発行元：MDPI AG

We carried out a molecular biological analysis of extended-spectrum β-lactamase (ESBL)-producing E. coli strains and their sensitivity to flomoxef (FMOX). Sequence type (ST) analysis by multilocus sequence typing (MLST) and classification of ESBL genotypes by multiplex PCR were performed on ESBL-producing E. coli strains isolated from urine samples collected from patients treated at our institution between 2008 and 2018. These sequences were compared with results for antimicrobial drug susceptibility determined using a micro-liquid dilution method. We also analyzed cases treated with FMOX at our institution to examine its clinical efficacy. Of the 911 E. coli strains identified, 158 (17.3%) were ESBL-producing. Of these, 67.7% (107/158) were strain ST-131 in ST analysis. Nearly all (154/158; 97.5%) were CTX-M genotypes, with M-14 and M-27 predominating. The isolated strains were sensitive to FMOX in drug susceptibility tests. Among the patient samples, 33 cases received FMOX, and of these, 5 had ESBL-producing E. coli. Among these five cases, three received FMOX for surgical prophylaxis as urinary carriers of ESBL-producing E. coli, and postoperative infections were prevented in all three patients. The other two patients received FMOX treatment for urinary tract infections. FMOX treatment was successful for one, and the other was switched to carbapenem. Our results suggest that FMOX has efficacy for perioperative prophylactic administration in urologic surgery involving carriers of ESBL-producing bacteria and for therapeutic administration for urinary tract infections. Use of FMOX avoids over-reliance on carbapenems or β-lactamase inhibitors and thus is an effective antimicrobial countermeasure.

DOI： 10.3390/antibiotics12030522

Novel extracellular role of REIC/Dkk-3 protein in PD-L1 regulation in cancer cells. 査読国際誌

Yuma Gohara, Nahoko Tomonobu, Rie Kinoshita, Junichiro Futami, Léna Audebert, Youyi Chen, Ni Luh Gede Yoni Komalasari, Fan Jiang, Chikako Yoshizawa, Hitoshi Murata, Ken-Ichi Yamamoto, Masami Watanabe, Hiromi Kumon, Masakiyo Sakaguchi

Journal of molecular medicine (Berlin, Germany) 101 ( 4 ) 431 - 447 2023年3月

記述言語：英語掲載種別：研究論文（学術雑誌）

The adenovirus-REIC/Dkk-3 expression vector (Ad-REIC) has been the focus of numerous clinical studies due to its potential for the quenching of cancers. The cancer-suppressing mechanisms of the REIC/DKK-3 gene depend on multiple pathways that exert both direct and indirect effects on cancers. The direct effect is triggered by REIC/Dkk-3-mediated ER stress that causes cancer-selective apoptosis, and the indirect effect can be classified in two ways: (i) induction, by Ad-REIC-mis-infected cancer-associated fibroblasts, of the production of IL-7, an important activator of T cells and NK cells, and (ii) promotion, by the secretory REIC/Dkk-3 protein, of dendritic cell polarization from monocytes. These unique features allow Ad-REIC to exert effective and selective cancer-preventative effects in the manner of an anticancer vaccine. However, the question of how the REIC/Dkk-3 protein leverages anticancer immunity has remained to be answered. We herein report a novel function of the extracellular REIC/Dkk-3-namely, regulation of an immune checkpoint via modulation of PD-L1 on the cancer-cell surface. First, we identified novel interactions of REIC/Dkk-3 with the membrane proteins C5aR, CXCR2, CXCR6, and CMTM6. These proteins all functioned to stabilize PD-L1 on the cell surface. Due to the dominant expression of CMTM6 among the proteins in cancer cells, we next focused on CMTM6 and observed that REIC/Dkk-3 competed with CMTM6 for PD-L1, thereby liberating PD-L1 from its complexation with CMTM6. The released PD-L1 immediately underwent endocytosis-mediated degradation. These results will enhance our understanding of not only the physiological nature of the extracellular REIC/Dkk-3 protein but also the Ad-REIC-mediated anticancer effects. KEY MESSAGES: • REIC/Dkk-3 protein effectively suppresses breast cancer progression through an acceleration of PD-L1 degradation. • PD-L1 stability on the cancer cell membrane is kept high by binding with mainly CMTM6. • Competitive binding of REIC/Dkk-3 protein with CMTM6 liberates PD-L1, leading to PD-L1 degradation.

DOI： 10.1007/s00109-023-02292-w

Etiology of recurrent cystitis in postmenopausal women based on vaginal microbiota and the role of Lactobacillus vaginal suppository. 査読国際誌

Takanori Sekito, Koichiro Wada, Ayano Ishii, Takehiro Iwata, Takehiro Matsubara, Shuta Tomida, Masami Watanabe, Motoo Araki, Takuya Sadahira

Frontiers in microbiology 14 1187479 - 1187479 2023年

記述言語：英語掲載種別：研究論文（学術雑誌）

BACKGROUND: The vaginal microbiota can be altered by uropathogenic bacteria associated with recurrent cystitis (RC), and the vaginal administration of Lactobacillus have suggested certain effects to prevent RC. The relationship between vaginal microbiota and the development of RC has not been elucidated. We aimed to clarify the etiology of RC from vaginal microbiota and importance of vaginal Lactobacillus. METHODS: Vaginal samples obtained from 39 postmenopausal women were classified into four groups: healthy controls; uncomplicated cystitis; RC; and prevention (prevented RC by Lactobacillus crispatus-containing vaginal suppositories). Principal coordinate analysis and beta-diversity analysis was used to assess 16S rRNA gene sequencing data from the vaginal microbiome. RESULTS: Cluster analysis divided the vaginal bacterial communities among 129 vaginal samples into three clusters (A, B, and C). Fourteen of 14 (100%) samples from the RC group and 51 of 53 (96%) samples from the prevention group were in clusters B and C, while 29 of 38 (76%) samples from the healthy group and 14 of 24 (58%) samples from the uncomplicated cystitis group were in cluster A. The principal coordinate analysis showed that plots in the uncomplicated cystitis group were similar to the healthy group, indicating a large separation between the RC group and the uncomplicated cystitis group. On beta-diversity analysis, there were significant differences between the healthy group and the uncomplicated cystitis group (p = 0.045), and between the RC group and the uncomplicated cystitis group or the healthy group (p = 0.001, p = 0.001, respectively). There were no significant differences between the RC group and the prevention group (p = 0.446). The top six taxa were as follows: Prevotella, Lactobacillus, Streptococcus, Enterobacteriaceae, Anaerococcus, and Bifidobacterium. Among patients with RC, Lactobacillus was undetectable before administration of suppositories, while the median relative abundance of Lactobacillus was 19% during administration of suppositories (p = 0.0211), reducing the average cystitis episodes per year (6.3 vs. 2.4, p = 0.0015). CONCLUSION: The vaginal microbiota of postmenopausal women with RC is differed from healthy controls and uncomplicated cystitis in terms of lack of Lactobacillus and relatively dominant of Enterobacteriaceae. Vaginal administration of Lactobacillus-containing suppositories can prevent RC by stabilizing vaginal dysbiosis and causing a loss of pathogenic bacteria virulence.

DOI： 10.3389/fmicb.2023.1187479

Canine Mammary Tumor Cell Lines Derived from Metastatic Foci Show Increased RAD51 Expression but Diminished Radioresistance via p21 Inhibition. 査読国際誌

Kei Shimakawa, Kazuhiko Ochiai, Sachi Hirose, Eri Tanabe, Masaki Michishita, Motoharu Sakaue, Yasunaga Yoshikawa, Masami Morimatsu, Tsuyoshi Tajima, Masami Watanabe, Yoshikazu Tanaka

Veterinary sciences 9 ( 12 ) 2022年12月

記述言語：英語掲載種別：研究論文（学術雑誌）

Due to the high incidence of mammary tumors in dogs, it is important to elucidate the pathogenesis of these tumors in veterinary medicine. Radiation therapy is often used to treat mammary tumors that target DNA lesions. RAD51 is a key molecule that repairs DNA damage via homologous recombination. We examined the relationship between RAD51 expression and radiosensitivity in mammary tumor cell lines. CHMp and CHMm from the same individual were selected based on the differences in RAD51 expression. The radiosensitivity of both cell lines was examined using MTT and scratch assays; CHMm, which has high RAD51 expression, showed higher sensitivity to radiation than CHMp. However, the nuclear focus of RAD51 during DNA repair was formed normally in CHMp, but not in most of CHMm. Since irradiation resulted in the suppression of cell cycle progression in CHMp, the expression of p21, a cell cycle regulatory factor, was detected in CHMp after 15 Gy irradiation but not in CHMm. These results indicate that functional expression is more important than the quantitative expression of RAD51 in canine mammary tumor cells in response to DNA damage.

DOI： 10.3390/vetsci9120703

Protocol for the generation of human induced hepatic stem cells using Sendai virus vectors. 査読国際誌

Hirofumi Noguchi, Yoshiki Nakashima, Masami Watanabe, Masayuki Matsushita, Masayoshi Tsukahara, Issei Saitoh, Chika Miyagi-Shiohira

STAR protocols 3 ( 4 ) 101884 - 101884 2022年12月

記述言語：英語掲載種別：研究論文（学術雑誌）

Our recent study demonstrated the generation of induced tissue-specific stem/progenitor (iTS/iTP) cells by the transient overexpression of reprogramming factors combined with tissue-specific selection. Here, we present a protocol to reprogram human hepatocytes to generate human induced tissue-specific liver stem (iTS-L) cells. Human hepatocytes are transfected with Sendai virus vectors (SeV) expressing OCT3/4, SOX2, KLF4, and c-MYC. iTS-L cells continuously express mRNA of hepatocyte-specific markers (HNF1β and HNF4α) and do not form teratomas. For complete details on the use and execution of this protocol, please refer to Nakashima et al. (2022).1.

DOI： 10.1016/j.xpro.2022.101884

Toll-like receptor 4 promotes bladder cancer progression upon S100A8/A9 binding, which requires TIRAP-mediated TPL2 activation 査読

Acosta Gonzalez Herik Rodrigo, Nahoko Tomonobu, Haruka Yoneda, Rie Kinoshita, Yosuke Mitsui, Takuya Sadahira, Shin-ichi Terawaki, Yuma Gohara, Ni Luh Gede Yoni Komalasari, Fan Jiang, Hitoshi Murata, Ken-ichi Yamamoto, Junichiro Futami, Akira Yamauchi, Futoshi Kuribayashi, Yusuke Inoue, Eisaku Kondo, Shinichi Toyooka, Masahiro Nishibori, Masami Watanabe, Yasutomo Nasu, Masakiyo Sakaguchi

Biochemical and Biophysical Research Communications 634 83 - 91 2022年12月

Induced hepatic stem cells are suitable for human hepatocyte production. 査読国際誌

Yoshiki Nakashima, Chika Miyagi-Shiohira, Issei Saitoh, Masami Watanabe, Masayuki Matsushita, Masayoshi Tsukahara, Hirofumi Noguchi

iScience 25 ( 10 ) 105052 - 105052 2022年10月

記述言語：英語掲載種別：研究論文（学術雑誌）

Human hepatocytes were transfected with Sendai virus vectors (SeV) expressing OCT3/4, SOX2, KLF4, and C-MYC to produce hepatocyte-derived induced pluripotent stem cells (iPSCs). The messenger RNA (mRNA) expression of undifferentiated markers (passage 19-21) and hepatocyte-specific markers (HSMs) (passage 0-20) in 48 established hepatocyte-derived iPSC-like colonies was examined. Among the 48 clones, 10 clones continuously expressed HSM mRNA (HNF1β and HNF4α) in passage 0-20. The colonies which expressed HSMs (iTS-L cells: induced tissue-specific stem cells from liver) showed a different tendency in microarray and methylation analyses to fibroblast-derived iPSCs (strain: 201B7). iTS-L cells were less likely to form teratomas in mice than iPSCs (He). The iTS-L cells were differentiated into hepatocyte-like cells more efficiently than iPSCs (He) or iPSCs (201B7). These data suggest that SeV expressing OCT3/4, SOX2, KLF4, and C-MYC induce the generation of iPSCs and iTS-L cells.

DOI： 10.1016/j.isci.2022.105052

AP39, a Mitochondrial-Targeted H2S Donor, Improves Porcine Islet Survival in Culture. 査読国際誌

Misaki Shinzato, Chika Miyagi-Shiohira, Kazuho Kuwae, Kai Nishime, Yoshihito Tamaki, Tasuku Yonaha, Mayuko Sakai-Yonaha, Ikuo Yamasaki, Ryusei Otsuka, Issei Saitoh, Masami Watanabe, Hirofumi Noguchi

Journal of clinical medicine 11 ( 18 ) 2022年9月

記述言語：英語掲載種別：研究論文（学術雑誌）

The rapid deterioration of transplanted islets in culture is a well-established phenomenon. We recently reported that pancreas preservation with AP39 reduces reactive oxygen species (ROS) production and improves islet graft function. In this study, we investigated whether the addition of AP39 to the culture medium could reduce isolated islet deterioration and improve islet function. Isolated islets from porcine pancreata were cultured with 400 nM AP39 or without AP39 at 37 °C. After culturing for 6-72 h, the islet equivalents of porcine islets in the AP39(+) group were significantly higher than those in the AP39(-) group. The islets in the AP39(+) group exhibited significantly decreased levels of ROS production compared to the islets in the AP39(-) group. The islets in the AP39(+) group exhibited significantly increased mitochondrial membrane potential compared to the islets in the AP39(-) group. A marginal number (1500 IEs) of cultured islets from each group was then transplanted into streptozotocin-induced diabetic mice. Culturing isolated islets with AP39 improved islet transplantation outcomes in streptozotocin-induced diabetic mice. The addition of AP39 in culture medium reduces islet deterioration and furthers the advancements in β-cell replacement therapy.

DOI： 10.3390/jcm11185385

Pancreas Preservation in Modified Histidine-lactobionate Solution Is Superior to That in University of Wisconsin Solution for Porcine Islet Isolation. 査読国際誌

Mayuko Sakai-Yonaha, Chika Miyagi-Shiohira, Kazuho Kuwae, Yoshihito Tamaki, Kai Nishime, Tasuku Yonaha, Issei Saitoh, Masami Watanabe, Hirofumi Noguchi

Transplantation 106 ( 9 ) 1770 - 1776 2022年9月

記述言語：英語掲載種別：研究論文（学術雑誌）

BACKGROUND: We previously reported that modified extracellular-type trehalose-containing Kyoto (MK) solution, which contains a trypsin inhibitor (ulinastatin), significantly improved the islet yield compared with University of Wisconsin (UW) preservation, which is the gold standard for organ preservation for islet isolation. In this study, we evaluated the efficiency of a modified histidine-lactobionate (MHL) solution in addition to UW or MK solution. The MHL solution has a high sodium-low potassium composition with low viscosity compared with the UW solution. Moreover, similar to MK solution, MHL solution also contains ulinastatin. METHODS: Porcine pancreata were preserved in UW, MK, or MHL solution, followed by islet isolation. An optimized number (1500 IE) of isolated islets from each group were then transplanted into streptozotocin-induced diabetic mice. RESULTS: The islet yield before and after purification was significantly higher in the MHL group than in the UW group. On the contrary, the islet yield before and after purification was not significantly different between the MHL and MK groups. Preserving the porcine pancreata in MHL solution improved the outcome of islet transplantation in streptozotocin-induced diabetic mice compared with that in UW solution. CONCLUSIONS: Pancreas preservation with MHL solution preserves islet function better than UW solution. The effect of MHL solution is similar to that of MK solution, suggesting that MHL solution can be used as an alternative to MK solution for pancreatic islet transplantation.

DOI： 10.1097/TP.0000000000003636

Pancreas Preservation with a Neutrophil Elastase Inhibitor, Alvelestat, Contributes to Improvement of Porcine Islet Isolation and Transplantation. 査読国際誌

Ryusei Otsuka, Chika Miyagi-Shiohira, Kazuho Kuwae, Kai Nishime, Yoshihito Tamaki, Tasuku Yonaha, Mayuko Sakai-Yonaha, Ikuo Yamasaki, Misaki Shinzato, Issei Saitoh, Masami Watanabe, Hirofumi Noguchi

Journal of clinical medicine 11 ( 15 ) 2022年7月

記述言語：英語掲載種別：研究論文（学術雑誌）

For pancreatic islet transplantation, pancreas procurement, preservation, and islet isolation destroy cellular and non-cellular components and activate components such as resident neutrophils, which play an important role in the impairment of islet survival. It has been reported that inhibitors of neutrophil elastase (NE), such as sivelestat and α1-antitrypsin, could contribute to improvement of islet isolation and transplantation. In this study, we investigated whether pancreatic preservation with alvelestat, a novel NE inhibitor, improves porcine islet yield and function. Porcine pancreata were preserved with or without 5 μM alvelestat for 18 h, and islet isolation was performed. The islet yields before and after purification were significantly higher in the alvelestat (+) group than in the alvelestat (-) group. After islet transplantation into streptozotocin-induced diabetic mice, blood glucose levels reached the normoglycemic range in 55% and 5% of diabetic mice in the alvelestat (+) and alvelestat (-) groups, respectively. These results suggest that pancreas preservation with alvelestat improves islet yield and graft function and could thus serve as a novel clinical strategy for improving the outcome of islet transplantation.

DOI： 10.3390/jcm11154290

Intravesical indwelling lidocaine‑releasing devices for IC/BPS (Review) 査読

Tomofumi Watanabe, Takuya Sadahira, Masami Watanabe, Yuki Maruyama, Koichiro Wada, Takanori Sekito, Kohei Edamura, Yasuyuki Kobayashi, Motoo Araki, Toyohiko Watanabe

World Academy of Sciences Journal 4 ( 4 ) 2022年6月

In vitro anticancer effects of alpelisib against PIK3CA‑mutated canine hemangiosarcoma cell lines. 査読国際誌

Marika Maeda, Kazuhiko Ochiai, Masaki Michishita, Masami Morimatsu, Hiroki Sakai, Nayuta Kinoshita, Motoharu Sakaue, Eri Onozawa, Daigo Azakami, Masami Yamamoto, Katsumi Ishioka, Takuya Sadahira, Masami Watanabe, Yoshikazu Tanaka

Oncology reports 47 ( 4 ) 2022年4月

記述言語：英語掲載種別：研究論文（学術雑誌）

Hemangiosarcoma (HSA) is a malignant neoplasm that occurs in humans and canines with a poor prognosis owing to metastatic spread, despite effective treatment. The frequency of spontaneous HSA development is higher in canines than in humans. Therefore, canine HSA is a useful model of intractable human disease, which requires early detection and an effective therapeutic strategy. A high frequency of the p110α phosphatidylinositol‑4,5‑bisphosphate 3‑kinase catalytic subunit alpha (PIK3CA) mutations is detected in a comprehensive genome‑wide analysis of canine cases of HSA. The present cloned the full‑length cDNA of canine PIK3CA and identified a mutation in codon 1047 from canine cases of HSA and cell lines that were established from these. The enforced expression of the 1047th histidine residue (H1047)R or L mutants of canine PIK3CA in HeLa cells enhanced epidermal growth factor receptor (EGFR) signaling via Akt phosphorylation. PIK3CA mutant canine HSA cell lines exhibited the hyperphosphorylation of Akt upon EGF stimulation as well. Alpelisib, a molecular targeted drug against PIK3CA activating mutations, exerted a significant antitumor effect in canine PIK3CA‑mutated HSA cell lines. By contrast, it had no significant effect on canine mammary gland tumor cell lines harboring PIK3CA mutations. On the whole, the findings of the present study suggest that alpelisib may be highly effective against PIK3CA mutations that occur frequently in canine HSA.

DOI： 10.3892/or.2022.8295

Dkk3/REIC Deficiency Impairs Spermiation, Sperm Fibrous Sheath Integrity and the Sperm Motility of Mice. 査読国際誌

Ruizhi Xue, Wenfeng Lin, Hirofumi Fujita, Jingkai Sun, Rie Kinoshita, Kazuhiko Ochiai, Junichiro Futami, Masami Watanabe, Hideyo Ohuchi, Masakiyo Sakaguchi, Zhengyan Tang, Peng Huang, Yasutomo Nasu, Hiromi Kumon

Genes 13 ( 2 ) 2022年1月

記述言語：英語掲載種別：研究論文（学術雑誌）

The role of Dickkopf-3 (Dkk3)/REIC (The Reduced Expression in Immortalized Cells), a Wnt-signaling inhibitor, in male reproductive physiology remains unknown thus far. To explore the functional details of Dkk3/REIC in the male reproductive process, we studied the Dkk3/REIC knock-out (KO) mouse model. By examining testicular sections and investigating the sperm characteristics (count, vitality and motility) and ultrastructure, we compared the reproductive features between Dkk3/REIC-KO and wild-type (WT) male mice. To further explore the underlying molecular mechanism, we performed RNA sequencing (RNA-seq) analysis of testicular tissues. Our results showed that spermiation failure existed in seminiferous tubules of Dkk3/REIC-KO mice, and sperm from Dkk3/REIC-KO mice exhibited inferior motility (44.09 ± 8.12% vs. 23.26 ± 10.02%, p < 0.01). The Ultrastructure examination revealed defects in the sperm fibrous sheath of KO mice. Although the average count of Dkk3/REIC-KO epididymal sperm was less than that of the wild-types (9.30 ± 0.69 vs. 8.27 ± 0.87, ×106), neither the gap (p > 0.05) nor the difference in the sperm vitality rate (72.83 ± 1.55% vs. 72.50 ± 0.71%, p > 0.05) were statistically significant. The RNA-seq and GO (Gene Oncology) enrichment results indicated that the differential genes were significantly enriched in the GO terms of cytoskeleton function, cAMP signaling and calcium ion binding. Collectively, our research demonstrates that Dkk3/REIC is involved in the process of spermiation, fibrous sheath integrity maintenance and sperm motility of mice.

DOI： 10.3390/genes13020285

Medical uses for silver nitrate in the urinary tract (Review) 査読

Takanori Sekito, Takuya Sadahira, Toyohiko Watanabe, Yuki Maruyama, Tomofumi Watanabe, Takehiro Iwata, Koichiro Wada, Kohei Edamura, Motoo Araki, Masami Watanabe

World Academy of Sciences Journal 4 ( 1 ) 2022年1月

p Identification of MICALL2 as a Novel Prognostic Biomarker Correlating with Inflammation and T Cell Exhaustion of Kidney Renal Clear Cell Carcinoma 査読

Wenfeng Lin, Wenwei Chen, Jisheng Zhong, Hideo Ueki, Abai Xu, Masami Watanabe, Motoo Araki, Chunxiao Liu, Yasutomo Nasu, Peng Huang

JOURNAL OF CANCER 13 ( 3 ) 1214 - 1228 2022年

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：IVYSPRING INT PUBL

Purpose: The interplay of inflammation and immunity affects all stages from tumorigenesis to progression, and even tumor response to therapy. A growing interest has been attracted from the biological function of MICALL2 to its effects on tumor progression. This study was designed to verify whether MICALL2 could be a prognostic biomarker to predict kidney renal clear cell carcinoma (KIRC) progression, inflammation, and immune infiltration within tumor microenvironment (TME). Methods: We firstly analyzed MICALL2 expressions across 33 cancer types from the UCSC Xena database and verified its expression in KIRC through GEPIA platform and GEO datasets. The clinicopathological characteristics were further analyzed based on the median expression. Kaplan-Meier method, univariate and multivariate analyses were applied to compare survival outcomes. ESTIMATE and CIBERSORT algorithms were performed to assess immune infiltration, and a co-expression analysis was conducted to evaluate the correlation between MICALL2 and immunoregulatory genes. Enrichment analysis was finally performed to explore the biological significance of MICALL2. Results: MICALL2 was highly expressed in 16 types of cancers compared with normal tissues. MICALL2 expression increased with advanced clinicopathological parameters and was an independent predictor for poor prognosis in KIRC. Moreover, MICALL2 closely correlated with inflammation-promoting signatures and immune infiltration including T cell exhaustion markers. Consistently, MICALL2 involved in the regulation of signaling pathways associated with tumor immunity, tumor progression, and impaired metabolic activities. Conclusion: MICALL2 can function as a prognostic biomarker mediating inflammation, immune infiltration, and T cell exhaustion within the microenvironment of KIRC.

DOI： 10.7150/jca.66922

Testosterone Recovery after Neoadjuvant Gonadotropin-Releasing Hormone Antagonist versus Agonist on Permanent Iodine-125 Seed Brachytherapy in Prostate Cancer Patients: A Propensity Score Analysis.

Takehiro Iwata, Yuki Maruyama, Tatsushi Kawada, Takuya Sadahira, Satoshi Katayama, Atsushi Takamoto, Tomoko Sako, Koichiro Wada, Kohei Edamura, Yasuyuki Kobayashi, Motoo Araki, Masami Watanabe, Toyohiko Watanabe, Yasutomo Nasu

Acta medica Okayama 75 ( 6 ) 705 - 711 2021年12月

記述言語：英語掲載種別：研究論文（学術雑誌）

Optimal neoadjuvant hormone therapy (NHT) for reducing prostate cancer (PC) patients' prostate volume pre-brachytherapy is controversial. We evaluated the differential impact of neoadjuvant gonadotropin-releasing hormone (GnRH) antagonist versus agonist on post-brachytherapy testosterone recovery in 112 patients treated pre-brachytherapy with NHT (GnRH antagonist, n=32; GnRH agonists, n=80) (Jan. 2007-June 2019). We assessed the effects of patient characteristics and a GnRH analogue on testosterone recovery with logistic regression and a propensity score analysis (PSA). There was no significant difference in the rate of testosterone recovery to normal levels (> 300 ng/dL) between the GnRH antagonist and agonists (p=0.07). The GnRH agonists induced a significantly more rapid testosterone recovery rate at 3 months post-brachytherapy versus the GnRH antagonist (p<0.0001); there was no difference in testosterone recovery at 12 months between the GnRH antagonist/agonists (p=0.8). In the multivariate analysis, no actor was associated with testosterone recovery. In the PSA, older age and higher body mass index (BMI) were significantly associated with longer testosterone recovery. Post-brachytherapy testosterone recovery was quicker with the neoadjuvant GnRH agonists than the antagonist, and the testosterone recovery rate was significantly associated with older age and higher BMI. Long-term follow-ups are needed to determine any differential effects of GnRH analogues on the quality of life of brachytherapy-treated PC patients.

DOI： 10.18926/AMO/62810

Clinical Efficacy and Safety of Sitafloxacin 200 mg Once Daily for Refractory Genitourinary Tract Infections. 査読

Takehiro Iwata, Takuya Sadahira, Yuki Maruyama, Takanori Sekito, Kasumi Yoshinaga, Shogo Watari, Kentaro Nagao, Tatsushi Kawada, Yusuke Tominaga, Shingo Nishimura, Atsushi Takamoto, Tomoko Sako, Kohei Edamura, Yasuyuki Kobayashi, Motoo Araki, Ayano Ishii, Masami Watanabe, Toyohiko Watanabe, Yasutomo Nasu, Koichiro Wada

Acta medica Okayama 75 ( 6 ) 763 - 766 2021年12月

記述言語：英語掲載種別：研究論文（学術雑誌）

The aim of this ongoing trial is to evaluate the clinical efficacy and safety of sitafloxacin (STFX) 200 mg once daily (QD) for 7 days in patients with refractory genitourinary tract infections, which include recurrent or complicated cystitis, complicated pyelonephritis, bacterial prostatitis, and epididymitis. The primary endpoint is the microbiological efficacy at 5-9 days after the last administration of STFX. Recruitment began in February 2021, and the target total sample size is 92 participants.

DOI： 10.18926/AMO/62820

In vivo evaluation of GG2-GG1/A2 element activity in the insulin promoter region using the CRISPR-Cas9 system. 国際誌

Hirofumi Noguchi, Chika Miyagi-Shiohira, Takao Kinjo, Issei Saitoh, Masami Watanabe

Scientific reports 11 ( 1 ) 20290 - 20290 2021年10月

Efficacy of Lactobacillus vaginal suppositories for the prevention of recurrent cystitis: A phase II clinical trial. 国際誌

Takuya Sadahira, Koichiro Wada, Motoo Araki, Ritsuko Mitsuhata, Masumi Yamamoto, Yuki Maruyama, Takehiro Iwata, Masami Watanabe, Toyohiko Watanabe, Reiko Kariyama, Yasutomo Nasu, Ayano Ishii

International journal of urology : official journal of the Japanese Urological Association 28 ( 10 ) 1026 - 1031 2021年10月

Impact of Sarcopenia on Erectile Function after Nerve-Sparing Robot-Assisted Radical Prostatectomy. 国際誌

Yosuke Mitsui, Takuya Sadahira, Yuki Maruyama, Ryota Sato, Acosta Gonzalez, Herik Rodrigo, Koichiro Wada, Motoo Araki, Masami Watanabe, Toyohiko Watanabe, Yasutomo Nasu

The world journal of men's health 39 ( 4 ) 673 - 682 2021年10月

Blood concentrations of tacrolimus upon conversion from rabeprazole to vonoprazan in renal transplant recipients: Correlation with cytochrome P450 gene polymorphisms. 国際誌

Shogo Watari, Motoo Araki, Jun Matsumoto, Kasumi Yoshinaga, Takanori Sekito, Yuki Maruyama, Yosuke Mitsui, Takuya Sadahira, Risa Kubota, Shingo Nishimura, Koichiro Wada, Yasuyuki Kobayashi, Hidemi Takeuchi, Katsuyuki Tanabe, Masashi Kitagawa, Hiroshi Morinaga, Shinji Kitamura, Hitoshi Sugiyama, Noritaka Ariyoshi, Jun Wada, Masami Watanabe, Toyohiko Watanabe, Yasutomo Nasu

Drug metabolism and pharmacokinetics 40 100407 - 100407 2021年10月

Presence of decoy cells for 6 months on urine cytology efficiently predicts BK virus nephropathy in renal transplant recipients. 国際誌

Takanori Sekito, Motoo Araki, Kasumi Yoshinaga, Yuki Maruyama, Takuya Sadahira, Shingo Nishimura, Koichiro Wada, Masami Watanabe, Toyohiko Watanabe, Katsuyuki Tanabe, Hidemi Takeuchi, Hiroshi Morinaga, Masashi Kitagawa, Shinji Kitamura, Hitoshi Sugiyama, Jun Wada, Hiroyuki Yanai, Yasutomo Nasu

International journal of urology : official journal of the Japanese Urological Association 28 ( 12 ) 1240 - 1246 2021年9月

The Cell Cycle Checkpoint Gene, RAD17 rs1045051, Is Associated with Prostate Cancer Risk. 査読国際共著

Jingkai Sun, Wenfeng Lin, Qixu Wang, Akiko Sakai, Ruizhi Xue, Masami Watanabe, Chunxiao Liu, Takuya Sadahira, Yasutomo Nasu, Abai Xu, Peng Huang

Acta medica Okayama 75 ( 4 ) 415 - 421 2021年8月

Successful deceased donor kidney transplantation to a recipient with a history of COVID-19 treatment. 国際誌

Kasumi Yoshinaga, Motoo Araki, Koichiro Wada, Kou Hasegawa, Takanori Sekito, Shuji Miyake, Shogo Watari, Yuki Maruyama, Takuya Sadahira, Shingo Nishimura, Katsuyuki Tanabe, Hidemi Takeuchi, Yuri Nakashima, Masaru Kinomura, Herik Acosta, Yosuke Mitsui, Risa Kubota, Hirochika Nakajima, Kohei Edamura, Yasuyuki Kobayashi, Masami Watanabe, Toyohiko Watanabe, Fumio Otsuka, Jun Wada, Yasutomo Nasu

Journal of infection and chemotherapy : official journal of the Japan Society of Chemotherapy 27 ( 7 ) 1097 - 1101 2021年7月

Cytopathic effects and local immune responses in repeated neoadjuvant HSV-tk + ganciclovir gene therapy for prostate cancer. 国際誌

Nobuyuki Yanagisawa, Takefumi Satoh, Ken-Ichi Tabata, Hideyasu Tsumura, Yasutomo Nasu, Masami Watanabe, Timothy C Thompson, Isao Okayasu, Yoshiki Murakumo, Shiro Baba, Masatsugu Iwamura

Asian journal of urology 8 ( 3 ) 280 - 288 2021年7月

Feasible kidney donation with living marginal donors, including diabetes mellitus. 国際誌

Kasumi Yoshinaga, Motoo Araki, Koichiro Wada, Takanori Sekito, Shogo Watari, Yuki Maruyama, Yosuke Mitsui, Takuya Sadahira, Risa Kubota, Shingo Nishimura, Kohei Edamura, Yasuyuki Kobayashi, Katsuyuki Tanabe, Hidemi Takeuchi, Masashi Kitagawa, Shinji Kitamura, Jun Wada, Masami Watanabe, Toyohiko Watanabe, Yasutomo Nasu

Immunity, inflammation and disease 9 ( 3 ) 1061 - 1068 2021年6月

Comparison of intracorporeal versus extracorporeal urinary diversion after robot-assisted radical cystectomy at a medium-sized facility.

Takehiro Iwata, Yasuyuki Kobayashi, Yuki Maruyama, Tatsushi Kawada, Takuya Sadahira, Yuko Oiwa, Satoshi Katayama, Shingo Nishimura, Atsushi Takamoto, Tomoko Sako, Koichiro Wada, Kohei Edamura, Motoo Araki, Masami Watanabe, Toyohiko Watanabe, Yasutomo Nasu

International journal of clinical oncology 26 ( 9 ) 1714 - 1721 2021年6月

The number of glutamines in the N-terminal of the canine androgen receptor affects signalling intensities. 国際誌

Kazuhiko Ochiai, Samak Sutijarit, Mitsuki Uemura, Masami Morimatsu, Masaki Michishita, Eri Onozawa, Marika Maeda, Takanori Sasaki, Masami Watanabe, Yoshikazu Tanaka, Toshinori Omi

Veterinary and comparative oncology 19 ( 2 ) 399 - 403 2021年6月

A Case of Metastatic Fumarate Hydratase-Deficient-like Renal Cell Carcinoma Successfully Managed by Ipilimumab plus Nivolumab.

Takanori Sekito, Atsushi Takamoto, Yasuyuki Kobayashi, Masao Mitsui, Shogo Watari, Risa Kubota, Takuya Sadahira, Takehiro Iwata, Shingo Nishimura, Kohei Edamura, Tomoko Sako, Motoo Araki, Masami Watanabe, Toyohiko Watanabe, Rei Shibata, Daisuke Ennishi, Yasutomo Nasu

Acta medica Okayama 75 ( 3 ) 397 - 402 2021年6月

ABO Blood Incompatibility Positively Affects Early Graft Function: Single-Center Retrospective Cohort Study. 国際誌

Shogo Watari, Motoo Araki, Koichiro Wada, Kasumi Yoshinaga, Yuki Maruyama, Yosuke Mitsui, Takuya Sadahira, Risa Kubota, Shingo Nishimura, Yasuyuki Kobayashi, Hidemi Takeuchi, Katsuyuki Tanabe, Masashi Kitagawa, Hiroshi Morinaga, Shinji Kitamura, Hitoshi Sugiyama, Jun Wada, Masami Watanabe, Toyohiko Watanabe, Yasutomo Nasu

Transplantation proceedings 53 ( 5 ) 1494 - 1500 2021年6月

Evaluation of Neutrophil Dynamics Change by Protective Effect of Tadalafil After Renal Ischemia/Reperfusion Using In Vivo Real-time Imaging. 国際誌

Yuki Maruyama, Motoo Araki, Kengo Kidokoro, Yuji Sogawa, Kasumi Yoshinaga, Yosuke Mitsui, Takuya Sadahira, Koichiro Wada, Masami Watanabe, Toyohiko Watanabe, Naoki Kashihara, Yasutomo Nasu

Transplantation 106 ( 2 ) 280 - 288 2021年4月

Pancreas preservation with amphotericin B deteriorates islet yield for porcine islet isolation. 国際誌

Sayaka Sawada, Chika Miyagi-Shiohira, Kazuho Kuwae, Yoshihito Tamaki, Kai Nishime, Mayuko Sakai-Yonaha, Tasuku Yonaha, Issei Saitoh, Masami Watanabe, Hirofumi Noguchi

Xenotransplantation 28 ( 4 ) e12690 2021年4月

A second opinion pathology review improves the diagnostic concordance between prostate cancer biopsy and radical prostatectomy specimens

Takanori Maehara, Takuya Sadahira, Yuki Maruyama, Koichiro Wada, Motoo Araki, Masami Watanabe, Toyohiko Watanabe, Hiroyuki Yanai, Yasutomo Nasu

Urology Annals 13 ( 2 ) 119 - 124 2021年4月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：Wolters Kluwer Medknow Publications

Objectives: The Gleason scoring system is an essential tool for determining the treatment strategy in prostate cancer (PCa). However, the Gleason grade group (GGG) often differs between needle-core biopsy (NCB) and radical prostatectomy (RP) specimens. We investigated the diagnostic value of a second opinion pathology review using NCB specimens in PCa. Materials and Methods: We retrospectively evaluated 882 patients who underwent robot-assisted RP from January 2012 to September 2019. Of these, patients whose original biopsy specimens were obtained from another hospital and reviewed by the urological pathology expert at our institution were included in the study. Patients who received neoadjuvant hormonal therapy were excluded from the study. Weighted kappa (k) coefficients were used to evaluate the diagnostic accuracy of each review. Results: A total of 497 patients were included in this study. Substantial agreement (weighted k = 0.783) in the GGG between initial- and second-opinion diagnoses based on NCB specimens was observed in 310 cases (62.4%). Although diagnoses based on a single opinion showed moderate agreement with the GGG of RP specimens (initial: 35.2%, weighted k = 0.522
second opinion
38.8%, weighted k = 0.560), matching initial and second opinion diagnoses improved the concordance (42.9%, 133/310 cases) to substantial agreement (weighted k = 0.626). Conclusions: A second opinion of PCa pathology helps to improve the diagnostic accuracy of NCB specimens. However, over half of diagnoses that matched between the initial and second opinions differed from the diagnosis of RP specimens.

DOI： 10.4103/UA.UA_81_20

Impact of paclitaxel, cisplatin, and gemcitabine as first-line chemotherapy in cisplatin-fit and -unfit patients with advanced/metastatic urothelial carcinoma. 国際誌

Satoshi Katayama, Yasuyuki Kobayashi, Atsushi Takamoto, Kohei Edamura, Takuya Sadahira, Takehiro Iwata, Shingo Nishimura, Tomoko Sako, Koichiro Wada, Motoo Araki, Masami Watanabe, Toyohiko Watanabe, Yasutomo Nasu

Urologic oncology 39 ( 10 ) 731.e25-731.e32 2021年3月

Photodynamic diagnostic ureteroscopy using the VISERA ELITE video system for diagnosis of upper-urinary tract urothelial carcinoma: a prospective cohort pilot study. 国際誌

Koichiro Wada, Motoo Araki, Ryuta Tanimoto, Takuya Sadahira, Shogo Watari, Yuki Maruyama, Yosuke Mitsui, Hirochika Nakajima, Herik Acosta, Satoshi Katayama, Takehiro Iwata, Shingo Nishimura, Atsushi Takamoto, Tomoko Sako, Kohei Edamura, Yasuyuki Kobayashi, Masami Watanabe, Toyohiko Watanabe, Yasutomo Nasu

BMC urology 21 ( 1 ) 45 - 45 2021年3月

Pancreas preservation in extracellular-type p38 inhibitor-containing solution improves islet yield for porcine islet isolation. 国際誌

Tasuku Yonaha, Chika Miyagi-Shiohira, Kazuho Kuwae, Yoshihito Tamaki, Kai Nishime, Mayuko Sakai-Yonaha, Issei Saitoh, Masami Watanabe, Hirofumi Noguchi

Xenotransplantation 28 ( 2 ) e12661 2021年3月

Gene Expression in Pancreatic Cancer-Like Cells and Induced Pancreatic Stem Cells Generated by Transient Overexpression of Reprogramming Factors. 国際誌

Chika Miyagi-Shiohira, Issei Saitoh, Masami Watanabe, Hirofumi Noguchi

Journal of clinical medicine 10 ( 3 ) 2021年1月

The role of Wnt signaling in male reproductive physiology and pathology. 国際共著国際誌

Ruizhi Xue, Wenfeng Lin, Jingkai Sun, Masami Watanabe, Abai Xu, Motoo Araki, Yasutomo Nasu, Zhengyan Tang, Peng Huang

Molecular human reproduction 27 ( 1 ) 2021年1月

The role of Wnt signaling in male reproductive physiology and pathology. 国際誌

Ruizhi Xue, Wenfeng Lin, Jingkai Sun, Masami Watanabe, Abai Xu, Motto Akira, Yasutomo Nasu, Zhengyan Tang, Peng Huang

Molecular human reproduction 2021年1月

Long-term ureteroscopic management of upper tract urothelial carcinoma: 28-year single-centre experience. 国際誌

Yuki Maruyama, Motoo Araki, Koichiro Wada, Kasumi Yoshinaga, Yosuke Mitsui, Takuya Sadahira, Shingo Nishimura, Kohei Edamura, Yasuyuki Kobayashi, Masami Watanabe, Toyohiko Watanabe, Manoj Monga, Yasutomo Nasu, Hiromi Kumon

Japanese journal of clinical oncology 51 ( 1 ) 130 - 137 2021年1月

Dual-Functional PLGA Nanoparticles Co-Loaded with Indocyanine Green and Resiquimod for Prostate Cancer Treatment. 国際共著国際誌

Wenfeng Lin, Chaoming Li, Naijin Xu, Masami Watanabe, Ruizhi Xue, Abai Xu, Motoo Araki, Ruifen Sun, Chunxiao Liu, Yasutomo Nasu, Peng Huang

International journal of nanomedicine 16 2775 - 2787 2021年

Discovery and Validation of Nitroxoline as a Novel STAT3 Inhibitor in Drug-resistant Urothelial Bladder Cancer. 国際共著国際誌

Wenfeng Lin, Jingkai Sun, Takuya Sadahira, Naijin Xu, Koichiro Wada, Chunxiao Liu, Motoo Araki, Abai Xu, Masami Watanabe, Yasutomo Nasu, Peng Huang

International journal of biological sciences 17 ( 12 ) 3255 - 3267 2021年

A Clinical Trial Evaluating the Usefulness of Tailored Antimicrobial Prophylaxis Using Rectal-culture Screening Media Prior to Transrectal Prostate Biopsy: A Multicenter, Randomized Controlled Trial.

Takuya Sadahira, Yuki Maruyama, Yoshiki Hiyama, Hiroyuki Kitano, Hiroki Yamada, Takayuki Goto, Tsubasa Kondo, Katsumi Shigemura, Yosuke Mitsui, Takehiro Iwata, Kohei Edamura, Motoo Araki, Masami Watanabe, Tadasu Takenaka, Jun Teishima, Yasuyoshi Miyata, Kiyohito Ishikawa, Ei-Ichiro Takaoka, Jun Miyazaki, Satoshi Takahashi, Naoya Masumori, Hiroshi Kiyota, Masato Fujisawa, Shingo Yamamoto, Takafumi Sakuma, Norihiro Kusumi, Takaharu Ichikawa, Toyohiko Watanabe, Yoshitsugu Nasu, Masaya Tsugawa, Yasutomo Nasu, Koichiro Wada

Acta medica Okayama 75 ( 5 ) 663 - 667 2021年

Medical uses for phenol in the urinary tract: A possible forgotten treatment (Review). 国際誌

Takuya Sadahira, Yuki Maruyama, Toyohiko Watanabe, Takanori Sekito, Yosuke Mitsui, Koichiro Wada, Motoo Araki, Masami Watanabe

Medicine international 1 ( 4 ) 13 - 13 2021年

記述言語：英語掲載種別：研究論文（学術雑誌）

Phenol is a chemical compound that was first used medically as an antiseptic. At low concentrations, phenol exerts local anesthetic effects achieved through denervation; at high concentrations, it exerts a potent protein-denaturing effect that induces apoptosis. Phenol injection therapy has a long history of use in urology. It is reportedly effective for hemorrhagic cystitis, benign prostate hyperplasia, overactive bladder, hydrocele, bladder tumors, interstitial cystitis and other benign urologic diseases, and it is also used as a tool to decrease bleeding during prostate surgery. The present review article summarizes the medical applications of phenol in urological field. The articles available on the medical uses of phenol are primarily older and retrospective, involving small numbers of patients. In the absence of comparative studies with other treatments, it is impossible to determine the relative benefit of phenol. However, the treatment outcomes of phenol injection are fairly well-established. Phenol therapy may be an option for patients who are poor candidates for invasive treatment. Further studies are required, however, as are improvements in the injection technique to reduce the rate of complications.

DOI： 10.3892/mi.2021.13

Novel canine isocitrate dehydrogenase 1 mutation Y208C attenuates dimerization ability. 国際誌

Shota Kawakami, Masaki Michishita, Motoharu Sakaue, Masami Morimatsu, Mitsuki Uemura, Nobuaki Kashiwagi, Marika Maeda, Yukino Machida, Daigo Azakami, Ai S Egusa, Eri Onozawa, Katsumi Ishioka, Masami Watanabe, Yoshikazu Tanaka, Toshinori Omi, Kazuhiko Ochiai

Oncology letters 20 ( 6 ) 351 - 351 2020年12月

Low-dose rituximab induction therapy is effective in immunological high-risk renal transplantation without increasing cytomegalovirus infection. 国際誌

Kasumi Yoshinaga, Motoo Araki, Koichiro Wada, Yuki Maruyama, Yosuke Mitsui, Takuya Sadahira, Risa Kubota, Shingo Nishimura, Yasuyuki Kobayashi, Hidemi Takeuchi, Katsuyuki Tanabe, Masashi Kitagawa, Hiroshi Morinaga, Haruhito Adam Uchida, Shinji Kitamura, Hitoshi Sugiyama, Jun Wada, Masami Watanabe, Toyohiko Watanabe, Yasutomo Nasu

International journal of urology : official journal of the Japanese Urological Association 27 ( 12 ) 1136 - 1142 2020年12月

Preservation of pancreas in the University of Wisconsin solution supplemented with AP39 reduces reactive oxygen species production and improves islet graft function. 国際誌

Kai Nishime, Chika Miyagi-Shiohira, Kazuho Kuwae, Yoshihito Tamaki, Tasuku Yonaha, Mayuko Sakai-Yonaha, Issei Saitoh, Masami Watanabe, Hirofumi Noguchi

American journal of transplantation : official journal of the American Society of Transplantation and the American Society of Transplant Surgeons 21 ( 8 ) 2698 - 2708 2020年11月

Kyoto probe-1 reveals phenotypic differences between mouse ES cells and iTS-P cells. 国際誌

Chika Miyagi-Shiohira, Issei Saitoh, Masami Watanabe, Hirofumi Noguchi

Scientific reports 10 ( 1 ) 18084 - 18084 2020年10月

Dynamin 1 is important for microtubule organization and stabilization in glomerular podocytes. 国際誌

The Mon La, Hiromi Tachibana, Shun-Ai Li, Tadashi Abe, Sayaka Seiriki, Hikaru Nagaoka, Eizo Takashima, Tetsuya Takeda, Daisuke Ogawa, Shin-Ichi Makino, Katsuhiko Asanuma, Masami Watanabe, Xuefei Tian, Shuta Ishibe, Ayuko Sakane, Takuya Sasaki, Jun Wada, Kohji Takei, Hiroshi Yamada

FASEB journal : official publication of the Federation of American Societies for Experimental Biology 34 ( 12 ) 16449 - 16463 2020年10月

Internalization of AMPA-type Glutamate Receptor in the MIN6 Pancreatic β-cell Line. 査読

The Mon La, Hiroshi Yamada, Sayaka Seiriki, Shun-Ai Li, Kenshiro Fujise, Natsuho Katsumi, Tadashi Abe, Masami Watanabe, Kohji Takei

Cell structure and function 45 ( 2 ) 121 - 130 2020年8月

Mutations in the C1 element of the insulin promoter lead to diabetic phenotypes in homozygous mice. 国際誌

Hirofumi Noguchi, Chika Miyagi-Shiohira, Yoshiki Nakashima, Takao Kinjo, Issei Saitoh, Masami Watanabe

Communications biology 3 ( 1 ) 309 - 309 2020年6月

Dkk3/REIC, an N-glycosylated Protein, Is a Physiological Endoplasmic Reticulum Stress Inducer in the Mouse Adrenal Gland.

Hirofumi Fujita, Tetsuya Bando, Seiichi Oyadomari, Kazuhiko Ochiai, Masami Watanabe, Hiromi Kumon, Hideyo Ohuchi

Acta medica Okayama 74 ( 3 ) 199 - 208 2020年6月

Tumor suppressor REIC/Dkk‑3 and its interacting protein SGTA inhibit glucocorticoid receptor to nuclear transport 査読

Takehiro Iwata, Takuya Sadahira, Kazuhiko Ochiai, Hideo Ueki, Takanori Sasaki, Peng Haung, Motoo Araki, Toyohiko Watanabe, Yasutomo Nasu, Masami Watanabe

Experimental and Therapeutic Medicine 2020年5月

Correlation between lumbar skeletal muscle size and urinary incontinence after radical prostatectomy. 査読国際誌

Yosuke Mitsui, Takuya Sadahira, Toyohiko Watanabe, Motoo Araki, Yuki Maruyama, Ryota Sato, Acosta Gonzalez, Herik Rodrigo, Koichiro Wada, Masami Watanabe, Michael B Chancellor, Yasutomo Nasu

Lower urinary tract symptoms 12 ( 3 ) 245 - 252 2020年5月

Novel cell-permeable p38-MAPK inhibitor efficiently prevents porcine islet apoptosis and improves islet graft function. 国際誌

Hirofumi Noguchi, Chika Miyagi-Shiohira, Yoshiki Nakashima, Issei Saitoh, Masami Watanabe

American journal of transplantation : official journal of the American Society of Transplantation and the American Society of Transplant Surgeons 20 ( 5 ) 1296 - 1308 2020年5月

Factors predicting pathological upgrading after prostatectomy in patients with Gleason grade group 1 prostate cancer based on opinion-matched biopsy specimens. 査読国際誌

Yuki Maruyama, Takuya Sadahira, Motoo Araki, Yosuke Mitsui, Koichiro Wada, Acosta Gonzalez, Herik Rodrigo, Kazuaki Munetomo, Yasuyuki Kobayashi, Masami Watanabe, Hiroyuki Yanai, Toyohiko Watanabe, Yasutomo Nasu

Molecular and clinical oncology 12 ( 4 ) 384 - 389 2020年4月

Quality of Life and Mental Satisfaction Improve Slowly in Preemptive Kidney Transplantation Compared With Nonpreemptive Kidney Transplantation. 査読国際誌

Yosuke Mitsui, Motoo Araki, Yuki Maruyama, Kasumi Yoshinaga, Takuya Sadahira, Koichiro Wada, Katsuyuki Tanabe, Masashi Kitagawa, Yasuyuki Kobayashi, Masami Watanabe, Toyohiko Watanabe, Yasutomo Nasu

Transplantation proceedings 52 ( 3 ) 740 - 747 2020年3月

Robotic Renal Autotransplantation: A Feasibility Study in a Porcine Model. 査読

Risa Kubota, Motoo Araki, Koichiro Wada, Kasumi Kawamura, Yuki Maruyama, Yosuke Mitsui, Takuya Sadahira, Yuichi Ariyoshi, Takehiro Iwata, Shingo Nishimura, Atsushi Takamoto, Tomoko Sako, Kohei Edamura, Yasuyuki Kobayashi, Yuzuki Kano, Masashi Kitagawa, Katsuyuki Tanabe, Hitoshi Sugiyama, Jun Wada, Masami Watanabe, Toyohiko Watanabe, Yasutomo Nasu

Acta medica Okayama 74 ( 1 ) 53 - 58 2020年2月

Promising Gene Therapy Using an Adenovirus Vector Carrying REIC/Dkk-3 Gene for the Treatment of Biliary Cancer. 査読国際誌

Emi Tanaka, Daisuke Uchida, Hidenori Shiraha, Hironari Kato, Atsushi Ohyama, Masaya Iwamuro, Masami Watanabe, Hiromi Kumon, Hiroyuki Okada

Current gene therapy 20 ( 1 ) 64 - 70 2020年

Nitroxoline inhibits bladder cancer progression by reversing EMT process and enhancing anti-tumor immunity. 国際共著国際誌

Naijin Xu, Wenfeng Lin, Jingkai Sun, Takuya Sadahira, Abai Xu, Masami Watanabe, Kai Guo, Motoo Araki, Gonghui Li, Chunxiao Liu, Yasutomo Nasu, Peng Huang

Journal of Cancer 11 ( 22 ) 6633 - 6641 2020年

Preventive efficacy and safety of lactobacillus vaginal suppositories in women with recurrent cystitis: A phase 2 study 査読

Sadahira T, Wada K, Ishii A, Maruyama Y, Iwata T, Araki M, Watanabe M, Watanabe T, Nasu Y

Eur Urol Open Sci 19 ( 2 ) 2036 - 2037 2020年

Clinical pharmacokinetics of oral azithromycin in epididymal tissue. 査読国際誌

Sadahira T, Wada K, Ikawa K, Morikawa N, Mitsui M, Araki M, Fujiyoshi M, Ishii A, Watanabe M, Watanabe T, Nasu Y

Journal of infection and chemotherapy : official journal of the Japan Society of Chemotherapy 25 ( 10 ) 832 - 834 2019年10月

Excellent Islet Yields after 18-h Porcine Pancreas Preservation by Ductal Injection, Pancreas Preservation with MK Solution, Bottle Purification, and Islet Purification Using Iodixanol with UW Solution and Iodixanol with MK Solution. 査読国際誌

Kuwae K, Miyagi-Shiohira C, Hamada E, Tamaki Y, Nishime K, Sakai M, Yonaha T, Makishi E, Saitoh I, Watanabe M, Noguchi H

Journal of clinical medicine 8 ( 10 ) 2019年9月

The canine RAD51 mutation leads to the attenuation of interaction with PALB2. 査読国際誌

Uemura M, Ochiai K, Morimatsu M, Michishita M, Onozawa E, Azakami D, Uno Y, Yoshikawa Y, Sasaki T, Watanabe M, Omi T

Veterinary and comparative oncology 18 ( 2 ) 247 - 255 2019年9月

Metastatic ureteral tumor arising from gastrointestinal cancer: A report of two cases

Kasumi Kawamura, Motoo Araki, Takuya Sadahira, Yuki Maruyama, Yosuke Mitsui, Shingo Nishimura, Atsushi Takamoto, Koichiro Wada, Ryuta Tanimoto, Morito Sugimoto, Yasuyuki Kobayashi, Masami Watanabe, Toyohiko Watanabe, Yasutomo Nasu, Hiroyuki Yanai

Nishinihon Journal of Urology 81 ( 4 ) 461 - 466 2019年8月

記述言語：日本語掲載種別：研究論文（学術雑誌）出版者・発行元：The West Japan Urological Association

We report two cases where gastrointestinal cancer gave rise to a metastatic ureteral tumor. Case one was a 64-year-old man who was operated on for descending colon cancer in September, 2007. In 2014, he was referred to our hospital because computed tomography (CT) scan showed a tumor of the right ureter. Ureteroscopic biopsy showed a metastatic ureteral tumor arising from descending colon cancer. The patient underwent open tumor resection. Transurethral resection of a bladder tumor (TUR- BT) was required three times for treatment of tumor recurrence from the ureteral stump in an effort to control gross hematuria Case two was a 62-year-old man who underwent pylorus-preserving gastrectomy for gastric cancer in December, 2014. Follow-up CT showed right hydronephrosis and thickening of the right ureteral wall in 2017. Selective urine cytology of the right ureter revealed urothelial carcinoma, class V. Ureteroscopic biopsy of the right ureter revealed invasive urothelial carcinoma. The patient underwent retroperitoneal laparoscopic nephroureterectomy. Histological examination revealed the right ureteral tumor to be metastasis of gastric cancer.

Robotic renal auto transplantation

Motoo Araki, Koichiro Wada, Shingo Nishimura, Risa Kubota, Kasumi Kawamura, Yuki Maruyama, Yosuke Mitsui, Takuya Sadahira, Atsushi Takamoto, Tomoko Sako, Kohei Edamura, Yasuyuki Kobayashi, Ayano Ishii, Masami Watanabe, Toyohiko Watanabe, Yasutomo Nasu

Nishinihon Journal of Urology 81 ( 3 ) 314 - 320 2019年6月

記述言語：日本語掲載種別：研究論文（国際会議プロシーディングス）出版者・発行元：The West Japan Urological Association

Renal autotransplantation (RAT) is a great surgical procedure for renal preservation, however it is underutilized due to its invasiveness. Robotic RATx has been in use since 2014. We conducted robotic RATx following a feasibility study using a porcine model. A 38-year-old woman with left ureteral stenosis was referred to us for urinary tract reconstruction. She had undergone an emergency Caesarean section for intraperitoneal bleeding secondary to left ovarian rupture four years earlier. After the surgery, she developed hypovolemic shock and disseminated intravascular coagulation (DIC) due to postoperative bleeding. Left internal iliac artery embolization saved her life. However, it resulted in a left ureteral stenosis measuring 2.7 cm that was located 5 cm from the left ureteral orifice, which required chronic ureteral stent exchange. Neither balloon dilation nor laser incision was successful. In addition, retrograde pyelography showed a possible left ureteropelvic junction obstruction. The patient was hesitant about the surgery because of its invasiveness. However, four years later, the patient elected to undergo robotic RATx. To date, only four cases of robotic RATx have been reported, including the current case. The indication of all four cases was extensive ureteral stenosis. Ileal ureter is frequently used for the treatment of extensive ureteral stenosis, but it can entail many problems including urine reabsorption resulting in metabolic acidosis. Ileal ureter is contraindicated for patients with a serum creatinine (sCr) level greater than 2 mg/dl. Even if sCr is less than 2 mg/dl, it is questionable whether sCr will remain less than 2 mg/ dl for the remainder of someone's life. Boari flap can give rise to vesicoureteral reflux causing urinary tract infection. RATx does not have those problems. Robotic RATx is a new, minimally invasive approach to renal preservation.

Comparison of the predictive value among inflammation-based scoring systems for bleomycin pulmonary toxicity in patients with germ cell tumors. 査読国際誌

Maruyama Y, Sadahira T, Araki M, Mitsui Y, Wada K, Edamura K, Kobayashi Y, Watanabe M, Watanabe T, Nasu Y

International journal of urology : official journal of the Japanese Urological Association 26 ( 8 ) 813 - 819 2019年5月

Comparison of longitudinal health-related quality-of-life outcomes between anterior and posterior surgical approaches to robot-assisted radical prostatectomy. 査読国際誌

Maruyama Y, Sadahira T, Araki M, Mitsui Y, Wada K, Tanimoto R, Kobayashi Y, Watanabe M, Watanabe T, Nasu Y

Journal of robotic surgery 14 ( 2 ) 255 - 260 2019年5月

Identification of Proteins Differentially Expressed by Adipose-derived Mesenchymal Stem Cells Isolated from Immunodeficient Mice. 査読国際誌

Nakashima Y, Nahar S, Miyagi-Shiohira C, Kinjo T, Kobayashi N, Kitamura S, Saitoh I, Watanabe M, Fujita J, Noguchi H

Int J Mol Sci 20 ( 11 ) E2672 2019年5月

Upregulation of mobility in pancreatic cancer cells by secreted S100A11 through activation of surrounding fibroblasts. 査読国際誌

Mitsui Y, Tomonobu N, Watanabe M, Kinoshita R, Sumardika W, Youyi C, Murata H, Yamamoto KI, Sadahira T, Rodrigo AGH, Takamatsu H, Araki K, Yamauchi A, Yamamura M, Fujiwara H, Inoue Y, Futami J, Saito K, Iioka H, Kondo E, Nishibori M, Toyooka S, Yamamoto Y, Nasu Y, Sakaguchi M

Oncology research 27 ( 8 ) 945 - 956 2019年4月

Very late relapse and management of clinical stage I seminoma after orchiectomy: A case report

Shogo Watari, Takuya Sadahira, Atsushi Takamoto, Yuki Maruyama, Yosuke Mitsui, Koichiro Wada, Ryuta Tanimoto, Morito Sugimoto, Yasuyuki Kobayashi, Motoo Araki, Masami Watanabe, Toyohiko Watanabe, Yasutomo Nasu, Hiroyuki Yanai

Nishinihon Journal of Urology 81 ( 1 ) 45 - 48 2019年2月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：The West Japan Urological Association

The patient was a 44-year-old man with a history of right radical orchiectomy for clinical stage I seminoma 14 years earlier followed by 6 years of surveillance, who presented with right lower leg edema. A thorough examination showed high testicular cancer-related tumor marker values, and based on computed tomography, a diagnosis of right pelvic lymph node metastasis was made. Salvage chemotherapy, consisting of bleomycin, etoposide, and cisplatin was administered followed by secondline chemotherapy with vinblastine, ifosfamide, and cisplatin. The tumor markers were confirmed to be negative, and the patient underwent retroperitoneal lymph node dissection. One year after surgery, there was no evidence of recurrence. Recurrence of clinical stage I seminoma more than 10 years after the initial diagnosis is rarely experienced. We report this case and review the management of clinical stage I seminoma.

A Novel Preservation Solution Containing a JNK Inhibitory Peptide Efficiently Improves Islet Yield for Porcine Islet Isolation. 査読国際誌

Noguchi H, Miyagi-Shiohira C, Nakashima Y, Ebi N, Hamada E, Tamaki Y, Kuwae K, Kitamura S, Kobayashi N, Saitoh I, Watanabe M

Transplantation 103 ( 2 ) 344 - 352 2019年2月

Robotic-assisted renal autotransplantation: Preliminary studies and future directions

Motoo Araki, Koichiro Wada, Kasumi Kawamura, Yuuki Maruyama, Yosuke Mitsui, Takuya Sadahira, Risa Kubota, Shingo Nishimura, Takashi Yoshioka, Yuichi Ariyoshi, Kei Fujio, Atsushi Takamoto, Morito Sugimoto, Katsumi Sasaki, Yasuyuki Kobayashi, Shin Ebara, Amano Hiroyuki, Masashi Inui, Masami Watanabe, Toyohiko Watanabe, Yasutomo Nasu

Endourology Progress: Technique, Technology and training 143 - 148 2019年1月

記述言語：英語掲載種別：論文集(書籍)内論文出版者・発行元：Springer Singapore

The indications for renal autotransplantation (RATx) include renal vascular trauma, thrombosis, stenosis, aneurysm, complex ureteral injuries, renal cell carcinoma, urolithiasis, retroperitoneal fibrosis, and loin pain-hematuria syndrome. Unfortunately, RATx is underutilized because of its invasiveness. The current gold standard approach to RATx is laparoscopic nephrectomy and open autotransplantation, which requires a large pelvic incision. Robotic-assisted renal autotransplantation (robotic RATx) is a new, minimally invasive approach that has been used since 2014. The first completely intracorporeal robotic RATx, used to repair a ureteral injury, was reported in 2014. Since then, only three cases have been reported, all from North America. After an initial porcine study, we conducted the fourth robotic RATx procedure. Robotic surgery has multiple advantages, such as providing a three-dimensional magnified view, navigating in narrow spaces, and fine suturing and dissection, which are particularly helpful in cases with desmoplastic changes. Most robotic RATx procedures have been used to repair ureteral injuries. These cases tend to have desmoplastic changes due to previous surgeries. Nephrectomy and RATx are technically challenging, hence robotic surgery may be the best option in patients with complex and severe desmoplastic changes. The disadvantages of robotic RATx include the length of surgery and cost. However, with continued use, both the operative time and the cost should decrease. In conclusion, robotic RATx is a new, minimally invasive approach to renal preservation.

DOI： 10.1007/978-981-13-3465-8_18

A Liquid Chromatography with Tandem Mass Spectrometry-Based Proteomic Analysis of Primary Cultured Cells and Subcultured Cells Using Mouse Adipose-Derived Mesenchymal Stem Cells. 査読国際誌

Nakashima Y, Nahar S, Miyagi-Shiohira C, Kinjo T, Kobayashi N, Saitoh I, Watanabe M, Fujita J, Noguchi H

Stem Cells Int 2019 7274057 - 7274057 2019年1月

Induction of Expandable Tissue-Specific Progenitor Cells from Human Pancreatic Tissue through Transient Expression of Defined Factors. 査読

Noguchi H, Miyagi-Shiohira C, Nakashima Y, Kinjo T, Kobayashi N, Saitoh I, Watanabe M, Shapiro AMJ, Kin T

Mol Ther Methods Clin Dev 13 243 - 252 2019年1月

The Novel Combination of Nitroxoline and PD-1 Blockade, Exerts a Potent Antitumor Effect in a Mouse Model of Prostate Cancer. 査読国際共著国際誌

Naijin Xu, Linglong Huang, Xiezhao Li, Masami Watanabe, Chaoming Li, Abai Xu, Chunxiao Liu, Qiang Li, Motoo Araki, Koichiro Wada, Yasutomo Nasu, Peng Huang

International journal of biological sciences 15 ( 5 ) 919 - 928 2019年

Pelvic magnetic resonance imaging parameters predict urinary incontinence after robot-assisted radical prostatectomy. 査読国際誌

Sadahira T, Mitsui Y, Araki M, Maruyama Y, Wada K, Edamura K, Kobayashi Y, Watanabe M, Watanabe T, Nasu Y

Lower urinary tract symptoms 11 ( 3 ) 122 - 126 2018年12月

Burned-out Prostate Cancer ? Primary Metastatic Cancer Not Detected on Repeat Biopsy. 査読

Mitsui Y, Sadahira T, Maruyama Y, Wada K, Tanimoto R, Sugimoto M, Araki M, Watanabe M, Yanai H, Watanabe T, Nasu Y

Acta medica Okayama 72 ( 6 ) 605 - 609 2018年12月

The 3-D Volumetric Measurement Including Resected Specimen for Predicting Renal Function AfterRobot-assisted Partial Nephrectomy. 査読国際誌

Mitsui Y, Sadahira T, Araki M, Maruyama Y, Nishimura S, Wada K, Kobayashi Y, Watanabe M, Watanabe T, Nasu Y

Urology 125 104 - 110 2018年12月

A Comparison of Proteins Expressed between Human and Mouse Adipose-Derived Mesenchymal Stem Cells by a Proteome Analysis through Liquid Chromatography with Tandem Mass Spectrometry. 査読国際誌

Nahar S, Nakashima Y, Miyagi-Shiohira C, Kinjo T, Kobayashi N, Saitoh I, Watanabe M, Noguchi H, Fujita J

Int J Mol Sci 19 ( 11 ) E3497 2018年11月

Induction of Expandable Adipose-Derived Mesenchymal Stem Cells from Aged Mesenchymal Stem Cells by a Synthetic Self-Replicating RNA. 査読

Miyagi-Shiohira C, Nakashima Y, Kobayashi N, Kitamura S, Saitoh I, Watanabe M, Noguchi H

Int J Mol Sci 19 ( 11 ) E3489 2018年11月

Loss of psoas major muscle volume during systemic chemotherapy is related to worse prognosis in testicular cancer. 査読国際誌

Mitsui Y, Sadahira T, Araki M, Maruyama Y, Wada K, Tanimoto R, Kobayashi Y, Watanabe M, Watanabe T, Nasu Y

Japanese journal of clinical oncology 49 ( 2 ) 183 - 189 2018年11月

Cytokines in adipose-derived mesenchymal stem cells promote the healing of liver disease. 査読

Nahar S, Nakashima Y, Miyagi-Shiohira C, Kinjo T, Toyoda Z, Kobayashi N, Saitoh I, Watanabe M, Noguchi H, Fujita J

World J Stem Cells 10 ( 11 ) 146 - 159 2018年11月

Induction of Expandable Adipose-Derived Mesenchymal Stem Cells from Aged Mesenchymal Stem Cells by a Synthetic Self-Replicating RNA

Chika Miyagi-Shiohira, Yoshiki Nakashima, Naoya Kobayashi, Shinji Kitamura, Issei Saitoh, Masami Watanabe, Hirofumi Noguchi

INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES 19 ( 11 ) 2018年11月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：MDPI

Adipose-derived mesenchymal stem cells (ADSCs) have attracted attention due to their potential for use in the treatment of various diseases. However, the self-renewal capacity of ADSCs is restricted and their function diminishes during passage. We previously generated induced tissue-specific stem cells from mouse pancreatic cells using a single synthetic self-replicating Venezuelan Equine Encephalitis (VEE)-reprogramming factor (RF) RNA replicon (SR-RNA) expressing the reprogramming factors POU class 5 homeobox 1 (OCT4), Krueppel-like factor 4 (KLF4), Sex determining region Y-box 2 (SOX2), and Glis Family Zinc Finger 1 (GLIS1). This vector was used to generate induced pluripotent stem (iPS) cells. Here, we applied this SR-RNA vector to generate human iTS cells from aged mesenchymal stem cells (hiTS-M cells) deficient in self-renewal that were derived from adipose tissue. These hiTS-M cells transfected with the SR-RNA vector survived for 15 passages. The hiTS-M cells expressed cell surface markers similar to those of human adipose-derived mesenchymal stem cells (hADSCs) and differentiated into fat cells and osteoblasts. Global gene expression profiling showed that hiTS-M cells were transcriptionally similar to hADSCs. These data suggest that the generation of iTS cells has important implications for the clinical application of autologous stem cell transplantation.

DOI： 10.3390/ijms19113489

Red nodular melanoma of the penile foreskin: A case report and literature review. 査読国際誌

Maruyama Y, Sadahira T, Mitsui Y, Wada K, Tanimoto R, Kobayashi Y, Araki M, Watanabe M, Watanabe T, Nasu Y

Molecular and clinical oncology 9 ( 4 ) 449 - 452 2018年10月

A Liquid Chromatography with Tandem Mass Spectrometry-Based Proteomic Analysis of the Proteins Secreted by Human Adipose-Derived Mesenchymal Stem Cells. 査読国際誌

Nakashima Y, Nahar S, Miyagi-Shiohira C, Kinjo T, Toyoda Z, Kobayashi N, Saitoh I, Watanabe M, Fujita J, Noguchi H

Cell Transplant 27 ( 10 ) 1469 - 1494 2018年10月

A Comparison of the Preservation of Mouse Adipose Tissue-Derived Mesenchymal Stem Cells Using the University of Wisconsin Solution and Hank's Balanced Salt Solution. 査読国際誌

Nahar S, Nakashima Y, Miyagi-Shiohira C, Kinjo T, Kobayashi N, Saitoh I, Watanabe M, Noguchi H, Fujita J

Stem Cells Int 2018 1625464 - 1625464 2018年9月

Bufalin suppresses the proliferation and metastasis of renal cell carcinoma by inhibiting the PI3K/Akt/mTOR signaling pathway. 査読国際誌

Jinlin Xie, Wenfeng Lin, Linglong Huang, Naijin Xu, Abai Xu, Binshen Chen, Masami Watanabe, Chunxiao Liu, Peng Huang

Oncology letters 16 ( 3 ) 3867 - 3873 2018年9月

記述言語：英語掲載種別：研究論文（学術雑誌）

Bufalin, one of the active ingredients of the Chinese drug Chan su, exhibits significant antitumor activity against various cancer types. However, the role of bufalin in renal cell carcinoma (RCC) remains unclear. In the present study, it was demonstrated that bufalin inhibited cell proliferation, blocked the cell cycle in the G2/M phase, and reduced the metastasis of human RCC ACHN cells via the upregulation of p21waf/cip1 and E-cadherin and the downregulation of cyclin dependent kinase 1, cyclin B1, N-cadherin, and hypoxia-inducible factor-1α (HIF-1α). Further mechanistic study revealed that bufalin reduced the expression of phosphorylated (phospho)-Akt and phospho-mammalian target of rapamycin (mTOR). Moreover, HIF-1α expression may be regulated through the inhibition of the phosphoinositide 3-kinase (PI3K)/protein kinase B (Akt)/mTOR signaling pathway. Thus, the present results suggest that bufalin induces cell cycle arrest and suppresses metastasis; this process may be associated with the PI3K/Akt/mTOR signaling pathway. Accordingly, it is suggested that bufalin is a therapeutic agent for RCC.

DOI： 10.3892/ol.2018.9111

Clinical impact of abdominal fat distribution measured by 3-D computed tomography volumetry on post-transplant renal function in recipients after living kidney transplantation: a retrospective study. 査読

Mitsui Y, Sadahira T, Araki M, Maruyama Y, Wada K, Tanimoto R, Kobayashi Y, Watanabe M, Watanabe T, Nasu Y

Clinical and experimental nephrology 23 ( 3 ) 415 - 424 2018年9月

排尿障害を伴う女性尿道憩室に対して手術治療を行った1例

松尾聡子, 杉本盛人, 佐久間貴文, 坪井一朗, 本郷智拡, 三井將雄, 河村香澄, 和田里章悟, 丸山雄樹, 光井洋介, 定平卓也, 前原貴典, 大岩裕子, 西村慎吾, 高本篤, 佐古智子, 和田耕一郎, 谷本竜太, 小林泰之, 荒木元朗, 石井亜矢乃, 渡部昌実, 渡邉豊彦, 那須保友, 中村あや, 津島知靖

西日本泌尿器科 80 ( 9 ) 495 - 495 2018年9月

Comparison Between Modified Extracellular-Type Trehalose-Containing Kyoto Solution and University of Wisconsin Solution in 18-Hour Pancreas Preservation for Islet Transplantation. 査読

Hamada E, Ebi N, Miyagi-Shiohira C, Tamaki Y, Nakashima Y, Kobayashi N, Saitoh I, Watanabe M, Kinjo T, Noguchi H

Pancreas 47 ( 7 ) e46 - e47 2018年8月

Characterization of induced tissue-specific stem cells from pancreas by a synthetic self-replicative RNA. 査読国際誌

Miyagi-Shiohira C, Nakashima Y, Kobayashi N, Saitoh I, Watanabe M, Noguchi H

Sci Rep 8 ( 1 ) 12341 - 12341 2018年8月

Endogenous Leu332Gln mutation in p53 disrupts the tetramerization ability in a canine mammary gland tumor cell line 査読

Kazuhiko Ochiai, Daigo Azakami, Masami Morimatsu, Hinako Hirama, Shota Kawakami, Takayuki Nakagawa, Masaki Michishita, Ai S. Egusa, Takanori Sasaki, Masami Watanabe, Toshinori Omi

Oncology Reports 40 ( 1 ) 488 - 494 2018年7月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：Spandidos Publications

Mutations in the p53 gene are associated with more than half of all human cancers. These mutations often cause a disruption of the tumor-suppressor function of p53 and induce genomic instabilities. Wild-type p53 requires tetramerization to function as an initiator of cell cycle arrest and apoptosis. Although alterations in p53 tetramerization caused by mutation have been well studied, there are few cell lines containing an endogenous mutation in the tetramerization domain of p53. Here, we report the discovery of a canine mammary gland tumor cell line CTB-m2, which contains the Leu332Gln (L332Q) mutation corresponding to Leu344 in the tetramerization domain of human p53. Although CTB-m2 cells are genetically heterozygous for the Leu332Gln mutation, the mutant mRNA was almost exclusively expressed. CTB-m2 cells showed enhanced cell proliferation compared to wild-type p53-expressing CTB-m cells of the same lineage. A p53 tetramerization reporter assay showed that the ability of the p53 mutant to form tetramers was signifcantly lower than that of wild-type p53. An immunoblot analysis of cross-linked p53 oligomerized forms demonstrated that the L332Q mutant lacked the ability to form tetramers but retained the ability to form dimers. These data suggest that the p53 mutant cell line CTB-m2 could be a useful tool for analyzing the precise tetramerization mechanisms of p53 and verifying the effects of therapeutic agents against tumors expressing p53 mutants that lack the ability to tetramerize.

DOI： 10.3892/or.2018.6409

Modified cell-permeable JNK inhibitors efficiently prevents islet apoptosis and improves the outcome of islet transplantation. 査読

Noguchi H, Miyagi-Shiohira C, Nakashima Y, Ebi N, Hamada E, Tamaki Y, Kuwae K, Kobayashi N, Saitoh I, Watanabe M

Sci Rep 8 ( 1 ) 11082 2018年7月

A Liquid Chromatography with Tandem Mass Spectrometry-Based Proteomic Analysis of Cells Cultured in DMEM 10% FBS and Chemically Defined Medium Using Human Adipose-Derived Mesenchymal Stem Cells. 査読

Nakashima Y, Nahar S, Miyagi-Shiohira C, Kinjo T, Kobayashi N, Saitoh I, Watanabe M, Fujita J, Noguchi H

Int J Mol Sci 19 ( 7 ) E2042 2018年7月

膵がん進展に導く膵がん細胞間質線維芽細胞クロストークを介在する分泌性S100A11 受容体RAGE連携の役割

光井洋介, 山本健一, Sumardika I Wayan, 木下理恵, 村田等, 二見淳一郎, 高松仁, 山本靖彦, 西堀正洋, 豊岡伸一, 渡部昌実, 那須保友, 阪口政清

日本がん免疫学会総会プログラム・抄録集 22回 156 - 156 2018年7月

新規抗癌免疫療法としてのAd-REIC遺伝子治療とその臨床効果

定平卓也, 渡部昌実, 那須保友

日本がん免疫学会総会プログラム・抄録集 22回 69 - 69 2018年7月

A Liquid Chromatography with Tandem Mass Spectrometry-Based Proteomic Analysis of Cells Cultured in DMEM 10% FBS and Chemically Defined Medium Using Human Adipose-Derived Mesenchymal Stem Cells

Yoshiki Nakashima, Saifun Nahar, Chika Miyagi-Shiohira, Takao Kinjo, Naoya Kobayashi, Issei Saitoh, Masami Watanabe, Jiro Fujita, Hirofumi Noguchi

INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES 19 ( 7 ) 2018年7月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：MDPI

Human adipose-derived mesenchymal stem cells (hADSCs) are representative cell sources for cell therapy. Classically, Dulbecco's Modified Eagle's medium (DMEM) containing 10% fetal bovine serum (FBS) has been used as culture medium for hADSCs. A chemically defined medium (CDM) containing no heterologous animal components has recently been used to produce therapeutic hADSCs. However, how the culture environment using a medium without FBS affects the protein expression of hADSC is unclear. We subjected hADSCs cultured in CDM and DMEM (10% FBS) to a protein expression analysis by tandem mass spectrometry liquid chromatography and noted 98.2% agreement in the proteins expressed by the CDM and DMEM groups. We classified 761 proteins expressed in both groups by their function in a gene ontology analysis. Thirty-one groups of proteins were classified as growth-related proteins in the CDM and DMEM groups, 16 were classified as antioxidant activity-related, 147 were classified as immune system process-related, 557 were involved in biological regulation, 493 were classified as metabolic process-related, and 407 were classified as related to stimulus responses. These results show that the trend in the expression of major proteins related to the therapeutic effect of hADSCs correlated strongly in both groups.

DOI： 10.3390/ijms19072042

Comparison of Tissue Loading Before and After the Creation of a Continuous Density Gradient in Porcine Islet Purification 査読

Miyagi-Shiohira Chika, Nakashima Yoshiki, Ebi Nana, Hamada Eri, Tamaki Yoshihito, Kuwae Kazuho, Kobayashi Naoya, Saitoh Issei, Watanabe Masami, Kinjo Takao, Noguchi Hirofumi

CELL MEDICINE 10 2018年6月

A Comparison of Pancreatic Islet Purification using Iodixanol with University of Wisconsin Solution and with Na-Lactobionate and Histidine Solution 査読

Nakashima Yoshiki, Miyagi-Shiohira Chika, Ebi Nana, Hamada Eri, Tamaki Yoshihito, Kuwae Kazuho, Kobayashi Naoya, Saitoh Issei, Watanabe Masami, Kinjo Takao, Noguchi Hirofumi

CELL MEDICINE 10 2018年6月

Prognostic impact of bleomycin pulmonary toxicity on the outcomes of patients with germ cell tumors 査読

Yuki Maruyama, Takuya Sadahira, Yosuke Mitsui, Motoo Araki, Koichiro Wada, Ryuta Tanimoto, Yasuyuki Kobayashi, Masami Watanabe, Toyohiko Watanabe, Yasutomo Nasu

Medical Oncology 35 ( 6 ) 80 2018年6月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：Humana Press Inc.

Bleomycin pulmonary toxicity (BPT) has been well described in patients with germ cell tumors treated with bleomycin etoposide and cisplatin chemotherapy (BEP). To assess the prognostic impact of BPT, we retrospectively identified 52 patients who underwent bleomycin etoposide and cisplatin chemotherapy from 2008 to 2017 in our institution, and evaluated the risk factors of BPT and its effect on prognosis. Patients who had received chemotherapy at another institution were excluded. BPT was defined as bleomycin discontinuation in response to pulmonary function test decline, pulmonary symptoms, or interstitial pneumonia on computed tomography without infection. We divided the patients into two groups according to this definition: BPT and non-BPT. Their median age was 34.2 years, and their median body mass index was 22.8 kg/m2. Twenty patients had a smoking history, 37 were diagnosed with non-seminoma, and 20 had lung metastasis. The median cumulative bleomycin dose was 270 mg/body. Fifteen patients were classified into the BPT group and 37 into the non-BPT group. Only body mass index &lt
22 was identified as a predictor of BPT in multivariable logistic models. Age or use of granulocyte-colony stimulating factor did not have a significant impact. Kaplan–Meier analysis revealed that the presence of BPT had no significant impact on either 5-year overall survival or progression-free survival. Lower body mass index can increase the risk of BPT in patients with germ cell tumors undergoing BEP. However, discontinuation of bleomycin with BPT does not adversely influence the survival outcomes.

DOI： 10.1007/s12032-018-1140-5

The Development of Cancer through the Transient Overexpression of Reprogramming Factors 査読

Miyagi-Shiohira Chika, Nakashima Yoshiki, Kobayashi Naoya, Saitoh Issei, Watanabe Masami, Noguchi Yasufumi, Kinjo Takao, Noguchi Hirofumi

CELL MEDICINE 10 2018年5月

Role of Egr1 on Pancreatic Endoderm Differentiation 査読

Tsugata Takako, Nikoh Naruo, Kin Tatsuya, Miyagi-Shiohira Chika, Nakashima Yoshiki, Saitoh Issei, Noguchi Yasufumi, Ueki Hideo, Watanabe Masami, Kobayashi Naoya, Shapiro Andrew, M. James, Noguchi Hirofumi

CELL MEDICINE 10 2018年5月

Evaluation of Islet Purification Methods for Making a Continuous Density Gradient and Loading Tissue 査読

Ebi Nana, Miyagi-Shiohira Chika, Hamada Eri, Tamaki Yoshihito, Masamoto Mariko, Makishi Erika, Nakashima Yoshiki, Kobayashi Naoya, Saitoh Issei, Watanabe Masami, Noguchi Yasufumi, Kinjo Takao, Noguchi Hirofumi

CELL MEDICINE 10 2018年5月

Gene therapy for prostate cancer: Current status and future prospects

Yasutomo Nasu, Masami Watanabe

Hormone Therapy and Castration Resistance of Prostate Cancer 397 - 406 2018年5月

記述言語：英語掲載種別：論文集(書籍)内論文出版者・発行元：Springer Singapore

Prostate is an ideal target organ for gene therapy as a translational research. It has some advantages as follows: prostate is not a life keeping organ, can be approached easily by ultrasound as a routine clinical practice and PSA is a sensitive and useful tumor marker for the evaluation of clinical response. Intraprostatic therapeutic gene transduction (in situ gene therapy) is one of the potent therapeutic options for prostate cancer gene therapy aiming at antimetastatic benefits through the generation of immune cell-mediated cytotoxic activities that affect not only the primary tumor but also metastatic lesion. In this chapter, current outcome and future prospect of prostate cancer gene therapy are discussed.

DOI： 10.1007/978-981-10-7013-6_40

Adhesion characteristics of porcine pancreatic islets and exocrine tissue to coating materials 査読

Yoshiki Nakashima, Chika Miyagi-Shiohira, Naoya Kobayashi, Issei Saitoh, Masami Watanabe, Hirofumi Noguchi

Islets 10 ( 3 ) 2018年5月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：Taylor and Francis Inc.

Since the report of the Edmonton protocol in 2000, islet transplantation has been implemented worldwide, and xenotransplantation using porcine islets has also been reported. In addition, many basic experiments using pancreatic islets and exocrine tissue after isolation have been reported. Recently, exocrine cells have been found to be essential for inducing the differentiation of pancreatic islets. Therefore, the importance of the culture conditions for pancreatic tissue when conducting experiments using pancreatic tissue is also increasing. In this study, we focused on the coat material and examined the adhesive properties of porcine pancreatic islets and exocrine tissue after isolation. Porcine islet isolation was performed, and isolated islets (purity ≥95%) and exocrine tissue (purity ≥99%) were used to achieve adhesion to several extracellular matrixes, fibronectin, collagen type I, collagen type IV, laminin I, fibrinogen, and bovine serum albumin (BSA). DMEM with 0.5% FBS was used as the assay medium. For exocrine tissue, the adhesion was promoted in fibronectin, collagen type I, laminin I, and fibrinogen. The adhesive ability to fibronectin was more than twice that to BSA, while the adhesive ability to collagen type I, laminin I, and fibrinogen was less than twice that to BSA. For islets, the adhesive ability to fibronectin was weaker than that of exocrine tissue. Furthermore, the adhesion effect in fibronectin was obtained within 30 minutes and in medium containing little serum for both islets and exocrine tissues. These data suggest that fibronectin may be useful for the adhesion of pancreatic tissue.

DOI： 10.1080/19382014.2018.1460294

Programmed death ligand 1 expression in bladder. 査読

Peng Zhou, Qiongren Wang, Chongshan Wang, Xiezhao L, Wei Du, Chunxiao Liu, Masami Watanabe, Peng Huang, Abai Xu

Int J Clin Exp Pathol 2018年4月

R132 mutations in canine isocitrate dehydrogenase 1 (IDH1) lead to functional changes 査読

Shota Kawakami, Kazuhiko Ochiai, Daigo Azakami, Yuiko Kato, Masaki Michishita, Masami Morimatsu, Toshina Ishiguro-Oonuma, Eri Onozawa, Masami Watanabe, Toshinori Omi

Veterinary Research Communications 42 ( 1 ) 49 - 56 2018年3月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：Springer Netherlands

Glioma is the second most common intracranial neoplasia in dogs, but the pathogenic mechanisms remain unclear. In humans, isocitrate dehydrogenase 1 (IDH1) is frequently mutated in gliomas. Although almost all human IDH1 mutations have been identified as involving the Arg132 codon, few studies have reported structural, functional, and mutational information for canine IDH1. Therefore, in this study, we cloned the canine IDH1 homologue and used PCR mutagenesis to substitute the wildtype (WT) Arg132 with His (R132H) or Ser (R132S). WT and mutated IDH1 were overexpressed in HeLa cells, and their presence was confirmed by immunoblotting and immunocytochemistry using mutation-specific antibodies. The IDH1 activity between WT, R132H, and R132S transfectants was compared by measuring the production of NADH and NADPH. NADPH production in R132H and R132S transfectants was lower than that in WT, but NADH levels were not significantly different. Finally, we detected increased expression of hypoxia inducible factor 1 alpha (HIF-1α) in the R132H and R132S transfectants. These results indicated that the canine IDH1 Arg132 mutation has the potential to induce carcinogenesis in canine somatic cells.

DOI： 10.1007/s11259-017-9707-8

Induction of cells with prostate cancer stem-like properties from mouse induced pluripotent stem cells via conditioned medium. 査読国際誌

Naijin Xu, Xiezhao Li, Masami Watanabe, Hideo Ueki, Hao Hu, Na Li, Motoo Araki, Koichiro Wada, Abai Xu, Chunxiao Liu, Yasutomo Nasu, Peng Huang

American journal of cancer research 8 ( 8 ) 1624 - 1632 2018年

記述言語：英語掲載種別：研究論文（学術雑誌）

Cancer stem cells (CSCs) that closely correlated with tumor growth, metastasis, provide a plausible explanation for chemoresistance and cancer relapse. CSCs are usually isolated and enriched from carcinoma cells, which is inconvenient, low-efficient, and even unreliable. Here, we converted mouse induced pluripotent stem cells (miPSCs) into prostate cancer stem-like cells with carcinoma microenvironment following exposure to conditioned medium (CM) derived from RM9, a mouse prostate cancer cell line. These transformed cells, termed as miPS-RM9CM, displayed CSCs properties, including spheroids morphology and expression of both stemness genes and cancer stem cells surface markers, such as Oct3/4, Sox2, Nanog, Klf-4, c-Myc, CD44, and CD133. In addition, in vivo transplantation experiment was performed to confirm the tumorigenicity. Furthermore, we used the model to assess conventional chemotherapeutic agent, docetaxel. The results showed that miPS-RM9CM cells exhibited increased resistance to docetaxel, however, high susceptibility to the cancer cell stemness inhibitor I (BBI-608). Our current study demonstrates that CM from cultured RM9 cells play a crucial role in the determination of cell fate from miPSCs to cancer stem-like cells and provide a potentially valuable system for the study of CSCs.

Glaucocalyxin A induces G2/M cell cycle arrest and apoptosis through the PI3K/Akt pathway in human bladder cancer cells. 査読国際誌

Wenfeng Lin, Jinlin Xie, Naijin Xu, Linglong Huang, Abai Xu, Hulin Li, Chaoming Li, Yubo Gao, Masami Watanabe, Chunxiao Liu, Peng Huang

International journal of biological sciences 14 ( 4 ) 418 - 426 2018年

記述言語：英語掲載種別：研究論文（学術雑誌）

Glaucocalyxin A (GLA), a major component isolated from Rabdosia japonica, has been proven to show anti-bacterial and anti-tumor biological characteristics according to previous studies. However, its potential effect on bladder cancer remains unknown. The present research aims to investigate the underlying mechanism in treating bladder cancer in vivo and in vitro. Cell proliferation was analyzed by CCK-8 assay and colony formation. Flow cytometry was used to measure the cell cycle distribution and apoptosis. The expressions of the cell cycle and apoptosis-related proteins were detected by western blotting and immunofluorescence staining. Meanwhile, the in vivo study was performed to evaluate the anti-tumor effect on a UMUC3 subcutaneous tumor of NOD/SCID mice model. GLA suppressed colony-formation ability, triggered G2/M arrest and promoted apoptosis of UMUC3 cells in a dose-dependent manner. Furthermore, western blotting showed that GLA downregulated the expressions of PI3K p85, p-Akt, Bcl-2, CDK1, Cyclin B1 whereas upregulated the levels of PTEN, Bax, Cleaved Caspase-3. In vivo, GLA at a dosage of 20 mg/kg significantly inhibited tumor growth compared with the control group by intraperitoneal injection. These results suggested that GLA-related G2/M arrest and apoptosis in UMUC3 cells were mediated by a suppressed PI3K/Akt signaling pathway, which regulated p21Waf1/Cip1 as well as intrinsic caspase cascade. Collectively, our observations could help to develop new drugs targeting the PI3K/Akt pathway for the treatment of bladder cancer.

DOI： 10.7150/ijbs.23602

Functional alteration of canine isocitrate dehydrogenase 2 (IDH2) via an R174K mutation 査読

Shota Kawakami, Kazuhiko Ochiai, Yuiko Kato, Masaki Michishita, Hinako Hirama, Ryo Obara, Daigo Azakami, Masami Watanabe, Toshinori Omi

Journal of Veterinary Medical Science 80 ( 1 ) 85 - 91 2018年

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：Japanese Society of Veterinary Science

Gliomas are common intracranial neoplasias in dogs. However, the underlying pathogenic mechanisms remain unclear. In humans, isocitrate dehydrogenase 2 (IDH2) is often mutated in gliomas. Although almost human IDH2 mutations have been identified at the Arg172 codon, few studies have reported structural, functional or mutational information for canine IDH2. In this study, we cloned the full-length canine IDH2 (cIDH2) cDNA and substituted wild type Arg174 (cIDH2 WT: corresponding to R172 of human IDH2) with Lys (cIDH2 R174K). The cIDH2 WT and R174K proteins were overexpressed in HeLa cells, and their presence was confirmed using an anti-human IDH2-WT mAb (clone: KrMab-3) and an anti-IDH2-R172K mAb (clone: KMab-1). The IDH2 activity between cIDH2 WT and cIDH2 R174K transfectants was compared by measuring the production of NADH and NADPH. NADPH production was lower for cIDH2 R174K than that for cIDH2 WT transfectants. Finally, we detected increased expression of hypoxia inducible factor-1 alpha (HIF-1α) in cIDH2 R174K transfectants. This indicates that mutations at R174 can potentially induce carcinogenesis in canine somatic cells.

DOI： 10.1292/jvms.17-0362

Properties of the feline tumour suppressor reduced expression in immortalized cells (REIC/Dkk-3) 査読

K. Ochiai, H. Oda, S. Shono, Y. Kato, S. Sugihara, S. Nakazawa, D. Azakami, M. Michishita, E. Onozawa, M. Bonkobara, T. Sako, L. Shun-Ai, H. Ueki, M. Watanabe, T. Omi

VETERINARY AND COMPARATIVE ONCOLOGY 15 ( 4 ) 1181 - 1186 2017年12月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：WILEY

Reduced expression in immortalized cells (REIC/Dkk-3), a member of the human Dickkopf (Dkk) family, is a growth suppressor in human and canine mammary tumours. Mammary gland tumours are common neoplasms with high malignancy in female cats. The purpose of this study was to clone the feline REIC/Dkk-3 homolog, investigate its expression in cell lines established from feline mammary gland tumours, and test its tumour suppressor function. Western blot analysis revealed that expression of the REIC/Dkk-3 protein was reduced in feline mammary carcinoma cell lines. Forced expression of REIC/Dkk-3 induced apoptosis in feline mammary tumour cell lines. These results demonstrate that REIC/Dkk-3 expression, which is downregulated in feline mammary tumour cell lines, results in the induction of apoptosis in these cells. Our findings suggest that feline REIC/Dkk-3 represents a potential molecular target for the development of therapies against feline mammary cancers.

DOI： 10.1111/vco.12254

Exogenous DKK-3/REIC inhibits Wnt/β-catenin signaling and cell proliferation in human kidney cancer KPK1. 査読

Xu J, Sadahira T, Kinoshita R, Li S, Huang P, Wada K, Araki M, Ochiai K, Noguchi H, Sakaguchi M, Nasu Y, Watanabe M

Oncol Lett 14 ( 5 ) 5638 - 5642 2017年11月

Promising therapeutic efficacy of a novel reduced expression in immortalized cells/dickkopf-3 expressing adenoviral vector for hepatocellular carcinoma 査読

Hiroaki Sawahara, Hidenori Shiraha, Daisuke Uchida, Hironari Kato, Ryo Kato, Atsushi Oyama, Teruya Nagahara, Masaya Iwamuro, Shigeru Horiguchi, Koichiro Tsutsumi, Mari Mandai, Tetsushige Mimura, Nozomu Wada, Yasuto Takeuchi, Kenji Kuwaki, Hideki Onishi, Shinichiro Nakamura, Masami Watanabe, Masakiyo Sakaguchi, Akinobu Takaki, Kazuhiro Nouso, Takahito Yagi, Yasutomo Nasu, Hiromi Kumon, Hiroyuki Okada

JOURNAL OF GASTROENTEROLOGY AND HEPATOLOGY 32 ( 10 ) 1769 - 1777 2017年10月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：WILEY

Background and Aim: Reduced expression in immortalized cells (REIC)/dickkopf-3 (Dkk-3) is a tumor suppressor gene that is downregulated in various cancers. In our previous study of prostate cancer, the REIC/Dkk-3-expressing adenoviral vector (Ad-REIC) was found to induce cancer-selective apoptosis. This study recently developed a novel super gene expression (SGE) system and used this system to re-construct an Ad-REIC vector, termed the Ad-SGE-REIC, to achieve more effective therapeutic outcomes. In this study, the therapeutic effects of Ad-SGE-REIC on hepatocellular carcinoma (HCC) was assessed.
Methods: Human HCC cell lines (HLE, Huh7, HepG2, HLF, SK-Hep1, and PLC), human HCC tissues, and mouse HCC cell line (Hepa1-6) were used in this study. REIC/Dkk-3 expression was assessed by immunoblotting and immunohistochemistry. The relative cell viability and the apoptotic effect were examined in vitro, and the anti-tumor effects of Ad-SGE-REIC treatment were analyzed in the mouse xenograft model. This study additionally assessed anti-tumor immunological effects on the immunocompetent mice.
Results: REIC/Dkk-3 expression was decreased in HCC cell lines and HCC tissues. Ad-SGE-REIC reduced cell viability and induced apoptosis in HCC cell lines (HLE and Huh7), inhibited tumor growth in the mouse xenograft model, and demonstrated in vivo anti-cancer immunostimulatory effects on the HCC cell line (Hepa1-6).
Conclusions: Ad-SGE-REIC treatment not only enhanced cell killing effects in vitro but also elicited significant therapeutic effects, with tumor growth suppression, in vivo. REIC/Dkk-3 gene therapy using Ad-SGE-REIC potentially represents an innovative new therapeutic tool for HCC.

DOI： 10.1111/jgh.13757

Overexpression of REIC/Dkk-3 suppresses the expression of CD147 and inhibits the proliferation of human bladder cancer cells 査読

Yuhei Horikawa, Masami Watanabe, Takuya Sadahirai, Yuichi Ariyoshi, Yasuyuki Kobayashi, Motoo Araki, Koichiro Wada, Kazuhiko Ochiai, Shun-Ai Li, Yasutomo Nasu

ONCOLOGY LETTERS 14 ( 3 ) 3223 - 3228 2017年9月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：SPANDIDOS PUBL LTD

Our group previously developed an adenoviral vector encoding the REIC/Dkk-3 gene (Ad-REIC), a tumor suppressor, for cancer gene therapy. The Ad-REIC agent induces apoptosis and inhibits invasion in a number of cancer cell lines; however, the molecular mechanisms underlying its effects remain unclear. Cluster of differentiation 147 (CD147), also known as extracellular matrix metalloproteinase inducer (EMMPRIN), is a key molecule that promotes cancer proliferation and invasion. In order to elucidate the therapeutic mechanism of Ad-REIC, its effect on the expression of CD147 in human bladder cancer KK47 cells was investigated. Treatment with Ad-REIC markedly downregulated the expression of CD147 and significantly inhibited cellular proliferation. Since the expression of CD147 is reported to be under the positive control of mitogen-activated protein kinase (MAPK) signaling and the c-Myc protein, the correlations between the expression of CD147 and the activation of MAPKs or the expression of c-Myc were examined. Unexpectedly, no positive correlation was observed between the level of CD147 and the potential regulators that were assessed, indicating that another signaling pathway is responsible for the downregulation of CD147. The results from the present study demonstrate that Ad-REIC treatment can significantly downregulate the expression of CD147 in bladder cancer cells. Downregulation of the cancer-progression factor CD147 may be a novel mechanism that underlies the therapeutic effects of Ad-REIC treatment.

DOI： 10.3892/ol.2017.6548

Robotic Renal Autotransplantation: First Case Outside of North America 査読

Motoo Araki, Koichiro Wada, Yosuke Mitsui, Takuya Sadahira, Risa Kubota, Shingo Nishimura, Takashi Yoshioka, Yuichi Ariyoshi, Kei Fujio, Atsushi Takamoto, Morito Sugimoto, Katsumi Sasaki, Yasuyuki Kobayashi, Shin Ebara, Hideki Taninishi, Hiroyuki Amano, Masashi Inui, Masami Watanabe, Toyohiko Watanabe, Yasutomo Nasu

ACTA MEDICA OKAYAMA 71 ( 4 ) 351 - 355 2017年8月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：OKAYAMA UNIV MED SCHOOL

A 38-year-old woman with a 2.7-cm left ureteral stenosis requiring chronic ureteral stent exchange elected to undergo robotic renal autotransplantation. Left ureteropelvic junction obstruction (UPJO) was also suspected. Robotic donor nephrectomy contributed to the fine dissection for desmoplastic changes. The kidney was removed through a Gelport and examined on ice. UPJO was not seen. An end-to-side robotic anastomosis was created between the renal and external iliac vessels. The console time was 507 min, and the warm ischemia time was 4 min 5 sec. She became stent-free. Robotic renal autotransplantation is a new, minimally invasive approach to renal preservation.

Robust cancer-specific gene expression by a novel cassette with hTERT and CMV promoter elements 査読

Masakiyo Sakaguchi, Takuya Sadahira, Hideo Ueki, Rie Kinoshita, Hitoshi Murata, Ken-Ichi Yamamoto, Junichiro Futami, Yasutomo Nasu, Kazuhiko Ochiai, Hiromi Kumon, Nam-Ho Huh, Masami Watanabe

ONCOLOGY REPORTS 38 ( 2 ) 1108 - 1114 2017年8月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：SPANDIDOS PUBL LTD

We developed and validated a novel hTERT/CMV promoter element-driven gene expression cassette that can robustly enhance cancer-specific gene expression. The following gene expressional elements were located in tandem within the plasmid construct: [hTERT core promoter, cytomegalovirus (CMV) minimized promoter, RU5' sequence, an inserted gene, BGH polyA, hTERT enhancer]; this is hereafter referred to as the hT/Cm-R-hT construct. Using various human cancer cell lines and normal cells, the cancer-specific transcription of the green fluorescent protein (GFP) gene was examined by western blotting and fluorescence microscopy. Cancer-specific gene expression was robustly achieved in the hT/Cm-R-hT plasmid in comparison to the other control hT/Cm-driven construct. Notably, the expression level of GFP observed in the hT/Cm-RhT-driven construct was superior to that of the control plasmid with the conventional CMV promoter in HEK293 cells, which are known to possess higher hTERT activity than normal cells. We next examined the availability of hT/Cm-R-hT in detecting the target GFP expressing cancer cells from human peripheral blood mononuclear cells (PBMCs). The hT/Cm-R-hT plasmid successfully induced cancer-specific gene expression; the robust expression of GFP was observed in target HeLa cancer cells, whereas GFP was not visibly expressed in normal PBMCs. The plasmid allowed for the selective visualization of viable HeLa cancer cells in mixed cell cultures containing up to 10000-fold more PBMCs. These findings indicate that the hT/Cm-R-hT expressional system is a valuable tool for detecting viable cancer cells mixed with normal cells. The current system can therefore be applied to the in vitro detection of cancer cells that are disseminated in the blood and other types of body fluid in vivo. Since the current system can also be applied to other types of vectors, including virus vectors, this approach using the hTERT promoter-based construct is expected to become a valuable tool for enhancing cancer specific gene expression.

DOI： 10.3892/or.2017.5710

The assessment of renal cortex and parenchymal volume using automated CT volumetry for predicting renal function after donor nephrectomy 査読

Yosuke Mitsui, Takuya Sadahira, Motoo Araki, Koichiro Wada, Ryuta Tanimoto, Yuichi Ariyoshi, Yasuyuki Kobayashi, Masami Watanabe, Toyohiko Watanabe, Yasutomo Nasu

Clinical and Experimental Nephrology 22 ( 2 ) 1 - 6 2017年7月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：Springer Tokyo

Background: Contrast-enhanced CT is necessary before donor nephrectomy and is usually combined with a Tc-99m-mercapto-acetyltriglycine (MAG3) scan to check split renal function (SRF). However, all transplant programs do not use MAG3 because of its high cost and exposure to radiation. We examined whether CT volumetry of the kidney can be a new tool for evaluating SRF. Methods: Sixty-three patients underwent live donor nephrectomy. Patients without a 1.0 mm slice CT or follow-up for &lt
12 months were excluded leaving 34 patients’ data being analyzed. SRF was measured by MAG3. Split renal volume (SRV) was calculated automatically using volume analyzer software. The correlation between SRF and SRV was examined. The association between the donor’s postoperative estimated glomerular filtration rate (eGFR) and predicted eGFR calculated by MAG3 or CT volumetry was analyzed at 1, 3, and 12 months post nephrectomy. Results: Strong correlations were observed preoperatively in a Bland–Altman plot between SRF measured by MAG3 and either CT cortex or parenchymal volumetry. In addition, eGFR after donation correlated with SRF measured by MAG3 or CT volumetry. The correlation coefficients (R) for eGFR Mag3 split were 0.755, 0.615, and 0.763 at 1, 3 and 12 months, respectively. The corresponding R values for cortex volume split were 0.679, 0.638, and 0.747. Those for parenchymal volume split were 0.806, 0.592, and 0.764. Conclusion: Measuring kidney by CT volumetry is a cost-effective alternative to MAG3 for evaluating SRF and predicting postoperative donor renal function. Both cortex and parenchymal volumetry were similarly effective.

DOI： 10.1007/s10157-017-1454-1

Expression of tumor suppressor REIC/Dkk-3 by a newly improved adenovirus vector with insertion of a hTERT promoter at the 3'-side of the transgene 査読

Endy Widya Putranto, Rie Kinoshita, Masami Watanabe, Takuya Sadahira, Hitoshi Murata, Ken-Ichi Yamamoto, Junichiro Futami, Ken Kataoka, Yusuke Inoue, I. Made Winarsa Ruma, I. Wayan Sumardika, Chen Youyi, Miyoko Kubo, Yoshihiko Sakaguchi, Kenji Saito, Yasutomo Nasu, Hiromi Kumon, Nam-Ho Huh, Masakiyo Sakaguchi

ONCOLOGY LETTERS 14 ( 1 ) 1041 - 1048 2017年7月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：SPANDIDOS PUBL LTD

Reduced expression in immortalized cells (REIC)/Dickkopf-3 (Dkk-3) overexpression, induced using an adenovirus (Ad)-REIC, has been revealed to have a dramatic therapeutic effect on multiple types of cancer. To achieve an improved therapeutic effect from Ad-REIC on cancer, our group previously developed an enhanced gene expression system, the C-TSC cassette [cytomegalovirus (CMV)-RU5' located upstream (C); another promoter unit composed of triple tandem promoters, human telomerase reverse transcriptase (hTERT), simian virus 40 and CMV, located downstream of the cDNA (TSC); plus a polyadenylation (polyA) signal]. When applied to the conventional Ad-REIC, this novel system induced the development of an enhanced product, Ad-C-TSC-REIC, which exhibited a noticeable anticancer effect. However, there were difficulties in terms of Ad-C-TSC-REIC productivity in HEK293 cells, which are a widely used donor cell line for viral production. Productivity of Ad-C-TSC-REIC was significantly reduced compared with the conventional Ad-REIC, as the Ad-C-TSC-REIC had a significantly higher ability to induce apoptotic cell death of not only various types of cancer cell, but also HEK293 cells. The present study aimed to overcome this problem by modifying the C-TSC structure, resulting in an improved candidate: A C-T cassette (C: CMV-RU5' located upstream; T: another promoter unit composed of a single hTERT promoter, located downstream of the cDNA plus a polyA signal), which demonstrated gene expression comparable to that of the C-TSC system. The improved adenovirus REIC/Dkk-3 product with the C-T cassette, named Ad-C-T-REIC, exhibited a higher expression level of REIC/Dkk3, similar to that of Ad-C-TSC-REIC. Notably, the vector mitigated the cell death of donor HEK293 cells, resulting in a higher rate of production of its adenovirus. These results indicated that Ad-C-T-REIC has the potential to be a useful tool for application in cancer gene therapy.

DOI： 10.3892/ol.2017.6201

Distant Bystander Effect of REIC/DKK3 Gene Therapy Through Immune System Stimulation in Thoracic Malignancies. 査読

Suzawa K, Shien K, Peng H, Sakaguchi M, Watanabe M, Hashida S, Maki Y, Yamamoto H, Tomida S, Soh J, Asano H, Tsukuda K, Nasu Y, Kumon H, Miyoshi S, Toyooka S

Anticancer Res 37 ( 1 ) 301 - 307 2017年6月

Canine REIC/Dkk-3 interacts with SGTA and restores androgen receptor signalling in androgen-independent prostate cancer cell lines 査読

Yuiko Kato, Kazuhiko Ochiai, Shota Kawakami, Nobuhiro Nakao, Daigo Azakami, Makoto Bonkobara, Masaki Michishita, Masami Morimatsu, Masami Watanabe, Toshinori Omi

BMC VETERINARY RESEARCH 13 ( 1 ) 170 2017年6月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：BIOMED CENTRAL LTD

Background: The pathological condition of canine prostate cancer resembles that of human androgen-independent prostate cancer. Both canine and human androgen receptor (AR) signalling are inhibited by overexpression of the dimerized co-chaperone small glutamine-rich tetratricopeptide repeat-containing protein a (SGTA), which is considered to cause the development of androgen-independency. Reduced expression in immortalised cells (REIC/Dkk-3) interferes with SGTA dimerization and rescues AR signalling. This study aimed to assess the effects of REIC/Dkk-3 and SGTA interactions on AR signalling in the canine androgen-independent prostate cancer cell line CHP-1.
Results: Mammalian two-hybrid and Halo-tagged pull-down assays showed that canine REIC/Dkk-3 interacted with SGTA and interfered with SGTA dimerization. Additionally, reporter assays revealed that canine REIC/Dkk-3 restored AR signalling in both human and canine androgen-independent prostate cancer cells. Therefore, we confirmed the interaction between canine SGTA and REIC/Dkk-3, as well as their role in AR signalling.
Conclusions: Our results suggest that this interaction might contribute to the development of a novel strategy for androgen-independent prostate cancer treatment. Moreover, we established the canine androgen-independent prostate cancer model as a suitable animal model for the study of this type of treatment-refractory human cancer.

DOI： 10.1186/s12917-017-1094-4

RCAN-11R peptide provides immunosuppression for fully mismatched islet allografts in mice 査読

Hirofumi Noguchi, Koji Sugimoto, Chika Miyagi-Shiohira, Yoshiki Nakashima, Naoya Kobayashi, Issei Saitoh, Masami Watanabe, Yasufumi Noguchi

SCIENTIFIC REPORTS 7 ( 1 ) 3043 - 3043 2017年6月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：NATURE PUBLISHING GROUP

Calcineurin inhibitors have been used for transplant therapy. However, the inhibition of calcineurin outside the immune system has a number of side effects. We previously developed a cell-permeable inhibitor of NFAT (nuclear factor of activated T cells) using the polyarginine peptide delivery system. This peptide (11R-VIVIT) selectively interferes with calcineurin-NFAT interaction without affecting the activity of calcineurin phosphatase and provides immunosuppression for fully mismatched islet allografts in mice. However, our recent study showed that 11R-VIVIT affected cell viability in vitro when it was used at higher concentration because of the VIVIT sequence. The aim of this study is to develop a safer NFAT inhibitor (RCAN-11R) that does not affect cell viability, and which is less toxic than calcineurin inhibitors. The minimal sequence of the protein family of regulators of calcineurin (RCAN) that is responsible for the inhibition of calcineurin-NFAT signaling was recently characterized. The peptide could selectively interfere with the calcineurin-NFAT interaction without affecting the activity of calcineurin phosphatase, similar to 11R-VIVIT. RCAN-11R did not affect cell viability when it was used at a higher concentration than the toxic concentration of 11R-VIVIT. RCAN-11R could therefore be useful as a therapeutic agent that is less toxic than current drugs or 11R-VIVIT.

DOI： 10.1038/s41598-017-02934-3

The canine prostate cancer cell line CHP-1 shows over-expression of the co-chaperone small glutamine-rich tetratricopeptide repeat-containing protein α. 査読

Azakami D, Nakahira R, Kato Y, Michishita M, Kobayashi M, Onozawa E, Bonkobara M, Kobayashi M, Takahashi K, Watanabe M, Ishioka K, Sako T, Ochiai K, Omi T

Vet Comp Oncol. 15 ( 2 ) 557 - 562 2017年6月

Clinical pharmacokinetics of oral levofloxacin and sitafloxacin in epididymal tissue 査読

Takuya Sadahira, Koichiro Wada, Kazuro Ikawa, Norifumi Morikawa, Hiroaki Kurahashi, Takashi Yoshioka, Yuichi Ariyoshi, Yasuyuki Kobayashi, Motoo Araki, Ayano Ishii, Masami Watanabe, Shinya Uehara, Toyohiko Watanabe, Yasutomo Nasu

JOURNAL OF INFECTION AND CHEMOTHERAPY 23 ( 4 ) 214 - 217 2017年4月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：ELSEVIER SCIENCE BV

Objectives: This study aimed to investigate the penetration of fluoroquinolones into human epididymal tissue.
Methods: The penetration of levofloxacin (LVFX) 500 mg or sitafloxacin (STFX) 100 mg into epididymal tissue was examined. Patients with prostate cancer who were referred for orchiectomy were included. LVFX 500 mg (n = 9) or STFX 100 mg (n = 9) was administered orally 1 h before orchiectomy, and 0.5 g of epididymal tissue and blood samples were collected simultaneously during surgery. Drug concentrations were measured by high-performance liquid chromatography, and patient characteristics and adverse events were analyzed.
Results: The mean ratio of the epididymal concentration to the serum concentration was 1.48 +/- 0.45 for LVFX and 1.54 +/- 0.81 for STFX. For LVFX, the simulated curves estimated the following: maximum concentrations (Cmax) of 8.84 mu g/ml in serum and 14.1 mu g/g in epididymal tissue and area under the concentration-time curve for 24 h (AUC(24)) of 68.5 mu g h/ml in serum and 108.9 mu g h/g in epididymal tissue. For STFX, the Cmax was 1.22 mu g/ml in serum and 1.66 mu g/g in epididymal tissue, and the AUC24 was 9.58 mu g h/ml in serum and 13.1 mu g h/g in epididymal tissue. Neither treatment-related adverse events nor postoperative urogenital infections were observed.
Conclusions: The results of this study suggest that oral administration of LVFX 500 mg or STFX 100 mg achieves effective epididymal concentrations for treatment of epididymitis. (C) 2016 Japanese Society of Chemotherapy and The Japanese Association for Infectious Diseases. Published by Elsevier Ltd. All rights reserved.

DOI： 10.1016/j.jiac.2016.12.010

The Downregulation of the Expression of CD147 by Tumor Suppressor REIC/Dkk-3, and Its Implication in Human Prostate Cancer Cell Growth Inhibition 査読

Akihiro Mori, Masami Watanabe, Takuya Sadahira, Yasuyuki Kobayashi, Yuichi Ariyoshi, Hideo Ueki, Koichiro Wada, Kazuhiko Ochiai, Shun-Ai Li, Yasutomo Nasu

ACTA MEDICA OKAYAMA 71 ( 2 ) 135 - 142 2017年4月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：OKAYAMA UNIV MED SCHOOL

The cluster of differentiation 147 (CD147), also known as EMMPRIN, is a key molecule that promotes cancer progression. We previously developed an adenoviral vector encoding a tumor suppressor REIC/Dkk-3 gene (Ad-REIC) for cancer gene therapy. The therapeutic effects are based on suppressing the growth of cancer cells, but, the underlying molecular mechanism has not been fully clarified. To elucidate this mechanism, we investigated the effects of Ad-REIC on the expression of CD147 in LNCaP prostate cancer cells. Western blotting revealed that the expression of CD147 was significantly suppressed by Ad-REIC. Ad-REIC also suppressed the cell growth of LNCaP cells. Since other researchers have demonstrated that phosphorylated mitogen-activated protein kinases (MAPKs) and c-Myc protein positively regulate the expression of CD147, we investigated the correlation between the CD147 level and the activation of MAPK and c-Myc expression. Unexpectedly, no positive correlation was observed between CD147 and its possible regulators, suggesting that another signaling pathway was involved in the downregulation of CD147. This is the first study to show the downregulation of CD147 by Ad-REIC in prostate cancer cells. At least some of the therapeutic effects of Ad-REIC may be due to the downregulation of the cancer-progression factor, CD147.

DOI： 10.18926/AMO/54982

Synergistic effects of the immune checkpoint inhibitor CTLA-4 combined with the growth inhibitor lycorine in a mouse model of renal cell carcinoma 査読

Xiezhao Li, Peng Xu, Chongshan Wang, Naijin Xu, Abai Xu, Yawen Xu, Takuya Sadahira, Motoo Araki, Koichiro Wada, Eiji Matsuura, Masami Watanabe, Junxia Zheng, Pinghua Sun, Peng Huang, Yasutomo Nasu, Chunxiao Liu

ONCOTARGET 8 ( 13 ) 21177 - 21186 2017年3月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：IMPACT JOURNALS LLC

Renal cell carcinoma (RCC) management has undergone a major transformation over the past decade; immune checkpoint inhibitors are currently undergoing clinical trials and show promising results. However, the effectiveness of immune checkpoint inhibitors in patients with metastatic RCC (mRCC) is still limited. Lycorine, an alkaloid extracted from plants of the Amaryllidaceae family, is touted as a potential anti-cancer drug because of its demonstrative growth inhibition capacity (induction of cell cycle arrest and inhibition of vasculogenic mimicry formation). Moreover, T cell checkpoint blockade therapy with antibodies targeting cytotoxic T-lymphocyte associated protein 4 (CTLA-4) has improved outcomes in cancer patients. However, the anti-tumor efficacy of combined lycorine and anti-CTLA-4 therapy remains unknown. Thus, we investigated a combination therapy of lycorine hydrochloride and anti-CTLA-4 using a murine RCC model. As a means of in vitro confirmation, we found that lycorine hydrochloride inhibited the viability of various RCC cell lines. Furthermore, luciferase-expressing Renca cells were implanted in the left kidney and the lung of BALB/c mice to develop a RCC metastatic mouse model. Lycorine hydrochloride and anti-CTLA-4 synergistically decreased tumor weight, lung metastasis, and luciferin-staining in tumor images. Importantly, the observed anti-tumor effects of this combination were dependent on significantly suppressing regulatory T cells while upregulating effector T cells; a decrease in regulatory T cells by 31.43% but an increase in effector T cells by 31.59% were observed in the combination group compared with those in the control group). We suggest that a combination of lycorine hydrochloride and anti-CTLA-4 is a viable therapeutic option for RCC patients.

DOI： 10.18632/oncotarget.15505

Programmed death ligand 1 expression in bladder rhabdomyosarcoma and its association with clinicopathological features 査読

Peng Zhou, Qiongren Wang, Chongshan Wang, Xiezhao Li, Wei Du, Chunxiao Liu, Masami Watanabe, Peng Huang, Abai Xu

INTERNATIONAL JOURNAL OF CLINICAL AND EXPERIMENTAL PATHOLOGY 10 ( 10 ) 10565 - 10570 2017年

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：E-CENTURY PUBLISHING CORP

The aim of this study was to detect PD-L1 expression in bladder rhabdomyosarcoma and its association with clinicopathological features and patient prognosis. PD-L1 expression was detected in paraffin-embedded sections obtained from 34 patients with bladder rhabdomyosarcoma via immunohistochemistry. Immunohistochemistry results were statistically analyzed to determine their association with patient clinicopathological features and survival outcomes. PD-L1-positive staining was observed in 47.1% (16/34) of patients. Metastatic tumor cells in the lymph nodes of two patients were positive for PD-L1 expression. PD-L1 expression was significantly different with regard to muscularis invasion, but the expression did not affect patient survival outcomes. We confirmed PD-L1 expression in bladder rhabdomyosarcoma, suggesting that PD-1/PD-L1 inhibitors are potential therapeutic agents for patients with bladder rhabdomyosarcoma.

A proteome analysis of pig pancreatic islets and exocrine tissue by liquid chromatography with tandem mass spectrometry 査読

Yoshiki Nakashima, Chika Miyagi-Shiohira, Naoya Kobayashi, Issei Saitoh, Masami Watanabe, Hirofumi Noguchi

ISLETS 9 ( 6 ) 159 - 176 2017年

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：TAYLOR & FRANCIS INC

Liquid chromatography with tandem mass spectrometry (LC-MS/MS) is a proteome analysis method, and the shotgun analysis by LC-MS/MS comprehensively identifies proteins from tissues and cells with high resolving power. In this study, we analyzed the protein expression in pancreatic tissue by LC-MS/MS. Islets isolated from porcine pancreata (purity 95%) and exocrine tissue (purity 99%) were used in this study. LC-MS/MS showed that 13 proteins were expressed in pancreatic islets only (Group I), 43 proteins were expressed in both islets and exocrine tissue (Group I&E), and 102 proteins were expressed in exocrine tissue only (Group E). Proteins involved in islet differentiation and cell proliferation were identified in Group I (e.g. CLUS, CMGA, MIF). In addition, various functional proteins (e.g. SCG2, TBA1A) were identified in islet by using the new method of principal component analysis (PCA)'. However, the function of such proteins on islets remains unclear. EPCAM was identified in Group E. Group E was found to include proteins involved in clinical inflammatory diseases such as pancreatitis (e.g. CBPA1, CGL, CYTB, ISK1 and PA21B). Many of these identified proteins were reported less frequently in previous studies, and HS71B, NEC2, PRAF3 and SCG1 were newly detected in Group I while CPNS1, DPEP1, GANAB, GDIB, GGT1, HSPB1, ICTL, VILI, MUTA, NDKB, PTGR1, UCHL3, VAPB and VINC were newly detected in Group E. These results show that comprehensive expression analysis of proteins by LC-MS/MS is useful as a method to investigate new factors constructing cellular component, biological process, and molecular function.

DOI： 10.1080/19382014.2017.1389826

Efficacy and safety of 3 day versus 7 day cefditoren pivoxil regimens for acute uncomplicated cystitis: Multicentre, randomized, open-label trial 査読

Takuya Sadahira, Okayama Urological Research Group (OURG), Koichiro Wada, Motoo Araki, Ayano Ishii, Atsushi Takamoto, Yasuyuki Kobayashi, Masami Watanabe, Toyohiko Watanabe, Yasutomo Nasu, Hiromi Kumon, Teruaki Akaeda, Nobuyuki Akazawa, Naoki Akebi, Daiji Araki, Tohru Araki, Ryoji Arata, Yuichi Ariyoshi, Eiichi Ando, Nobuyoshi Ando, Kazushi Ishii, Tsutomu Ishikawa, Noritaka Ishito, Takaharu Ichikawa, Takaaki Inoue, Miyabi Inoue, Yosuke Inoue, Shin Irie, Takehiro Iwata, Tatsuya Uesugi, Shinya Uehara, Katsutoshi Uematsu, Satoshi Uno, Kohei Edamura, Shin Ebara, Yuko Oiwa, Tadashi Oeda, Teruhisa Ohashi, Yozo Ohashi, Hideo Ozawa, Junzo Ochi, Noriaki Ono, Seiji Kai, Haruki Kaku, Satoshi Katayama, Yasuhiro Katayama, Tetuzo Kaneshige, Keiichiro Kawauchi, Masashi Kawaguchi, Tatsushi Kawada, Taiki Kanbara, Mikio Kishi, Ryo Kishimoto, Nobuyuki Kusaka, Norihiro Kusumi, Kimito Kunitomi, Risa Kubota, Hiroaki Kurahashi, Hironori Kojima, Tomoko Kobayashi, Makoto Kobuke, Katsuyoshi Kondo, Takashi Saika, Kazuma Sakaeda, Koji Sakuramoto, Shinichi Sako, Tomoko Sako, Taketo Sasaoka, Katsumi Sasaki, Yoshimasa Jo, Morito Sugimoto, Yuko Seno, Akira Takao, Kosuke Takamura, Kousuke Takamura, Hitoshi Takamoto, Katsuji Takeda, Tadasu Takenaka, Daisuke Tanaka, Ryuta Tanimoto, Masaya Tsugawa, Tomoyasu Tsushima, Hiromu Tsuboi, Syunji Tojo, Kenji Tokinaga, Yusuke Tominaga, Keisuke Doi, Atsushi Nagai, Hirochika Nakajima, Tetsuya Nakada, Hirokazu Nakatsuka, Aya Nakamura, Yasuki Nakayama, Yoshitsugu Nasu, Daisuke Nishikawa, Jun Nishiguchi, Yoshio Nishitani, Shingo Nishimura, Motoichi Nishimura, Yasuhiro Nishiyama, Hajime Nibuno, Kunihiro Nozaki, Hiroyuki Nose, Gaku Noda, Hideaki Hashimoto, Kazuhiro Hata, Toshihide Hayashi, Nobuki Hayashi, Syunji Hayata, Ryoei Hara, Takeshi Hirata, Tomohiro Fujii, Kei Fujio, Ryuji Fujita, Kensuke Bekku, Takanori Maehara, Yoshio Maki, Yuko Matsumoto, Daisuke Manabe, Yusuke Mayumi, Yuki Maruyama, Yosuke Mitsui, Sadayuki Miyaji, Syuhei Munemasa, Wataru Murao, Takanori Murakami, Tadashi Murata, Akihiro Mori, Akira Morita, Koichi Monden, Tomoya Yamasaki, Masahiro Yamashita, Daiduke Yamada, Toyoko Yamato, Toru Yamane, Tomoaki Yamanoi, Yasuo Yamamoto, Kazuaki Yukari, Teruhiko Yokoyama, Takashi Yoshioka, Yuichi Watanabe

Journal of Antimicrobial Chemotherapy 72 ( 2 ) 529 - 534 2017年

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：Oxford University Press

Background: Fluoroquinolone-non-susceptible Escherichia coli isolated from patients with acute uncomplicated cystitis are a matter of increasing concern. Cefditoren pivoxil is an oral, β-lactamase-stable, extended-spectrum cephalosporin that is effective against fluoroquinolone-non-susceptible bacteria. Objectives: To evaluate the clinical and microbiological efficacies of cefditoren pivoxil against acute uncomplicated cystitis and to determine the optimal duration of cefditoren pivoxil treatment. Methods: We compared 3 and 7 day regimens of cefditoren pivoxil in a multicentre, randomized, open-label study. Results: A total of 104 female patients with acute uncomplicated cystitis were enrolled and randomized into 3 day (n=51) or 7 day (n=53) treatment groups. At first visit, 94 bacterial strains were isolated from the 104 participants of which 81.7% (85/104) were E. coli. Clinical and microbiological efficacies were evaluated 5-9 days following administration of the final dose of cefditoren pivoxil. The clinical efficacies of the 3 and 7 day groups were 90.9% (40/44) and 93.2% (41/44), respectively (P=1.000). The microbiological efficacies of the 3 and 7 day groups were 82.5% (33/40) and 90.2% (37/41), respectively (P=0.349). There were no adverse events due to cefditoren pivoxil treatment, with the exception of a mild allergic reaction in one patient, after which the cefditoren pivoxil was exchanged for another antimicrobial. Conclusions: Cefditoren pivoxil is safe and effective for uncomplicated cystitis, with no significant differences in clinical and microbiological efficacies between 3 and 7 day regimens.

DOI： 10.1093/jac/dkw424

Dynamin2 GTPase contributes to invadopodia formation in invasive bladder cancer cells 査読

Yubai Zhang, Maya Nolan, Hiroshi Yamada, Masami Watanabe, Yasutomo Nasu, Kohji Takei, Tetsuya Takeda

BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS 480 ( 3 ) 409 - 414 2016年11月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：ACADEMIC PRESS INC ELSEVIER SCIENCE

Cancer cell invasion is mediated by actin-based membrane protrusions termed invadopodia. Invadopodia consist of "core" F-actin bundles associated with adhesive and proteolytic machineries promoting cell invasion by degrading extracellular matrix (ECM). Formation of the F-actin core in invadopodia is regulated by various actin-binding proteins including Arp2/3 complex and cortactin. Dynamin GTPase localizes to the invadopodia and is implicated in cancer cell invasion, but its precise role at the invadopodia remained elusive.
In this study, we examined the roles of dynamin at the invadopodia of bladder cancer cells. Although all three dynamin isoforms (dynamin1, 2 and 3) are expressed in human bladder cancer cell line T24, only dynamin2 localizes to the invadopodia. Inhibition of dynamin2 function, using either RNA interference (RNAi) or the dynamin specific inhibitor Dynasore, caused defects in invadopodia formation and suppressed invasive activity of 124 bladder cancer cells. Structure-function analysis using dynamin2 deletion fragments identified the proline/arginine-rich domain (PRD) of dynamin2 as indispensable for invadopodia formation and invasiveness of 124 cells. Thus, dynamin2 contributes to bladder cancer invasion by controlling invadopodia formation in bladder cancer cells and may prove a valuable therapeutic target. (C) 2016 Elsevier Inc. All rights reserved.

DOI： 10.1016/j.bbrc.2016.10.063

Adenovirus vector carrying REICIDKK-3 gene: neoadjuvant intraprostatic injection for high-risk localized prostate cancer undergoing radical prostatectomy 査読

H. Kumon, Y. Ariyoshi, K. Sasaki, T. Sadahira, M. Araki, S. Ebara, H. Yanai, M. Watanabe, Y. Nasu

CANCER GENE THERAPY 23 ( 11 ) 400 - 409 2016年11月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：NATURE PUBLISHING GROUP

As the First-In-Human study of in situ gene therapy using an adenovirus vector carrying the human REIC (reduced expression in immortalized cell)/Dkk-3 gene (Ad-REIC), we conducted neoadjuvant intraprostatic injections in patients with high-risk localized prostate cancer undergoing radical prostatectomy (RP). Patients with recurrence probability of 35% or more within 5 years following RP, as calculated by Kaftan's nomogram, were enrolled. Patients received two ultrasound-guided intratumoral injections at 2-week intervals, followed by RP 6 weeks after the second injection. After confirming the safety of the therapeutic interventions with initially planned three escalating doses of 1.0 x 10(10), 1.0 x 10(11) and 1.0 x 10(12) viral particles (vp) in 1.0-1.2 ml (n=3, 3 and 6), an additional higher dose of 3.0 x 10(12) vp in 3.6 ml (n=6) Was further studied. All four DLs including the additional dose level-4 (DL-4) were feasible with no adverse events, except for grade 1 or 2 transient fever. Laboratory toxicities were grade 1 or 2 elevated aspartate transaminase/alanine transaminase (n=4). Regarding antitumor activities, cytopathic effects (tumor degeneration With cytolysis and pyknosis) and remarkable tumor-infiltrating lymphocytes in the targeted tumor areas were detected in a clear dose-dependent manner. Consequently, biochemical recurrence-free survival in DL-4 was significantly more favorable than in patient groups DL-1+2+2:

DOI： 10.1038/cgt.2016.53

Comparison of Purification Solutions With Different Osmolality for Porcine Islet Purification. 査読

Miyagi-Shiohira C, Kobayashi N, Saitoh I, Watanabe M, Noguchi Y, Matsushita M, Noguchi H

Cell Med 9 ( 1月2日 ) 53 - 59 2016年9月

The Evaluation of Islet Purification Methods That Use Large Bottles to Create a Continuous Density Gradient. 査読

Miyagi-Shiohira C, Kobayashi N, Saitoh I, Watanabe M, Noguchi Y, Matsushita M, Noguchi H

Cell Med 9 ( 1月2日 ) 45 - 51 2016年9月

Evaluation of Serum-Free, Xeno-Free Cryopreservation Solutions for Human Adipose-Derived Mesenchymal Stem Cells. 査読

Miyagi-Shiohira C, Kobayashi N, Saitoh I, Watanabe M, Noguchi Y, Matsushita M, Noguchi H

Cell Med 9 ( 1月2日 ) 15 - 20 2016年9月

A super gene expression system enhances the anti-glioma effects of adenovirus-mediated REIC/Dkk-3 gene therapy 査読

Tetsuo Oka, Kazuhiko Kurozumi, Yosuke Shimazu, Tomotsugu Ichikawa, Joji Ishida, Yoshihiro Otani, Toshihiko Shimizu, Yusuke Tomita, Masakiyo Sakaguchi, Masami Watanabe, Yasutomo Nasu, Hiromi Kumon, Isao Date

SCIENTIFIC REPORTS 6 2016年9月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：NATURE PUBLISHING GROUP

Reduced expression in immortalized cells/Dickkopf-3 (REIC/Dkk-3) is a tumor suppressor and therapeutic gene in many human cancers. Recently, an adenovirus REIC vector with the super gene expression system (Ad-SGE-REIC) was developed to increase REIC/Dkk-3 expression and enhance therapeutic effects compared with the conventional adenoviral vector (Ad-CAG-REIC). In this study, we investigated the in vitro and in vivo effects of Ad-SGE-REIC on malignant glioma. In U87 Delta EGFR and GL261 glioma cells, western blotting confirmed that robust upregulation of REIC/Dkk-3 expression occurred in Ad-SGE-REIC-transduced cells, most notably after transduction at a multiplicity of infection of 10. Cytotoxicity assays showed that Ad-SGE-REIC resulted in a time-dependent and significant reduction in the number of malignant glioma cells attaching to the bottom of culture wells. Xenograft and syngeneic mouse intracranial glioma models treated with Ad-SGE-REIC had significantly longer survival than those treated with the control vector Ad-LacZ or with Ad-CAG-REIC. This study demonstrated the anti-glioma effect of Ad-SGE-REIC, which may represent a promising strategy for the treatment of malignant glioma.

DOI： 10.1038/srep33319

MCAM, as a novel receptor for S100A8/A9, mediates progression of malignant melanoma through prominent activation of NF-κB and ROS formation upon ligand binding. 査読

Ruma IM, Putranto EW, Kondo E, Murata H, Watanabe M, Huang P, Kinoshita R, Futami J, Inoue Y, Yamauchi A, Sumardika IW, Youyi C, Yamamoto K, Nasu Y, Nishibori M, Hibino T, Sakaguchi M

Clin Exp Metastasis. 33 ( 6 ) 609 - 627 2016年8月

The Efficacy of Rituximab in High-risk Renal Transplant Recipients 査読

Motoo Araki, Koichiro Wada, Yosuke Mitsui, Risa Kubota, Takashi Yoshioka, Yuichi Ariyoshi, Yasuyuki Kobayashi, Masashi Kitagawa, Katsuyuki Tanabe, Hiroshi Sugiyama, Jun Wada, Masami Watanabe, Toyohiko Watanabe, Katsuyuki Hotta, Yasutomo Nasu

ACTA MEDICA OKAYAMA 70 ( 4 ) 295 - 297 2016年8月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：OKAYAMA UNIV MED SCHOOL

Although graft survival following renal transplantation (RTx) has improved, outcomes following highrisk RTx are variable. Preexisting antibodies, including donor-specific antibodies (DSA), play an important role in graft dysfunction and survival. We have designed a study to investigate the safety and efficacy of anti-CD20 monoclonal antibodies (rituximab) in high-risk RTx recipients. Major eligibility criteria include: 1) major and minor ABO blood group mismatch, 2) positive DSA. Thirty-five patients will receive 200 mg/body of rituximab. The primary endpoint is the incidence of B cell depletion. This study will clarify whether rituximab is efficacious in improving graft survival in high-risk RTx recipients.

DOI： 10.18926/AMO/54507

Novel REIC/Dkk-3-encoding adenoviral vector as a promising therapeutic agent for pancreatic cancer 査読

H. Sawahara, H. Shiraha, D. Uchida, H. Kato, T. Nagahara, M. Iwamuro, J. Kataoka, S. Horiguchi, M. Watanabe, M. Sakaguchi, A. Takaki, K. Nouso, Y. Nasu, H. Kumon, H. Okada

CANCER GENE THERAPY 23 ( 8 ) 278 - 283 2016年8月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：NATURE PUBLISHING GROUP

Reduced expression in immortalized cells (REIC)/dickkopf-3 (Dkk-3), a tumor suppressor gene, is downregulated in various cancers. We previously reported the tumor-inhibitory effects of the REIC/Dkk-3 gene, delivered by a conventional adenoviral vector (Ad-CAG-REIC) in pancreatic cancer. Here, we developed an Ad-REIC vector with a novel gene expression system, termed the super gene expression (SGE) system, and assessed its therapeutic effects relative to those of Ad-CAG-REIC in pantreatic cancer cells. Human pancreatic cancer cell lines ASPC1 and MIAPaCa2 were used. REIC/Dkk-3 expression was assessed by western blot analysis. Relative cell viability and apoptotic effects were examined in vitro. The anti-tumor effects of Ad-REIC treatment were assessed in the mouse xenograft model. Compared with Ad-CAG-REIC, Ad-SGE-REIC elicited a significant increase in REIC protein expression in the cells studied, Relative to Ad-CAG-REIC, Ad-SGE-REIC reduced cell viability and induced apoptosis in the ASPC1 and MIAPaCa2 cell lines in vitro, and achieved superior tumor growth inhibition in the mouse xenograft model. Compared with conventional Ad-REIC agents, Ad-SGE-REIC provided enhanced inhibitory effects against tumor growth. Our results indicate that Ad-SGE-REIC is an innovative therapeutic tool for pancreatic cancer.

DOI： 10.1038/cgt.2016.31

A Phase II Clinical Trial Evaluating the Preventive Effectiveness of Lactobacillus Vaginal Suppositories in Patients with Recurrent Cystitis 査読

Koichiro Wada, Shinya Uehara, Ayano Ishii, Takuya Sadahira, Masumi Yamamoto, Ritsuko Mitsuhata, Atsushi Takamoto, Motoo Araki, Yasuyuki Kobayashi, Masami Watanabe, Toyohiko Watanabe, Katsuyuki Hotta, Yasutomo Nasu

ACTA MEDICA OKAYAMA 70 ( 4 ) 299 - 302 2016年8月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：OKAYAMA UNIV MED SCHOOL

Urinary tract infections (UTIs) are the most common bacterial infections in women, and many patients experience frequent recurrence. The aim of this report is to introduce an on-going prospective phase II clinical trial performed to evaluate the preventive effectiveness of Lactobacillus vaginal suppositories for prevention of recurrent cystitis. Patients enrolled in this study are administered vaginal suppositories containing the GAI 98322 strain of Lactobacillus crispatus every 2 days or 3 times a week for one year. The primary endpoint is recurrence of cystitis and the secondary endpoints are adverse events. Recruitment began in December 2013 and target sample size is 20 participants.

DOI： 10.18926/AMO/54508

Study about the Efficacy of Metformin to Immune Function in Cancer Patients. 査読

Watanabe M, Yamamoto H, Eikawa S, Shien K, Shien T, Soh J, Hotta K, Wada J, Hinotsu S, Fujiwara T, Kiura K, Doihara H, Miyoshi S, Udono H, Toyooka S

Acta medica Okayama 70 ( 4 ) 327 - 30 2016年8月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：OKAYAMA UNIV MED SCHOOL

A study to evaluate the effect of metformin on the immune system was commenced in July 2014. Metformin is one of the most commonly prescribed drugs for type 2 diabetes, and previous studies have reported that metformin has an anti-tumor effect. The aim of this study is to evaluate the efficacy of metformin on the immune system in human cancer patients in vivo. The primary outcome parameter will be the rate change in the population of CD8＋ T cells, which produce multiple cytokines.

DOI： 10.18926/AMO/54514

Synergistic anti-pancreatic cancer immunological effects by treatment with reduced expression in immortalized cells/dickkopf-3 protein and peripheral blood mononuclear cells 査読

Daisuke Uchida, Hidenori Shiraha, Hironari Kato, Hiroaki Sawahara, Teruya Nagahara, Masaya Iwamuro, Junro Kataoka, Shigeru Horiguchi, Masami Watanabe, Akinobu Takaki, Kazuhiro Nouso, Yasutomo Nasu, Hiromi Kumon, Kazuhide Yamamoto

JOURNAL OF GASTROENTEROLOGY AND HEPATOLOGY 31 ( 6 ) 1154 - 1159 2016年6月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：WILEY-BLACKWELL

Background and AimReduced expression in immortalized cells/dickkopf-3 (REIC/DKK3) is a reported tumor suppressor gene and has potential to become an innovative therapy for various cancers. We examined the antitumor immunological effects of human REIC/DKK3 protein against pancreatic cancer.
MethodsActivation of extracellular signal-regulated kinases 1 and 2, mammalian target of rapamycin, and signal transducer and activator of transcription 3 by REIC/DKK3 protein was assessed in human peripheral blood mononuclear cells using immunoblotting. Pancreatic cancer cell lines (AsPC-1 and MIA Paca-2) were cocultured with peripheral blood mononuclear cells, and the anticancer effects of REIC/DKK3 protein were assessed using the methyl thiazole tetrazolium, cytotoxicity, and enzyme-linked immunospot assays. The antitumor immunological effects of the combined treatment with REIC/DKK3 protein and peripheral blood mononuclear cells were also assessed in a pancreatic cancer model using non-obese diabetic/severe combined immunodeficiency mice.
ResultsThe REIC/DKK3 protein activated extracellular signal-regulated kinases 1 and 2, mammalian target of rapamycin, and signal transducer and activator of transcription 3 in peripheral blood mononuclear cells. REIC/DKK3 protein inhibited in vitro cancer cell viability and enhanced cytotoxicity when incubated with peripheral blood mononuclear cells. REIC/DKK3 protein induced significant production of interferon gamma from lymphocytes incubated with pancreatic cancer cells, indicating that CD8+ T cells were activated in the peripheral blood mononuclear cells when cocultured with AsPC-1 and MIA Paca-2 in the presence of REIC/DKK3 protein. Combined treatment with REIC/DKK3 protein and peripheral blood mononuclear cells produced in vivo anticancer immunostimulatory effects on pancreatic cancer cells.
ConclusionsThe REIC/DKK3 protein and peripheral blood mononuclear cells synergistically enhanced anticancer immunological effects against pancreatic cancer cells. The observed immunomodulatory effect of combined treatment likely occurs in adenovirus-mediated REIC/DKK3 gene therapy and provides important clues to the therapeutic mechanisms involving immune cells.

DOI： 10.1111/jgh.13259

[Gene therapy]. 査読

Watanabe M, Sadahira T, Nasu Y

Nihon rinsho. Japanese journal of clinical medicine 74 Suppl 3 221 - 225 2016年5月

The induction of antigen-specific CTL by in situ Ad-REIC gene therapy 査読

Y. Ariyoshi, M. Watanabe, S. Eikawa, C. Yamazaki, T. Sadahira, T. Hirata, M. Araki, S. Ebara, Y. Nasu, H. Udono, H. Kumon

GENE THERAPY 23 ( 5 ) 408 - 414 2016年5月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：NATURE PUBLISHING GROUP

An adenovirus vector carrying the human Reduced Expression in Immortalized Cell (REIC)/Dkk-3 gene (Ad-REIC) mediates simultaneous induction of cancer-selective apoptosis and augmentation of anticancer immunity. In our preclinical and clinical studies, in situ Ad-REIC gene therapy showed remarkable direct and indirect antitumor effects to realize therapeutic cancer vaccines. We herein aimed to confirm the induction of tumor-associated antigen-specific cytotoxic T lymphocytes (CTLs) by Ad-REIC. Using an ovalbumin (OVA), a tumor-associated antigen, expressing E.G7 tumor-bearing mouse model, we investigated the induction and expansion of OVA-specific CTLs responsible for indirect, systemic effects of Ad-REIC. The intratumoral administration of Ad-REIC mediated clear antitumor effects with the accumulation of OVA-specific CTLs in the tumor tissues and spleen. The CD86-positive dendritic cells (DCs) were upregulated in the tumor draining lymph nodes of Ad-REIC-treated mice. In a dual tumor-bearing mouse model in the left and right back, Ad-REIC injection in one side significantly suppressed the tumor growth on both sides and significant infiltration of OVA-specific CTLs into non-injected tumor was also detected. Consequently, in situ Ad-REIC gene therapy is expected to realize a new-generation cancer vaccine via anticancer immune activation with DC and tumor antigen-specific CTL expansion.

DOI： 10.1038/gt.2016.7

Promising Therapeutic Efficacy of a Novel REIC/Dkk-3 Expressing Adenoviral Vector for Hepatocellular Carcinoma 査読

Hiroaki Sawahara, Shiraha Hidenori, Uchida Daisuke, Sakaguchi Masakiyo, Watanabe Masami, Nasu Yasutomo, Kumon Hiromi, Okada Hiroyuki

GASTROENTEROLOGY 150 ( 4 ) S1153 2016年4月

Development of a Novel REIC/DKK-3-Encoding Adenoviral Agent: Its Robust and Promising Therapeutic Effects in Pancreatic Cancer 査読

Uchida Daisuke, Shiraha Hidenori, Kato Hironari, Hiroaki Sawahara, Sakaguchi Masakiyo, Watanabe Masami, Nasu Yasutomo, Kumon Hiromi, Okada Hiroyuki

GASTROENTEROLOGY 150 ( 4 ) S294 - S294 2016年4月

Tumor suppressor REIC/DKK-3 and co-chaperone SGTA: Their interaction and roles in the androgen sensitivity 査読

Kazuhiko Ochiai, Masami Morimatsu, Yuiko Kato, Toshina Ishiguro-Oonuma, Chihiro Udagawa, Oumaporn Rungsuriyawiboon, Daigo Azakami, Masaki Michishita, Yuichi Ariyoshi, Hideo Ueki, Yasutomo Nasu, Hiromi Kumon, Masami Watanabe, Toshinori Omi

ONCOTARGET 7 ( 3 ) 3273 - 3286 2016年1月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：IMPACT JOURNALS LLC

REIC/DKK-3 is a tumor suppressor, however, its intracellular physiological functions and interacting molecules have not been fully clarified. Using yeast two-hybrid screening, we found that small glutamine-rich tetratricopeptide repeat-containing protein a (SGTA), known as a negative modulator of cytoplasmic androgen receptor (AR) signaling, is a novel interacting partner of REIC/DKK-3. Mammalian two-hybrid and pull-down assay results indicated that the SGTA-REIC/DKK-3 interaction involved the N-terminal regions of both REIC/DKK-3 and SGTA and that REIC/DKK-3 interfered with the dimerization of SGTA, which is a component of the AR complex and a suppressor of dynein motor-dependent AR transport and signaling. A reporter assay in human prostate cancer cells that displayed suppressed AR signaling by SGTA showed recovery of AR signaling by REIC/DKK-3 expression. Considering these results and our previous data that REIC/DKK-3 interacts with the dynein light chain TCTEX-1, we propose that the REIC/DKK-3 protein interferes with SGTA dimerization, promotes dynein-dependent AR transport and then upregulates AR signaling.

DOI： 10.18632/oncotarget.6488

Feasibility of Neoadjuvant Ad-REIC Gene Therapy in Patients with High-Risk Localized Prostate Cancer Undergoing Radical Prostatectomy 査読

Hiromi Kumon, Katsumi Sasaki, Yuichi Ariyoshi, Takuya Sadahira, Motoo Araki, Shin Ebara, Hiroyuki Yanai, Masami Watanabe, Yasutomo Nasu

CTS-CLINICAL AND TRANSLATIONAL SCIENCE 8 ( 6 ) 837 - 840 2015年12月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：WILEY-BLACKWELL

In a phase I/IIa study of in situ gene therapy using an adenovirus vector carrying the human REIC/Dkk-3 gene (Ad-REIC), we assessed the inhibitory effects of cancer recurrence after radical prostatectomy (RP), in patients with high risk localized prostate cancer (PCa). After completing the therapeutic interventions with initially planned three escalating doses of 1.0 x 10(10), 1.0 x 10(11), and 1.0 x 10(12) viral particles (VP) in 1.0-1.2 mL (n = 3, 3, and 6), an additional higher dose of 3.0 x 10(12) VP in 3.6 mL (n = 6) was further studied. Patients with recurrence probability of 35% or more within 5 years after RP as calculated by Kattan's nomogram, were enrolled. They received two ultrasound-guided intratumoral injections at 2-week intervals, followed by RP 6 weeks after the second injection. Based on the findings of MRI and biopsy mapping, as a rule, one track injection to the most prominent cancer area was given to initial 12 patients and 3 track injections to multiple cancer areas in additional 6 patients. As compared to the former group, biochemical recurrence-free survival of the latter showed a significantly favorable outcome. Neoadjuvant Ad-REIC, mediating simultaneous induction of cancer selective apoptosis and augmentation of antitumor immunity, is a feasible approach in preventing cancer recurrence after RP. (199)

DOI： 10.1111/cts.12362

Molecular cloning of canine co-chaperone small glutamine-rich tetratricopeptide repeat-containing protein α (SGTA) and investigation of its ability to suppress androgen receptor signalling in androgen-independent prostate cancer. 査読

Kato Y, Ochiai K, Michishita M, Azakami D, Nakahira R, Morimatsu M, Ishiguro-Oonuma T, Yoshikawa Y, Kobayashi M, Bonkobara M, Kobayashi M, Takahashi K, Watanabe M, Omi T

Vet J 206 ( 2 ) 143 - 148 2015年11月

REIC/Dkk-3遺伝子治療の胸部悪性腫瘍に対する免疫刺激を介する抗腫瘍効果

諏澤憲, 枝園和彦, 阪口政清, 渡部昌実, 橋田真輔, 宗淳一, 山本寛斉, 牧祐歩, 佃和憲, 那須保友, 公文裕巳, 三好新一郎, 豊岡伸一

日本癌学会総会記事 74回 J - 1224 2015年10月

Distant Bystander Effect of REIC/Dkk-3 Gene Therapy through Immune System Stimulation in a Murine Model of Thoracic Malignancies 査読

Suzawa Ken, Shien Kazuhiko, Huang Peng, Sakaguchi Masakiyo, Watanabe Masami, Hashida Shinsuke, Soh Junichi, Yamamoto Hiromasa, Maki Yuho, Asano Hiroaki, Tsukuda Kazunori, Nasu Yasutomo, Kumon Hiromi, Miyoshi Shinichiro, Toyooka Shinichi

JOURNAL OF THORACIC ONCOLOGY 10 ( 9 ) S599 2015年9月

Possible role of cortactin phosphorylation by protein kinase Cα in actin-bundle formation at growth cone. 査読

Yamada H, Kikuchi T, Masumoto T, Wei FY, Abe T, Takeda T, Nishiki T, Tomizawa K, Watanabe M, Matsui H, Takei K

Biol Cell 107 ( 9 ) 319 - 330 2015年9月

A novel in situ permeation system and its utility in cancer tissue ablation 査読

Masami Watanabe

INTERNATIONAL JOURNAL OF ONCOLOGY 47 ( 3 ) 875 - 883 2015年9月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：SPANDIDOS PUBL LTD

Focal ablation therapy is an emerging treatment modality for localized cancer lesions. It is an attractive strategy for inhibiting tumor progression and preventing morbidity associated with open surgery. As for intratissue drug delivery systems for use in local therapy, the convection-enhanced delivery (CED) of liquid drugs has been utilized, particularly for the treatment of malignant brain tumors. Although the conventional CED system is useful for providing drug/vehicle-based local therapy, there are several reported disadvantages in terms of the ability to control the extent of drug diffusion. We herein developed and validated a novel in situ permeation (ISP)-MW-1 system for achieving intratissue drug diffusion. The ISP system includes a perfusion catheter connected to an injector and aspirator, which enables intratissue perfusion of the solute diluted in the vehicle in the tip-inserted cavity. We subsequently evaluated the utility of the ISP-MW-1 system for in situ permeation in a subcutaneous tumor model in hamsters. Dehydrated ethanol, saline and 50% acetic acid were evaluated as the vehicle, and methylene blue was used as a dissolved substance for evaluating the diffusion of the agent. As a result, almost all of the tumor tissue within the capsule (tumor size: 3 cm) was permeated with the dehydrated ethanol and 50% acetic acid and partially with the saline. We further demonstrated that ISP treatment with 50% acetic acid completely ablated the subcutaneous tumors in all of the treated hamsters (n=3). Therefore, the ISP-MW-1 system is a promising approach for controlling the intratissue diffusion of therapeutic agents and for providing local ablation therapy for cancer lesions. We believe that this system may be applicable to a broad range of medicinal and industrial fields, such as regenerative medicine, drug delivery systems, biochemistry and material technologies as well as cancer therapy.

DOI： 10.3892/ijo.2015.3068

Islet Culture/Preservation Before Islet Transplantation. 査読

Noguchi H, Miyagi-Shiohira C, Kurima K, Kobayashi N, Saitoh I, Watanabe M, Noguchi Y, Matsushita M

Cell Med. 8 ( 1月2日 ) 25 - 29 2015年8月

Cryopreservation of Adipose-Derived Mesenchymal Stem Cells. 査読

Miyagi-Shiohira C, Kurima K, Kobayashi N, Saitoh I, Watanabe M, Noguchi Y, Matsushita M, Noguchi H

Cell Med. 8 ( 1月2日 ) 3 - 7 2015年8月

Testosterone replacement therapy and cross-sex hormone treatment

Morito Sugimoto, Hiroaki Kurahashi, Yuko Matsumoto, Masami Watanabe, Yasutomo Nasu, Hiromi Kumon, Kazushi Ishii, Atsushi Nagai

Nishinihon Journal of Urology 77 ( 6 ) 202 - 206 2015年6月

記述言語：日本語掲載種別：研究論文（国際会議プロシーディングス）出版者・発行元：The West Japan Urological Association

Currently, androgen replacement therapy (ART) which is provided clinically in the field of urology is a common treatment for many of the patients with LOH syndrome or gender identity disorder (GID) Female to Male (FTM). We herein describe the current situation and the challenges regarding ART for these two diseases. ART has been provided according to LOH syndrome treatment guidelines. However, the opinion of any individual clinician and/or the patient's own adherence in continuing treatment will be seen as influencing treatment decisions at any particular moment. With regard to ART in our department, cases where ART has been judged to be effective for LOH syndrome comprised 51% of cases (23 out of 45 cases). No significant difference was showen between cases which hoped treatment continuation and didn't hope. Guidelines for correctly evaluating the response to treatment and the feasibility of providing such treatment are awaited. ART provided for GID-FTM is particular in that its therapeutic effect is expected to differ greatly from that obtained when ART is provided for LOH syndrome. Order to obtain a cessation of the expected virilization and physiology of a patient's body, the hormone dose and interval need to be carefully considered. Data accumulated in our department have demonstrated that in most patients, a sufficient therapeutic effect is obtained by the administration of 125 mg testosterone enanthate for 2 weeks, and serious adverse events have not been observed. Guidelines regarding GID androgen replacement therapy are also awaited, in particular with regard to a more detailed evaluation of its predicted therapeutic effect.

The cysteine-rich core domain of REIC/Dkk-3 is critical for its effect on monocyte differentiation and tumor regression 査読

Rie Kinoshita, Masami Watanabe, Peng Huang, Shun-Ai Li, Masakiyo Sakaguchi, Hiromi Kumon, Junichiro Futami

ONCOLOGY REPORTS 33 ( 6 ) 2908 - 2914 2015年6月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：SPANDIDOS PUBL LTD

Reduced expression in immortalized cells (REIC)/Dickkopf (Dkk)-3 is a tumor-suppressor gene and has been studied as a promising therapeutic gene for cancer gene therapy. Intratumoral injection of an adenovirus vector carrying the human REIC/Dkk-3 gene (Ad-REIC) elicits cancer cell-specific apoptosis and anticancer immune responses. The cytokine-like effect of secretory REIC/Dkk-3 on the induction of dendritic cell (DC)-like cell differentiation from monocytes plays a role in systemic anticancer immunity. In the present study, we generated recombinant full-length and N-terminally truncated REIC/Dkk-3 to characterize the biological activity of the protein. During the purification procedure, we identified a 17 kDa cysteine-rich stable product (C17-REIC) showing limited degradation. Further analysis showed that the C17-REIC domain was sufficient for the induction of DC-like cell differentiation from monocytes. Concomitant with the differentiation of DCs, the REIC/Dkk-3 protein induced the phosphorylation of glycogen synthase kinase 3 beta (GSK-3 beta) and signal transducers and activators of transcription (STAT) at a level comparable to that of granulocyte/macrophage colony-stimulating factor. In a mouse model of subcutaneous renal adenocarcinoma, intraperitoneal injection of full-length and C17-REIC proteins exerted anticancer effects in parallel with the activation of immunocompetent cells such as DCs and cytotoxic T lymphocytes in peripheral blood. Taken together, our results indicate that the stable cysteine-rich core region of REIC/Dkk-3 is responsible for the induction of anticancer immune responses. Because REIC/Dkk-3 is a naturally circulating serum protein, the upregulation REIC/Dkk-3 protein expression could be a promising option for cancer therapy.

DOI： 10.3892/or.2015.3885

Potential Factors for the Differentiation of ESCs/iPSCs Into Insulin-Producing Cells. 査読

Takako Tsugata, Naruo Nikoh, Tatsuya Kin, Issei Saitoh, Yasufumi Noguchi, Hideo Ueki, Masami Watanabe, Andrew M, James Shapiro, Hirofumi Noguchi

Cell Medicine 7 ( 2 ) 83 - 93 2015年4月

A vaccine strategy with multiple prostatic acid phosphatase-fused cytokines for prostate cancer treatment 査読

Kei Fujio, Masami Watanabe, Hideo Ueki, Shun-Ai Li, Rie Kinoshita, Kazuhiko Ochiai, Junichiro Futami, Toyohiko Watanabe, Yasutomo Nasu, Hiromi Kumon

ONCOLOGY REPORTS 33 ( 4 ) 1585 - 1592 2015年4月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：SPANDIDOS PUBL LTD

Immunotherapy is one of the attractive treatment strategies for advanced prostate cancer. The US Food and Drug Administration (FDA) previously approved the therapeutic vaccine, sipuleucel-T, which is composed of autologous antigen-presenting cells cultured with a fusion protein [prostatic acid phosphatase (PAP) and granulocyte-macrophage colony-stimulating factor (GMCSF)]. Although sipuleucel-T has been shown to prolong the median survival of patients for 4.1 months, more robust therapeutic effects may be expected by modifying the vaccination protocol. In the present study, we aimed to develop and validate a novel vaccination strategy using multiple PAP-fused cytokines for prostate cancer treatment. Using a super gene expression (SGE) system that we previously established to amplify the production of a recombinant protein, significant amounts of PAP-fused cytokines [human GMCSF, interleukin-2 (IL2), IL4, IL7 and mouse GMCSF and IL4] were obtained. We examined the activity of the fusion proteins in vitro to validate their cytokine functions. A significant upregulation of dendritic cell differentiation from monocytes was achieved by PAP-GMCSF when used with the other PAP-fused cytokines. The PAP-fused human IL2 significantly increased the proliferation of lymphocytes, as determined by flow cytometry. We also investigated the in vivo therapeutic effects of multiple PAP-fused cytokines in a mouse prostate cancer model bearing prostate-specific antigen (PSA)- and PAP-expressing tumors. The simultaneous intraperitoneal administration of PAP-GMCSF, -IL2, -IL4 and -IL7 significantly prevented tumor induction and inhibited the tumor growth in the PAP-expressing tumors, yet not in the PSA-expressing tumors. The in vivo therapeutic effects with the multiple PAP-fused cytokines were superior to the effects of PAP-GMCSF alone. We thus demonstrated the advantages of the combined use of multiple PAP-fused cytokines including PAP-GMCSF, and propose a promising prostatic antigen-vaccination strategy to enhance the therapeutic effects.

DOI： 10.3892/or.2015.3770

Polymorphisms of canine BRCA2 BRC repeats affecting interaction with RAD51 査読

Kazuhiko Ochiai, Toshina Ishiguro-Oonuma, Yasunaga Yoshikawa, Chihiro Udagawa, Yuiko Kato, Masami Watanabe, Makoto Bonkobara, Masami Morimatsu, Toshinori Omi

BIOMEDICAL RESEARCH-TOKYO 36 ( 2 ) 155 - 158 2015年4月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：BIOMEDICAL RESEARCH PRESS LTD

Mutations in the breast cancer susceptibility gene BRCA2 leading to the failure of interactions with the recombinase RAD51 are associated with an increased risk of cancer in humans. This interaction depends on the eight BRC repeat (BRC1-8) sequences in BRCA2. We previously reported that canine BRC3 has two polymorphisms (T1425P and K1435R) influencing the interaction with RAD51, and 1435R was identified in mammary tumor dog samples. In this study, we investigated the sequence variations of BRC3 and 4 in 236 dogs of five breeds. Allele frequencies of 1425P and 1435R were 0.063 and 0.314, respectively, and there was no other polymorphism in the sequenced region. A mammalian two-hybrid assay using BRC3-4 sequences demonstrated that 1425P allele reduced the binding strength with RAD51 but 1435R had no effect. These results may provide an insight into the functions of not only individual but also multiple BRC repeats of BRCA2 in dogs.

DOI： 10.2220/biomedres.36.155

Ad-REIC gene therapy: Promising results in a patient with metastatic CRPC following chemotherapy 査読

Hiromi Kumon, Katsumi Sasaki, Yuichi Ariyoshi, Takuya Sadahira, Shin Ebara, Takao Hiraki, Susumu Kanazawa, Hiroyuki Yanai, Masami Watanabe, Yasutomo Nasu

Clinical Medicine Insights: Oncology 9 31 - 38 2015年3月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：Libertas Academica Ltd.

A 63-year-old man with metastatic castration-resistant prostate cancer (CRPC) was successfully treated for two years with in situ gene therapy using an adenovirus vector carrying the human REIC/Dkk‑3 gene (Ad-REIC), following chemotherapy. Ad-REIC mediates simultaneous induc-tion of cancer-selective apoptosis and augmentation of antitumor immunity, and a Phase I/IIa clinical study on Ad-REIC has been conducted at Okayama University Hospital since January 2011. At the time of enrollment in December 2012, the patient presented with rapid progression of lymph node (LN) metastases. Two scheduled Ad-REIC injections and 10 additional Ad-REIC injections into metastatic pelvic and para-aortic LNs under CT guidance, with an average four weeks’ interval, exhibited the potent direct and indirect effects of Ad-REIC as a therapeutic cancer vaccine. During the next 12 months, three additional injections into para-aortic LNs showing regrowth achieved adequate control of all metastatic LNs with prostate-specific antigen (PSA) decline, without any particular adverse events.

DOI： 10.4137/CMO.S23252

Integrin antagonist augments the therapeutic effect of adenovirus-mediated REIC/Dkk-3 gene therapy for malignant glioma 査読

Y. Shimazu, K. Kurozumi, T. Ichikawa, K. Fujii, M. Onishi, J. Ishida, T. Oka, M. Watanabe, Y. Nasu, H. Kumon, I. Date

GENE THERAPY 22 ( 2 ) 146 - 154 2015年2月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：NATURE PUBLISHING GROUP

Reduced expression in immortalized cells/Dickkopf-3 (REIC/Dkk-3) was identified as a gene whose expression is reduced in many human cancers. REIC/Dkk-3 expression is also downregulated in malignant glioma and regulates cell growth through caspase-dependent apoptosis. cRGD (EMD121974), an antagonist of integrins, has demonstrated preclinical efficacy against malignant glioma. In this study, we investigated the antiglioma effect of combination therapy using an adenovirus vector carrying REIC/Dkk-3 (Ad-REIC) and cRGD. Quantitative real-time reverse-transcription PCR revealed the reduction of REIC/Dkk-3 mRNA levels in malignant glioma cell lines. The reduction of REIC/Dkk-3 protein expression in malignant glioma cell lines was also confirmed with western blot analysis. After treatment with Ad-REIC and cRGD, the proliferative rate of malignant glioma cells was significantly reduced in a time-dependent manner. In vivo, there was a statistically significant increase in the survival of mice treated with Ad-REIC and cRGD combination therapy compared with Ad-REIC monotherapy. We identified an apoptotic effect following monotherapy with Ad-REIC. Moreover, cRGD augmented the antiglioma efficacy of Ad-REIC. These results may lead to a promising new approach for the treatment of malignant glioma.

DOI： 10.1038/gt.2014.100

Induction of tissue-specific stem cells by reprogramming factors, and tissue-specific selection 査読

H. Noguchi, I. Saitoh, T. Tsugata, H. Kataoka, M. Watanabe, Y. Noguchi

CELL DEATH AND DIFFERENTIATION 22 ( 1 ) 145 - 155 2015年1月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：NATURE PUBLISHING GROUP

Although induced pluripotent stem (iPS) cells have significant implications for overcoming most of the ethical issues associated with embryonic stem (ES) cells, there are still several unresolved issues related to the use of iPS cells for clinical applications, such as teratoma formation. In this study, we were able to generate tissue-specific stem (induced tissue-specific stem; iTS) cells from the pancreas (iTS-P) or liver (iTS-L) by transient overexpression of reprogramming factors, combined with tissue-specific selection. The generation of iTS cells was easier than that of iPS cells. The iTS-P/iTS-L cells express genetic markers of endoderm and pancreatic/hepatic progenitors and were able to differentiate into insulin-producing cells/hepatocytes more efficiently than ES cells. Subcutaneous transplantation of both types of iTS cells into immunodeficient mice resulted in no teratoma formation. The technology used for the transient overexpression of reprogramming factors and tissue-specific selection may be useful for the generation of other tissue-specific stem cells, and the generation of iTS cells could have important implications for the clinical application of stem cells.

DOI： 10.1038/cdd.2014.132

Potential Factors for the Differentiation of ESCs/iPSCs Into Insulin-Producing Cells. 査読

Tsugata T, Nikoh N, Kin T, Saitoh I, Noguchi Y, Ueki H, Watanabe M, James Shapiro AM, Noguchi H

Cell Med 7 ( 2 ) 83 - 93 2014年12月

Significant association between the Axin2 rs2240308 single nucleotide polymorphism and the incidence of prostate cancer 査読

Chao Ma, Chunxiao Liu, Peng Huang, Haruki Kaku, Jie Chen, Kai Guo, Hideo Ueki, Akiko Sakai, Yasutomo Nasu, Hiromi Kumon, Kenji Shimizu, Masami Watanabe

ONCOLOGY LETTERS 8 ( 2 ) 789 - 794 2014年8月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：SPANDIDOS PUBL LTD

The Wnt signaling pathway plays a crucial role in human cancer development, and axis inhibition protein 2 (Axin2) is a master scaffold protein involved in Wnt signaling. Axin2 negatively regulates Wnt signaling and acts as a tumor suppressor protein. The present study evaluated the association between the Axin2 single nucleotide polymorphism (SNP) rs2240308 [guanine (G)/adenine (A)] and the incidence of prostate cancer. In total, 103 patients with prostate cancer and 100 cancer-free control males were included in this case-control study, and were genotyped using the genomic DNA extracted from peripheral blood samples. The results revealed a higher incidence of prostate cancer in the subjects with the homozygous GG genotype and a reduced cancer incidence in the patients with the GA genotype of the rs2240308 SNP (G/A) in the Axin2 gene. T-he adjusted odds ratio for carriers with the GA genotype was 0.377 (95% CI, 0.206-0.688; P=0.001) and that for the AA genotype was 0.830 (95% CI, 0.309-2.232; P=0.712) compared with the GG genotype. Therefore, the GA genotype was found to exhibit a protective effect that decreased the risk of prostate cancer. To the best of our knowledge, this is the first study to demonstrate the significant association between this SNP (rs2240308, G/A) and the risk of prostate cancer. This association indicates the possibility that the variations in the Axin2 gene in this position may play a significant role in promoting the development of cancer in the prostate. We believe that the Axin2 SNP (rs2240308) could be a useful biomarker for the predisposition and early diagnosis of the disease.

DOI： 10.3892/ol.2014.2177

Immunological aspects of REIC/Dkk-3 gene therapy : the mechanism of the robust anti-tumor effects. 査読

Masami Watanabe, Peng Huang, Fernando Abarzua, Haruki Kaku, Katsumi Sasaki, Hideo Ueki, Toyohiko Wananabe, Yasutomo Nasu, Hiromi Kumon

JOURNAL OF GENE MEDICINE 16 ( 7-8 ) 226 - 227 2014年7月

Dramatic Increase in Expression of a Transgene by Insertion of Promoters Downstream of the Cargo Gene 査読

Masakiyo Sakaguchi, Masami Watanabe, Rie Kinoshita, Haruki Kaku, Hideo Ueki, Junichiro Futami, Hitoshi Murata, Yusuke Inoue, Shun-Ai Li, Peng Huang, Endy Widya Putranto, I. Made Winarsa Ruma, Yasutomo Nasu, Hiromi Kumon, Nam-ho Huh

MOLECULAR BIOTECHNOLOGY 56 ( 7 ) 621 - 630 2014年7月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：HUMANA PRESS INC

For expression of genes in mammalian cells, various vectors have been developed using promoters including CMV, EF-1 alpha, and CAG promoters and have been widely used. However, such expression vectors sometimes fail to attain sufficient expression levels depending on the nature of cargo genes and/or on host cell types. In the present study, we aimed to develop a potent promoter system that enables high expression levels of cargo genes ubiquitously in many different cell types. We found that insertion of an additional promoter downstream of a cargo gene greatly enhanced the expression levels. Among the constructs we tested, C-TSC cassette (C: CMV-RU5' located upstream; TSC: another promoter unit composed of triple tandem promoters, hTERT, SV40, and CMV, located downstream of the cDNA plus a polyadenylation signal) had the most potent capability, showing far higher efficiency than that of potent conventional vector systems. The results indicate that the new expression system is useful for production of recombinant proteins in mammalian cells and for application as a gene therapeutic measure.

DOI： 10.1007/s12033-014-9738-0

Potential of adenovirus-mediated REIC/Dkk-3 gene therapy for use in the treatment of pancreatic cancer 査読

Daisuke Uchida, Hidenori Shiraha, Hironari Kato, Teruya Nagahara, Masaya Iwamuro, Junro Kataoka, Shigeru Horiguchi, Masami Watanabe, Akinobu Takaki, Kazuhiro Nouso, Yasutomo Nasu, Takahito Yagi, Hiromi Kumon, Kazuhide Yamamoto

JOURNAL OF GASTROENTEROLOGY AND HEPATOLOGY 29 ( 5 ) 973 - 983 2014年5月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：WILEY-BLACKWELL

Background and AimThe reduced expression in immortalized cells REIC/the dickkopf 3 (Dkk-3) gene, tumor suppressor gene, is downregulated in various malignant tumors. In a prostate cancer study, an adenovirus vector carrying the REIC/Dkk-3 gene (Ad-REIC) induces apoptosis. In the current study, we examined the effects of REIC/Dkk-3 gene therapy in pancreatic cancer.
MethodsREIC/Dkk-3 expression was assessed by immunoblotting and immunohistochemistry in the pancreatic cancer cell lines (ASPC1, MIAPaCa2, Panc1, BxPC3, SUIT-2, KLM1, and T3M4) and pancreatic cancer tissues. The Ad-REIC agent was used to investigate the apoptotic effect in vitro and antitumor effects in vivo. We also assessed the therapeutic effects of Ad-REIC therapy with gemcitabine.
ResultsThe REIC/Dkk-3 expression was lost in the pancreatic cancer cell lines and decreased in pancreatic cancer tissues. Ad-REIC induced apoptosis and inhibited cell growth in the ASPC1 and MIAPaCa2 lines in vitro, and Ad-REIC inhibited tumor growth in the mouse xenograft model using ASPC1 cells. The antitumor effect was further enhanced in combination with gemcitabine. This synergistic effect may be caused by the suppression of autophagy via the enhancement of mammalian target of rapamycin signaling.
ConclusionsAd-REIC induces apoptosis and inhibits tumor growth in pancreatic cancer cell lines. REIC/Dkk-3 gene therapy is an attractive therapeutic tool for pancreatic cancer.

DOI： 10.1111/jgh.12501

Establishment of a pancreatic stem cell line from fibroblast-derived induced pluripotent stem cells 査読

Takashi Kuise, Hirofumi Noguchi, Hiroshi Tazawa, Takashi Kawai, Masaya Iwamuro, Issei Saitoh, Hitomi Usui Kataoka, Masami Watanabe, Yasufumi Noguchi, Toshiyoshi Fujiwara

BIOMEDICAL ENGINEERING ONLINE 13 2014年5月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：BIOMED CENTRAL LTD

Background: For cell therapies to treat diabetes, it is important to produce a sufficient number of pancreatic endocrine cells that function similarly to primary islets. Induced pluripotent stem (iPS) cells represent a potentially unlimited source of functional pancreatic endocrine cells. However, the use of iPS cells for laboratory studies and cell-based therapies is hampered by their high tumorigenic potential and limited ability to generate pure populations of differentiated cell types in vitro. The purpose of this study was to establish a pancreatic stem cell line from iPS cells derived from mouse fibroblasts.
Methods: Mouse iPS cells were induced to differentiate into insulin-producing cells by a multi-step differentiation protocol, which was conducted as described previously with minor modifications. Selection of the pancreatic stem cell was based on morphology and Pdx1 expression. The pancreatic potential of the pancreatic stem cells was evaluated using a reverse transcription PCR, real-time PCR, immunofluorescence, and a glucose challenge test. To assess potential tumorigenicity of the pancreatic stem cells, the cells were injected into the quadriceps femoris muscle of the left hindlimb of nude mice.
Results: The iPS-derived pancreatic stem cells expressed the transcription factor -Pdx1- a marker of pancreatic development, and continued to divide actively beyond passage 80. Endocrine cells derived from these pancreatic stem cells expressed insulin and pancreatic genes, and they released insulin in response to glucose stimulation. Mice injected with the pancreatic stem cells did not develop tumors, in contrast to mice injected with an equal number of iPS cells.
Conclusion: This strategy provides a new approach for generation of insulin-producing cells that is more efficient and safer than using iPS cells. We believe that this approach will help to develop a patient-specific cell transplantation therapy for diabetes in the near future.

DOI： 10.1186/1475-925X-13-64

リプログラミングファクターの導入による人工膵幹細胞の作成

野口洋文, 齊藤一誠, 片岡仁美, 渡部昌実, 野口寧文, 西郷健一, 長谷川正行, 丸山通広, 大月和宣, 青山博道, 松本育子, 浅野武秀, 圷尚武

糖尿病 57 ( Suppl.1 ) S - 440 2014年4月

A novel gene expression system strongly enhances the anticancer effects of a REIC/Dkk-3-encoding adenoviral vector 査読

Masami Watanabe, Masakiyo Sakaguchi, Rie Kinoshita, Haruki Kaku, Yuichi Ariyoshi, Hideo Ueki, Ryuta Tanimoto, Shin Ebara, Kazuhiko Ochiai, Junichiro Futami, Shun-Ai Li, Peng Huang, Yasutomo Nasu, Nam-Ho Huh, Hiromi Kumon

ONCOLOGY REPORTS 31 ( 3 ) 1089 - 1095 2014年3月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：SPANDIDOS PUBL LTD

Gene expression systems with various promoters, including the cytomegalovirus (CMV) promoter, have been developed to increase the gene expression in a variety of normal and cancer cells. In particular, in the clinical trials of cancer gene therapy, a more efficient and robust gene expression system is required to achieve sufficient therapeutic outcomes. By inserting the triple translational enhancer sequences of human telomerase reverse transcriptase (hTERT), Simian virus 40 (SV40) and CMV downstream of the sequence of the BGH polyA, we were able to develop a novel gene expression system that significantly enhances the expression of the genes of interest. We termed this novel gene expression cassette the super gene expression (SGE) system, and herein verify the utility of the SGE cassette for a replication-deficient adenoviral vector. We newly developed an adenoviral vector expressing the tumor suppressor, reduced expression in immortalized cells (REIC)/Dickkopf-3 (Dkk-3), based on the CMV promoter-driven SGE system (Ad-SGE-REIC) and compared the therapeutic utility of Ad-SGE-REIC with that of the conventional adenoviral vectors (Ad-CMV-REIC or Ad-CAG-REIC). The results demonstrated that the CMV promoter-SGE system allows for more potent gene expression, and that the Ad-SGE-REIC is superior to conventional adenoviral systems in terms of the REIC protein expression and therapeutic effects. Since the SGE cassette can be applied for the expression of various therapeutic genes using various vector systems, we believe that this novel system will become an innovative tool in the field of gene expression and gene therapy.

DOI： 10.3892/or.2013.2958

Adenovirus-mediated REIC/Dkk-3 gene therapy: Development of an autologous cancer vaccination therapy (Review) 査読

Masami Watanabe, Yasutomo Nasu, Hiromi Kumon

ONCOLOGY LETTERS 7 ( 3 ) 595 - 601 2014年3月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：SPANDIDOS PUBL LTD

Reduced expression in immortalized cells (REIC)/Dickkopf (Dkk)-3 is a tumor suppressor and therapeutic gene and has been studied with respect to the application of cancer gene therapy. Our previous studies demonstrated that the intratumoral injection of an adenovirus vector carrying the human REIC/Dkk-3 gene (Ad-REIC) suppresses tumor growth in mouse models of prostate, breast and testicular cancer and malignant mesothelioma. The mechanisms underlying these antitumor therapeutic effects have only been clarified recently. It has been demonstrated that Ad-REIC treatment inhibits cancer progression via the upregulation of systemic anticancer immunity. Under experimental conditions, autologous cancer vaccination via cancer-specific apoptosis and anticancer immune activation is a possible therapeutic mechanism. The robust anticancer effects observed in previous preclinical studies support the clinical utility of Ad-REIC. At present, a phase I-IIa study of Ad-REIC gene therapy in prostate cancer patients is ongoing. The current study reviews the observations of previous fundamental studies and summarizes the anticancer mechanisms of intratumoral Ad-REIC treatment in terms of cancer vaccination.

DOI： 10.3892/ol.2013.1777

Anti-Cancer Effects of REIC/Dkk-3-encoding Adenoviral Vector for the Treatment of Non-small Cell Lung Cancer 査読

Kazuhiko Shien, Norimitsu Tanaka, Masami Watanabe, Junichi Soh, Masakiyo Sakaguchi, Keitaro Matsuo, Hiromasa Yamamoto, Masashi Furukawa, Hiroaki Asano, Kazunori Tsukuda, Yasutomo Nasu, Nam-Ho Huh, Shinichiro Miyoshi, Hiromi Kumon, Shinichi Toyooka

PLOS ONE 9 ( 2 ) 2014年2月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：PUBLIC LIBRARY SCIENCE

Objectives: REIC/Dkk-3 is down-regulated in a broad range of human cancer cells and is considered to function as a tumor suppressor. We previously reported that REIC/Dkk-3-expressing adenovirus vector (Ad-REIC) induced endoplasmic reticulum (ER) stress and cancer-specific apoptosis in human prostate cancer. In this study, we examined the therapeutic impact of Ad-REIC on non-small cell lung cancer (NSCLC).
Materials and Methods: We examined the anti-tumor effect of Ad-REIC on 25 NSCLC cell lines in vitro and A549 cells in vivo. Two of these cell lines were artificially established as EGFR-tyrosine kinase inhibitor (TKI) resistant sublines.
Results: Ad-REIC-treatment inhibited the cell viability by 40% or more in 13 (52%) of the 25 cell lines at multiplicity of infection (MOI) of 20 (20 MOI). These cell lines were regarded as being highly sensitive cells. The cell viability of a nonmalignant immortalized cell line, OUMS-24, was not inhibited at 200 MOI of Ad-REIC. The effects of Ad-REIC on EGFR-TKI resistant sublines were equivalent to those in the parental cell lines. Here, we demonstrated that Ad-REIC treatment activated c-Jun N-terminal kinase (JNK) in NSCLC cell lines, indicating the induction of ER stress with GRP78/BiP (GRP78) up-regulation and resulting in apoptosis. A single intratumoral injection of Ad-REIC significantly inhibited the tumorigenic growth of A549 cells in vivo. As predictive factors of sensitivity for Ad-REIC treatment in NSCLC, we examined the expression status of GRP78 and coxsackievirus and adenovirus receptor (CAR). We found that the combination of the GRP78 and CAR expressional statuses may be used as a predictive factor for Ad-REIC sensitivity in NSCLC cells.
Conclusion: Ad-REIC induced JNK activation and subsequent apoptosis in NSCLC cells. Our study indicated that Ad-REIC has therapeutic potential against NSCLC and that the expression statuses of GRP78 and CAR may predict a potential therapeutic benefit of Ad-REIC.

DOI： 10.1371/journal.pone.0087900

N’-[4-(dipropylamino)benzylidene]-2-hydroxybenzohydrazide is a dynamin GTPase inhibitor that suppresses cancer cell migration and invasion by inhibiting actin polymerization . 査読

Hiroshi Yamada, Tadashi Abe, Shun-Ai Li, Shota Tago, Peng Huang, Masami Watanabe, Satoru Ikeda, Naohisa Ogo, Akira Asai, Kohji Takei

Biochem Biophys Res Commun 443 ( 2 ) 511 - 517 2014年1月

A rare complication: Misdirection of an indwelling urethral catheter into the ureter 査読

Tsutomu Ishikaw, Motoo Araki, Takeshi Hirata, Masami Watanabe, Shin Ebara, Toyohiko Watanabe, Yasutomo Nasu, Hiromi Kumon

Acta Medica Okayama 68 ( 1 ) 47 - 51 2014年

記述言語：英語掲載種別：研究論文（学術雑誌）

We report 3 patients with the rare complication of an indwelling urethral catheter misdirected into the ureter. This is the largest series to date. Patients were referred to us for a variety of reasons following exchange of their chronic indwelling urinary catheters. CT in all cases demonstrated the urinary catheters residing in the left ureter. The ages of the patients were 37, 67 and 81 years old. All patients suffered from neurogenic bladder. Two patients were female, one was male, and 2 of the 3 had a sensory disorder inhibiting their pain response. The catheters were replaced with open-end Foley catheters. Extensive follow-up CT scans were obtained in one case, demonstrating improvement of hydronephrosis and no evidence of ureteral stenosis. Cystoscopy in this patient demonstrated normally positioned and functioning ureteral orifices. Although the placement of an indwelling urethral catheter is a comparatively safe procedure, one must keep in mind that this complication can occur, particularly in female patients with neurogenic bladder. CT without contrast is a noninvasive, definitive diagnostic tool. © 2014 by Okayama University Medical School.

DOI： 10.18926/AMO/52144

Comparison of Incubation Solutions Prior to the Purification of Porcine Islet Cells. 国際誌

Takashi Kawai, Hirofumi Noguchi, Takashi Kuise, Atsuko Nakatsuka, Akihiro Katayama, Noriko Imagawa, Hitomi Usui Kataoka, Issei Saitoh, Yasufumi Noguchi, Masami Watanabe, Toshiyoshi Fujiwara

Cell medicine 6 ( 1-2 ) 9 - 14 2013年12月

記述言語：英語掲載種別：研究論文（学術雑誌）

For pancreatic islet transplantation, one of the most important steps of islet isolation is islet purification. The most common method of islet purification is density gradient centrifugation because there are differences in density between islets and acinar tissue. However, the density of islets/acinar tissue depends on several conditions, such as the incubation time before purification and the osmolality of the preincubation solution. In this study, we evaluated the impact of using two different preincubation solutions before purification. We used the University of Wisconsin (UW) solution and a new preservation solution (HN-1), which we recently developed. There were no significant differences between the two solutions in terms of the islet yield, rate of viability, and purity or stimulation index after purification. There were also no differences in the attainability and suitability of posttransplantation normoglycemia. Our study shows that the HN-1 solution is equivalent to the UW solution for preincubation before islet purification.

DOI： 10.3727/215517913X674180

Comparison of New Preservation Solutions, HN-1 and University of Wisconsin Solution, in Pancreas Preservation for Porcine Islet Isolation. 査読国際誌

Katayama Akihiro, Noguchi Hirofumi, Kuise Takashi, Nakatsuka Atsuko, Hirota Daisho, Kataoka Hitomi, Kawai Takashi, Inoue Kentaro, Imagawa Noriko, Saitoh Issei, Noguchi Yasufumi, Watanabe Masami, Wada Jun, Fujiwara Toshiyoshi

Cell Medicine 6 ( 1-2 ) 3 - 8 2013年12月

Testosterone replacement elevates the serum uric acid levels in patients with female to male gender identity disorder 査読

Hiroaki Kurahashi, Masami Watanabe, Morito Sugimoto, Yuichi Ariyoshi, Sabina Mahmood, Motoo Araki, Kazushi Ishii, Yasutomo Nasu, Atsushi Nagai, Hiromi Kumon

ENDOCRINE JOURNAL 60 ( 12 ) 1321 - 1327 2013年12月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：JAPAN ENDOCRINE SOC

Gender identity disorder (GID) results from a disagreement between a person's biological sex and the gender to which he or she identifies. With respect to the treatment of female to male GID, testosterone replacement therapy (TRT) is available. The uric acid (UA) level can be influenced by testosterone; however, the early effects and dose-dependency of TRT on the serum UA concentration have not been evaluated in this population. We herein conducted a dose-response analysis of TRT in 160 patients with female to male GID. The TRT consisted of three treatment groups who received intramuscular injections of testosterone enanthate: 125 mg every two weeks, 250 mg every three weeks and 250 mg every two weeks. Consequently, serum UA elevation was observed after three months of TRT and there was a tendency toward testosterone dose-dependency. The onset of hyperuricemia was more prevalent in the group who received the higher dose. We also demonstrated a positive correlation between increased levels of serum UA and serum creatinine. Since the level of serum creatinine represents an individual's muscle volume and the muscle is a major source of purine, which induces UA upregulation, the serum UA elevation observed during TRT is at least partially attributed to an increase in muscle mass. This is the first study showing an association between serum UA elevation and a TRT-induced increase in muscle mass. The current study provides important information regarding TRT for the follow-up and management of the serum UA levels in GID patients.

DOI： 10.1507/endocrj.EJ13-0203

Culture Conditions for Mouse Pancreatic Stem Cells. 国際誌

Hirofumi Noguchi, Issei Saitoh, Hitomi Usui Kataoka, Masami Watanabe, Yasufumi Noguchi, Toshiyoshi Fujiwara

Cell medicine 5 ( 2-3 ) 63 - 8 2013年11月

記述言語：英語掲載種別：研究論文（学術雑誌）

Recently, mouse pancreatic stem cells have been isolated from adult mouse pancreata. However, these pancreatic stem cells could be maintained only under specific culture conditions with lot-limited fetal bovine serum (FBS). For the efficient isolation and maintenance of mouse pancreatic stem cells, it is important to identify culture conditions that can be used independent of the FBS lot. In this study, we evaluated the culture conditions required to maintain mouse pancreatic stem cells. The mouse pancreatic stem cells derived from the pancreas of a newborn mouse, HN#101, were cultured under the following conditions: 1) Dulbecco's modified Eagle's medium (DMEM) with 20% lot-limited FBS, in which mouse pancreatic stem cells could be cultured without changes in morphology and growth activity; 2) complete embryonic stem (ES) cell media; and 3) complete ES cell media on feeder layers of mitomycin C-treated STO cells, which were the same culture conditions used for mouse ES cells. Under culture conditions #1 and #3, the HN#101 cells continued to form a flat "cobblestone" monolayer and continued to divide actively beyond the population doubling level (PDL) 100 without growth inhibition, but this did not occur under culture condition #2. The gene expression profile and differentiated capacity of the HN#101 cells cultured for 2 months under culture condition #3 were similar to those of HN#101 cells at PDL 50. These data suggest that complete ES cell media on feeder layers could be useful for maintaining the undifferentiated state of pancreatic stem cells.

DOI： 10.3727/215517913X666495

Isolation Efficiency of Mouse Pancreatic Stem Cells Is Age Dependent. 国際誌

Takashi Kuise, Hirofumi Noguchi, Issei Saitoh, Hitomi Usui Kataoka, Masami Watanabe, Yasufumi Noguchi, Toshiyoshi Fujiwara

Cell medicine 5 ( 2-3 ) 69 - 73 2013年11月

記述言語：英語掲載種別：研究論文（学術雑誌）

Mouse pancreatic stem cells have been isolated from mouse pancreata. This study evaluated the efficacy of isolating mouse pancreatic stem cells using mice of different ages. The pancreata of newborn mice, 8-week-old mice, and 24-week-old mice were harvested and digested by using collagenase. The "duct-like" cells in the digested pancreatic tissue were then inoculated into 96-well plates, cloned by limiting dilution, and cultured in DMEM with 20% FBS. Pancreatic stem cells were isolated from the pancreata of all newborn mice, while cells could only be isolated from 10% of the pancreata of 8-week-old mice and could not be isolated from the pancreata of any 24-week-old mice. These data suggest that young mice may have some pancreatic stem cells and that older mice may only have a few pancreatic stem cells. These data also indicate that it is extremely difficult to isolate pancreatic stem cells from older mice, suggesting that future research focus its efforts on finding methods of isolating pancreatic stem cells from adult mice.

DOI： 10.3727/215517913X666503

Comparison of incubation solutions prior to the purification of porcine islet cells. 査読

Kawai Takashi, Noguchi Hirofumi, Kuise Takashi, Nakatsuka Atsuko, Katayama Akihiro, Imagawa Noriko, Kataoka, Hitomi Usui, Saitoh Issei, Noguchi Yasufumi, Watanabe Masami, Fujiwara Toshiyoshi

Cell Medicine 2013年10月

Molecular cloning and tumour suppressor function analysis of canine REIC/Dkk-3 in mammary gland tumours 査読

Kazuhiko Ochiai, Masami Watanabe, Daigo Azakami, Masaki Michishita, Yasunaga Yoshikawa, Chihiro Udagawa, Pornphimon Metheenukul, Thippayarat Chahomchuen, Hiroshi Aoki, Hiromi Kumon, Masami Morimatsu, Toshinori Omi

VETERINARY JOURNAL 197 ( 3 ) 769 - 775 2013年9月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：ELSEVIER SCI LTD

REIC/Dkk-3, a member of the human Dickkopf (Dkk) family, plays a role as a suppressor of growth in several human cancers. In this study, the tumour suppression function of canine REIC/Dkk-3 was investigated. The full-length open reading frame of the canine REIC/Dkk-3 homologue was cloned and the tissue distribution of REIC/Dkk-3 mRNA was determined, along with the subcellular localisation of the REIC/Dkk-3 protein in canine cancer cell lines. Expression of REIC/Dkk-3 was lower in mammary gland tumours and in canine mammary carcinoma cell lines than in normal mammary gland tissue. Overexpression of REIC/Dkk-3 induced apoptosis in canine mammary carcinoma cell lines. These results show that expression of REIC/Dkk-3 is downregulated in canine mammary tumours and that one of the functions of this gene is induction of apoptosis. (C) 2013 Elsevier Ltd. All rights reserved.

DOI： 10.1016/j.tvjl.2013.04.024

Clinical significance of CD24 as a predictor of bladder cancer recurrence 査読

Chunxiao Liu, Shaobo Zheng, Haiyan Shen, Kai Xu, Jie Chen, Hulin Li, Yawen Xu, Abai Xu, Binshen Chen, Haruki Kaku, Yasutomo Nasu, Hiromi Kumon, Peng Huang, Masami Watanabe

ONCOLOGY LETTERS 6 ( 1 ) 96 - 100 2013年7月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：SPANDIDOS PUBL LTD

Cluster of differentiation (CD)24 was originally described as a B lymphocyte marker and has recently received considerable attention in cancer research as its overexpression has been observed in several types of carcinoma. The CD24 molecule is a glycosyl-phosphatidylinositol-linked cell surface protein that appears to be associated with aggressive cancers involving invasion and metastasis. However, the expression of CD24 in human bladder cancer and its clinical significance remains largely unknown and no association has been reported between CD24 overexpression and human bladder tumor recurrence. In the present study, the CD24 expression in cancer tissues obtained during transurethral surgery and the subsequent intra-bladder tumor recurrence following surgery were assessed. Immunohistochemical staining was performed and the intensity of CD24 staining was semi-quantitatively evaluated. CD24 expression was observed more frequently in high-grade bladder tumors (G2-G3) than low-grade tumors (G1). Positive CD24 expression was significantly associated with intra-bladder tumor recurrence following surgery and increased staining intensity was also correlated with recurrence. The positive association between CD24 expression and tumor recurrence was observed in each tumor category (stages Ta and Tl, low and high grade). The results demonstrated that CD24 expression is Significantly associated with bladder tumor recurrence. To the best of our knowledge, this is the first study to reveal the significance of CD24 as a predictor of bladder cancer recurrence. These insights may lead to future therapeutic strategies targeting CD24 to prevent the dissemination of bladder cancer cells and tumor recurrence.

DOI： 10.3892/ol.2013.1357

Stabilization of actin bundles by a dynamin 1/cortactin ring complex is necessary for growth cone filopodia. 査読国際誌

Hiroshi Yamada, Tadashi Abe, Ayano Satoh, Nana Okazaki, Shota Tago, Kinue Kobayashi, Yumi Yoshida, Yoshiya Oda, Masami Watanabe, Kazuhito Tomizawa, Hideki Matsui, Kohji Takei

The Journal of neuroscience : the official journal of the Society for Neuroscience 33 ( 10 ) 4514 - 26 2013年3月

記述言語：英語掲載種別：研究論文（学術雑誌）

Dynamin GTPase, a key molecule in endocytosis, mechanically severs the invaginated membrane upon GTP hydrolysis. Dynamin functions also in regulating actin cytoskeleton, but the mechanisms are yet to be defined. Here we show that dynamin 1, a neuronal isoform of dynamin, and cortactin form ring complexes, which twine around F-actin bundles and stabilize them. By negative-staining EM, dynamin 1-cortactin complexes appeared as "open" or "closed" rings depending on guanine nucleotide conditions. By pyrene actin assembly assay, dynamin 1 stimulated actin assembly in mouse brain cytosol. In vitro incubation of F-actin with both dynamin 1 and cortactin led to the formation of long and thick actin bundles, on which dynamin 1 and cortactin were periodically colocalized in puncta. A depolymerization assay revealed that dynamin 1 and cortactin increased the stability of actin bundles, most prominently in the presence of GTP. In rat cortical neurons and human neuroblastoma cell line, SH-SY5Y, both dynamin 1 and cortactin localized on actin filaments and the bundles at growth cone filopodia as revealed by immunoelectron microscopy. In SH-SY5Y cell, acute inhibition of dynamin 1 by application of dynamin inhibitor led to growth cone collapse. Cortactin knockdown also reduced growth cone filopodia. Together, our results strongly suggest that dynamin 1 and cortactin ring complex mechanically stabilizes F-actin bundles in growth cone filopodia. Thus, the GTPase-dependent mechanochemical enzyme property of dynamin is commonly used both in endocytosis and regulation of F-actin bundles by a dynamin 1-cortactin complex.

DOI： 10.1523/JNEUROSCI.2762-12.2013

Dose-response analysis of testosterone replacement therapy in patients with female to male gender identity disorder 査読

Aya Nakamura, Masami Watanabe, Morito Sugimoto, Tomoko Sako, Sabina Mahmood, Haruki Kaku, Yasutomo Nasu, Kazushi Ishii, Atsushi Nagai, Hiromi Kumon

ENDOCRINE JOURNAL 60 ( 3 ) 275 - 281 2013年3月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：JAPAN ENDOCRINE SOC

Gender identity disorder (GID) is a conflict between a person's actual physical gender and the one they identify him or herself with. Testosterone is the key agent in the medical treatment of female to male GID patients. We conducted a dose-response analysis of testosterone replacement therapy (TRT) in 138 patients to determine the onset of the therapeutic effects. The TRT consisted of intramuscular injection of testosterone enanthate and patients were divided into three groups; 250 mg every two weeks, 250 mg every three weeks and 125 mg every two weeks. The onset of deepening of voice, increase in facial hair and cessation of menses was evaluated in each group. At one month after the start of TRT, the onset of these physical changes was more prevalent in the group receiving the higher dose of testosterone, and there were dose-dependent effects observed between the three treatment groups. On the other hand, at six months after the start of TRT, most of the patients had achieved treatment responses and there were no dose-dependent effects with regard to the percentage of patients with therapeutic effects. No significant side effects were observed in any of the treatment groups. We demonstrated that the early onset of the treatment effects of TRT is dose-dependent, but within six months of starting TRT, all three doses were highly effective. Current study provides useful information to determine the initial dose of TRT and to suggest possible changes that should be made in the continuous dosage for long term TRT.

DOI： 10.1507/endocrj.EJ12-0319

In Vivo Imaging of Transplanted Islets Labeled with a Novel Cationic Nanoparticle 査読

Koichi Oishi, Yoshitaka Miyamoto, Hiroaki Saito, Katsutoshi Murase, Kenji Ono, Makoto Sawada, Masami Watanabe, Yasufumi Noguchi, Toshiyoshi Fujiwara, Shuji Hayashi, Hirofumi Noguchi

PLoS ONE 8 ( 2 ) 2013年2月

記述言語：英語掲載種別：研究論文（学術雑誌）

To monitor pancreatic islet transplantation efficiency, reliable noninvasive imaging methods, such as magnetic resonance imaging (MRI) are needed. Although an efficient uptake of MRI contrast agent is required for islet cell labeling, commercially-available magnetic nanoparticles are not efficiently transduced into cells. We herein report the in vivo detection of transplanted islets labeled with a novel cationic nanoparticle that allowed for noninvasive monitoring of islet grafts in diabetic mice in real time. The positively-charged nanoparticles were transduced into a β-cell line, MIN6 cells, and into isolated islets for 1 hr. MRI showed a marked decrease in the signal intensity on T1- and T2-weighted images at the implantation site of the labeled MIN 6 cells or islets in the left kidneys of mice. These data suggest that the novel positively-charged nanoparticle could be useful to detect and monitor islet engraftment, which would greatly aid in the clinical management of islet transplant patients. © 2013 Oishi et al.

DOI： 10.1371/journal.pone.0057046

Culture conditions for mouse pancreatic stem cells. 査読

Noguchi H, Saitoh I, Kataoka HU, Watanabe M, Fujiwara T

Cell Medicine. 5 63 - 68 2013年1月

Isolation efficiency of mouse pancreatic stem cells is age-dependent. 査読

Kuise T, Noguchi H, Saitoh I, Kataoka HU, Watanabe M, Fujiwara T

Cell Medicine. 5 69 - 73 2013年1月

Cancer stem cell-like characteristics of a CD133+ subpopulation in the J82 human bladder cancer cell line. 査読国際誌

Peng Huang, Masami Watanabe, Haruki Kaku, Hideo Ueki, Hirofumi Noguchi, Morito Sugimoto, Takeshi Hirata, Hiroshi Yamada, Kohji Takei, Shaobo Zheng, Kai Xu, Yasutomo Nasu, Yasuyuki Fujii, Chunxiao Liu, Hiromi Kumon

Molecular and clinical oncology 1 ( 1 ) 180 - 184 2013年1月

記述言語：英語掲載種別：研究論文（学術雑誌）

Cancer stem cells (CSCs) are thought to be crucial for understanding the biological roots of cancer, and are of increasing importance as a target for new anticancer agents. According to an expression analysis of the cell surface antigens of various types of cancer, CD133 is considered to be a potential marker of cancer stemness. In this study, a human urinary bladder cancer cell line (J82) was used to analyze the cancer stem cell-like characteristics of CD133+ bladder cancer cells in vitro and in vivo. The CD133 expression in the J82 cells was examined and the cells were immunomagnetically categorized into positive and negative subsets. The CD133- and CD133+ subsets were phenotypically divergent with regard to the cell growth pattern, while CD133+ cells tended to colonize during their growth. In CD133+ cells, the pluripotent stem cell factors Oct-4 and Sox-2 were upregulated, and a statistically significant proliferation increase was observed when compared to CD133- cells. The CD133+ subpopulation was more tolerant to the chemotherapeutic agent cisplatin, and Bacillus Calmette-Guérin (BCG), an agent instilled intravesically to treat bladder cancer. In addition, CD133+ J82 cells were more resistant to radiation treatment when compared to CD133- cells. The in vivo tumorigenesis of the CD133- and CD133+ subsets of J82 cancer cells was also examined by subcutaneously injecting them into nude mice. The tumor growth was more aggressive in the CD133+ subpopulation, showing a significant difference in the tumorigenic potential in these subsets. In conclusion, J82 human bladder cancer cells include CD133- and CD133+ subpopulations, while the CD133 molecule is a potential marker of the potential malignancy of human bladder cancer. In the present study, the CD133+ subpopulation was herein demonstrated to have certain characteristics consistent with those of cancer stem cells.

DIRECT THERAPEUTIC EFFECT OF REIC/DKK-3-ENCODING ADENOVIRAL VECTOR FOR NON-SMALL CELL LUNG CANCER 査読

Yamamoto Hiromasa, Tanaka Norimitsu, Toyooka Shinichi, Watanabe Masami, Sakaguchi Masakiyo, Soh Junichi, Shien Kazuhiko, Furukawa Masashi, Asano Hiroaki, Tsukuda Kazunori, Nasu Yasutomo, Huh Nam-ho, Kumon Hiromi, Miyoshi Shinichirou

JOURNAL OF THORACIC ONCOLOGY 7 ( 11 ) S463 2012年11月

Preclinical biodistribution and safety study of reduced expression in immortalized cells/Dickkopf-3-encoding adenoviral vector for prostate cancer gene therapy 査読

Morito Sugimoto, Masami Watanabe, Haruki Kaku, Shun-Ai Li, Hirofumi Noguchi, Hideo Ueki, Masakiyo Sakaguchi, Nam-Ho Huh, Yasutomo Nasu, Hiromi Kumon

ONCOLOGY REPORTS 28 ( 5 ) 1645 - 1652 2012年11月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：SPANDIDOS PUBL LTD

The biodistribution and safety of adenoviral vectors encoding the human REIC/Dkk-3 tumor suppressor gene (Ad-REIC) were examined in this preclinical study for in situ prostate cancer gene therapy. First, the in vitro apoptotic effects of Ad-REIC in normal and cancer cells derived from the prostate and liver were examined. Significant apoptotic effects were observed at 100 MOI (multiplicity of infection) in prostate cancer cells (LNCaP, PC3) and hepatoma cells (HEP3B and HEPG2); however, no effects were seen in normal cells. To analyze the safety of intraprostatic Ad-REIC administration, the biodistribution and histology after Ad-REIC injection were evaluated in various organs of normal male C57BL6 mice. In a supporting study, vector dissemination following intravenous injection of Ad-REIC into tail veins was determined. To evaluate whether Ad-REIC was present in the collected tissue specimens, human REIC gene detection was performed using DNA-PCR. Intraprostatic treatment administered at lower doses showed vector biodistribution into the colon, urinary bladder and prostate. At higher doses, vector dissemination was observed in tissues more distant from the prostate, including the lung, thymus, heart, liver and adrenal gland. After intravenous injection a: Ad-REIC, dissemination was observed in the liver and spleen. These results indicate that the biodistribution of Ad-REIC is determined by the dose and route of administration. Although acute inflammatory effects were observed in the prostate after intraprostatic administration at higher doses, no abnormal histological findings were noted in the other tissues, including those of intravenously treated mice. Regarding the safety of Ad-REIC administration, no deaths and no signs of toxicity or unusual behavior were observed in the mice in any treatment group. Based on these preclinical experiments, adenovirus-mediated in situ REIC/Dkk-3 gene therapy is considered to be safe for use as a treatment for human prostate cancer.

DOI： 10.3892/or.2012.2001

REIC/Dkk-3-encoding adenoviral vector as a potentially effective therapeutic agent for bladder cancer 査読

Takeshi Hirata, Masami Watanabe, Haruki Kaku, Yasuyuki Kobayashi, Hiroshi Yamada, Masakiyo Sakaguchi, Kohji Takei, Nam-Ho Huh, Yasutomo Nasu, Hiromi Kumon

INTERNATIONAL JOURNAL OF ONCOLOGY 41 ( 2 ) 559 - 564 2012年8月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：SPANDIDOS PUBL LTD

Bladder cancer is one of the most common urogenital malignancies. The intravesical instillation of anticancer agents is an attractive strategy to treat a superficial lesion or floating/disseminated cancer cells after transurethral operation. An adenovirus carrying REIC/Dkk-3, a tumor suppressor gene (Ad-REIC), exhibits cancer-specific apoptotic effects in various types of cancer cells. The aim of the present study was to examine the potential of Ad-REIC as a therapeutic agent for bladder cancer. KK47 and RT4 human bladder cancer cells were sensitive to the Ad-REIC treatment for apoptosis induction, but some human bladder cancer cell lines (T24, J82 and TccSup) were resistant. Significant cell growth inhibition was observed when these resistant cancer cell lines were treated with Ad-REIC in a condition of floating cells, which is clinically observed after transurethral operation and becomes a cause of intravesical cancer dissemination. The therapeutic potential of Ad-REIC for the treatment of multidrug-resistant bladder cancer was investigated. The adriamycin-resistant KK47 bladder cancer cells (KK47/ADM), which also present multidrug resistance, showed induction of significant apoptosis following Ad-REIC treatment. The Ad-REIC treatment induced downregulation of P-glycoprotein in KK47/ADM cells and restored the sensitivity to doxorubicin (adriamycin). Ad-REIC suppressed P-glycoprotein expression in a c-Jun-NH2-kinase (JNK)-dependent manner. Therefore, the current study indicated two therapeutic aspects of the Ad-REIC agent against human bladder cancer cells, as an apoptosis inducer/cell growth inhibitor and as a sensitizer of chemotherapeutic agents in multidrug-resistant cancer cells. The intravesical instillation of Ad-REIC could be an attractive therapeutic method in human bladder cancer where the treatment of superficial lesions and floating/disseminated or multidrug-resistant cancer cells is necessary.

DOI： 10.3892/ijo.2012.1503

A novel gene expression system for detecting viable bladder cancer cells 査読

Hideo Ueki, Masami Watanabe, Haruki Kaku, Peng Huang, Shun-Ai Li, Kazuhiko Ochiai, Takeshi Hirata, Hirofumi Noguchi, Hiroshi Yamada, Kohji Takei, Yasutomo Nasu, Yuji Kashiwakura, Hiromi Kumon

INTERNATIONAL JOURNAL OF ONCOLOGY 41 ( 1 ) 135 - 140 2012年7月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：SPANDIDOS PUBL LTD

A novel transcriptional system was developed that can robustly enhance cancer-specific gene expression. In the system, hTERT promoter-driven gene expression was enhanced by an advanced two-step transcriptional amplification (TSTA). This construct was used to develop a novel system for detection of bladder cancer cells. The current study evaluated the advanced TSTA system by examining the cancer-specific gene transcription in various bladder cancer cell lines. The system significantly enhanced cancer-specific luciferase gene expression in the bladder cancer cell lines in comparison to the previous expression system of one-step or conventional TSTA. The fold gain of the enhancement was significantly correlated to the telomerase activity of the cell lines. A green fluorescent protein (GFP) gene encoding plasmid vector was constructed where hTERT promoter-driving transcription is enhanced by the advanced TSTA to utilize the system for the imaging and detection of viable bladder cancer cells. The advanced TSTA-hTERT-GFP plasmid successfully induced cancer-specific gene expression, showing robust GFP expression in human bladder cancer cell lines, but no visible GFP expression in normal bladder urothelial cells. The control GFP plasm id with a CMV promoter yielded GFP expression in both normal bladder cells and cancer cells. The advanced TSTA-hTERT-GFP plasmid allowed selective visualization of viable human bladder cancer cells in mixed cell culture containing 10- and 100-fold more normal bladder urothelial cells. These findings indicate that the advanced TSTA-hTERT expressional system is a valuable tool for detecting viable bladder cancer cells. The current system can be applied for in vitro detection of bladder cancer cells in urine and other types of cancer cells disseminated in vivo.

DOI： 10.3892/ijo.2012.1417

Implications of transcriptional factor, OCT-4, in human bladder malignancy and tumor recurrence 査読

Peng Huang, Jie Chen, Lei Wang, Yanqun Na, Haruki Kaku, Hideo Ueki, Katsumi Sasaki, Ken Yamaguchi, Kai Zhang, Takashi Saika, Yasutomo Nasu, Masami Watanabe, Hiromi Kumon

MEDICAL ONCOLOGY 29 ( 2 ) 829 - 834 2012年6月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：HUMANA PRESS INC

OCT-4, which is also known as POU5f1, is a key regulator of self-renewal in embryonic stem cells. The new cancer stem cell concept proposes that the expression of such genes is potentially correlated with tumorigenesis and can affect some aspects of the cancer behavior, such as recurrence or metastasis. This study investigated the association between OCT-4 expression in cancer tissues obtained by transurethral surgery and the clinical data to clarify the involvement of OCT-4 in human bladder malignancy. Immunohistochemical analysis demonstrated that a positive rate of OCT-4 expression was significantly associated with the higher-grade cancer (G2 and G3) in comparison with that of the lower grade (G1). In addition, positive OCT-4 expression was significantly associated with the intra-bladder tumor recurrence after the operation. The staining intensity of OCT-4 expression was also correlated with tumor recurrence. These data indicate that positive OCT-4 expression may be involved in the development of high-grade bladder cancer and with the bladder cancer recurrence. This is the first study showing a correlation between the expression of OCT-4 and bladder cancer recurrence. OCT-4 may be a valuable clinical marker for the progression of bladder cancer and may be an attractive therapeutic target for the development of new medicines for the treatment of malignancy.

DOI： 10.1007/s12032-011-9962-4

非小細胞肺癌に対する新たな個別化治療戦略 REIC/Dkk-3遺伝子治療の可能性

宗淳一, 豊岡伸一, 田中則光, 渡部昌実, 山本寛斉, 阪口政清, 許南浩, 那須保友, 公文裕巳, 三好新一郎

日本呼吸器外科学会雑誌 26 ( 3 ) P17 - 01 2012年4月

Partial sensitization of human bladder cancer cells to a gene-therapeutic adenovirus carrying REIC/Dkk-3 by downregulation of BRPK/PINK1 査読

Yu Jin, Hitoshi Murata, Masakiyo Sakaguchi, Ken Kataoka, Masami Watanabe, Yasutomo Nasu, Hiromi Kumon, Nam-Ho Huh

ONCOLOGY REPORTS 27 ( 3 ) 695 - 699 2012年3月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：SPANDIDOS PUBL LTD

REIC/Dkk-3 is a tumor suppressor gene that was first identified as a gene downregulated in association with immortalization of normal human fibroblasts. We have demonstrated that an adenovirus carrying REIC/Dkk-3 (Ad-REIC) showed a tumor-specific killing effect on a wide range of cancers. However, some human cancers, bladder cancers in particular, are resistant to Ad-REIC. In this study, we investigated the combination effect of downregulation of BRPK/PINK1 (PINK1) and Ad-REIC on bladder cancer cells. Five bladder cancer cell lines among six cell lines examined were resistant to Ad-REIC. Among the cell lines, the resistance of two cell lines was probably due to low infection efficiency of the adenovirus. PINK1-specific siRNA remarkably downregulated Bcl-x(L) and TRAP1 proteins and upregulated BAX protein expression. Finally, downregulation of PINK1 partially sensitized the other three cell lines that were resistant to Ad-REIC. This sensitization was associated with increasing production of reactive oxygen species (ROS). These results indicate that PINK1 is one of the key molecules for the mitochondrial protection system and that PINK1 can be a new target molecule to sensitize bladder cancer cells that are resistant to Ad-REIC.

DOI： 10.3892/or.2011.1543

Preclinical Safety and Efficacy of in Situ REIC/Dkk-3 Gene Therapy for Prostate Cancer 査読

Keiichiro Kawauchi, Masami Watanabe, Haruki Kaku, Peng Huang, Kasumi Sasaki, Masakiyo Sakaguchi, Kazuhiko Ochiai, Nam-ho Huh, Yasutomo Nasu, Hiromi Kumon

ACTA MEDICA OKAYAMA 66 ( 1 ) 7 - 16 2012年2月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：OKAYAMA UNIV MED SCHOOL

The preclinical safety and therapeutic efficacy of adenoviral vectors that express the REIC/Dkk-3 tumor suppressor gene (Ad-REIC) was examined for use in prostate cancer gene therapy. The Ad-human (h) and mouse (m) REIC were previously demonstrated to induce strong anti-cancer effects in vitro and in vivo, and we herein report the results of two in vivo studies. First, intra-tumor Ad-hREIC administration was examined for toxicity and therapeutic effects in a subcutaneous tumor model using the PC3 prostate cancer cell line. Second, intra-prostatic Ad-mREIC administration was tested for toxicity in normal mice. The whole-body and spleen weights, hematological and serum chemistry parameters, and histological evaluation of tissues from throughout the body were analyzed. Both experiments indicated that there was no significant difference in the examined parameters between the Ad-REIC-treated group and the control (PBS- or Ad-LacZ-treated) group. In the in vitro analysis using PC3 cells, a significant apoptotic effect was observed after Ad-hREIC treatment. Confirming this observation, the robust anti-tumor efficacy of Ad-hREIC was demonstrated in the in vivo subcutaneous prostate cancer model. Based on the results of these preclinical experiments, we consider the adenovirus-mediated REIC/Dkk-3 in situ gene therapy to be safe and useful for the clinical treatment of prostate cancer.

DOI： 10.18926/AMO/48076

Single Nucleotide Polymorphism WRN Leu1074Phe Is Associated with Prostate Cancer Susceptibility in Chinese Subjects 査読

Lei Wang, Haruki Kaku, Peng Huang, Kexin Xu, Kai Yang, Jiheng Zhang, Ming Li, Liping Xie, Xiaofeng Wang, Akiko Sakai, Masami Watanabe, Yasutomo Nasu, Kenji Shimizu, Hiromi Kumon, Yanqun Na

ACTA MEDICA OKAYAMA 65 ( 5 ) 315 - 323 2011年10月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：OKAYAMA UNIV MED SCHOOL

Deficiencies in the human DNA repair gene WRN are the cause of Werner syndrome, a rare autosomal recessive disorder characterized by premature aging and a predisposition to cancer. This study evaluated the association of WRN Leu1074Phe (rs1801195), a common missense single nucleotide polymorphism in WRN, with prostate cancer susceptibility in Chinese subjects. One hundred and forty-seven prostate cancer patients and 111 male cancer-free control subjects from 3 university hospitals in China were included. Blood samples were obtained from each subject, and the single nucleotide polymorphism WRN Leu1074Phe was genotyped by using a Snapshot assay. The results showed that WRN Leu1074Phe was associated with the risk of prostate cancer in Chinese men and that the TG/GG genotype displayed a decreased prevalence of prostate cancer compared with the TT genotype (OR = 0.58, 95% CI: 0.35-0.97, p = 0.039). Through stratified analysis, more significant associations were revealed for the TG/GG genotype in the subgroup with diagnosis age <= 72 yr (OR = 0.27, 95% CI: 0.12-0.61, p = 0.002) and in patients with localized diseases (OR = 0.36, 95% CI: 0.19-0.70, p = 0.003). However, no statistically significant difference was found in the subgroup with age > 72 yr or in patients with advanced diseases. We concluded that the genetic variant Leu1074Phe in the DNA repair gene WRN might play a role in the risk of prostate cancer in Chinese subjects.

DOI： 10.18926/AMO/47013

Advanced two-step transcriptional amplification as a novel method for cancer-specific gene expression and imaging 査読

Masami Watanabe, Hideo Ueki, Kazuhiko Ochiai, Peng Huang, Yasuyuki Kobayashi, Yasutomo Nasu, Katsumi Sasaki, Haruki Kaku, Yuji Kashiwakura, Hiromi Kumon

ONCOLOGY REPORTS 26 ( 4 ) 769 - 775 2011年10月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：SPANDIDOS PUBL LTD

The two-step transcriptional amplification (TSTA) system was previously reported to enhance the tissue-specific gene expression driven by weak promoters, but the enhancement of the gene expression is limited to use in in vitro and in vivo experimental situations. To achieve robust tissue-specific gene expression using the TSTA system, we developed an advanced TSTA system which includes polyglutamines and rat glucocorticoid receptor sequences between the GAL4 and VP16 sequences in the region of the first step of transcription. We evaluated the advanced TSTA system as a method to enhance the human telomerase reverse transcriptase (hTERT) promoter-driving cancer-specific transcription in various cancer cell lines. As a result, the advanced TSTA enhanced cancer-specific luciferase gene expression in all of the examined cancer cell lines, when compared with both the one-step and conventional TSTA systems (an similar to 6- and similar to 17-fold enhancement, respectively). Notably, the enhancement of the hTERT driven expression by the conventional TSTA system was modest and even inferior to the one-step system in several cancer cell lines. We then constructed a luciferase gene encoding the adeno-associated virus vector in which the hTERT promoter-mediated expression was driven by the advanced TSTA or control systems. In an orthotopic liver tumor model, mice were treated with the vector via tail vein injection. An optical imaging device was used to visualize the in vivo luciferase expression in the orthotopic tumor. The advanced TSTA system significantly enhanced the luciferase expression compared with the one-step and conventional TSTA systems (18.0+/-1.0- and 15.9+/-0.85-fold gain, respectively). Therefore, the advanced TSTA system significantly improves hTERT-dependent cancer-specific gene expression both in vitro and in vivo when compared with the previous systems. Since the advanced TSTA method can also be applied to other site-specific gene expression systems using tissue-specific promoters, this approach is expected to become a valuable tool enabling in vivo site-specific targeting in the field of gene therapy and molecular imaging.

DOI： 10.3892/or.2011.1371

Tumor growth and metastasis suppression by Glipr1 gene-modified macrophages in a metastatic prostate cancer model 査読

K. Tabata, S. Kurosaka, M. Watanabe, K. Edamura, T. Satoh, G. Yang, E. Abdelfattah, J. Wang, A. Goltsov, D. Floryk, T. C. Thompson

GENE THERAPY 18 ( 10 ) 969 - 978 2011年10月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：NATURE PUBLISHING GROUP

We previously identified the mouse and human Glipr1 and GLIPR1/RTVP-1 genes, respectively, as direct p53 targets with proapoptotic activities in various cancer cell lines, including prostate cancer (PCa). Intratumoral injection of an adenoviral vector capable of efficient transduction and expression of Glipr1 (AdGlipr1) yielded promising therapeutic results in an orthotopic, metastatic mouse model of PCa. AdGlipr1-transduced macrophages (M phi/Glipr1) generated greater surface expression of CD40, CD80 and major histocompatibility complex class II molecules and greater production of interleukin 12 (IL-12) and IL-6 in vitro than control macrophages did. Mechanistic analysis indicated that increased production of IL-12 in M phi/Glipr1 depends on activation of the p38 signaling cascade. M phi/Glipr1 injected into orthotopic 178-2BMA tumors in vivo resulted in significantly suppressed prostate tumor growth and spontaneous lung metastases and longer survival relative to those observed in control-treated mice. Furthermore, these preclinical data indicate the generation of systemic natural killer cell activity and tumor-specific cytotoxic T lymphocyte responses. Trafficking studies confirmed that intratumorally injected M phi/Glipr1 could migrate to draining lymph nodes. Overall, our data suggest that this novel gene-modified cell approach is an effective treatment avenue that induces antitumor immune responses in preclinical studies. Gene Therapy (2011) 18, 969-978; doi:10.1038/gt.2011.51; published online 21 April 2011

DOI： 10.1038/gt.2011.51

Tumor suppressor REIC/Dkk-3 interacts with the dynein light chain, Tctex-1 査読

Kazuhiko Ochiai, Masami Watanabe, Hideo Ueki, Peng Huang, Yasuyuki Fujii, Yasutomo Nasu, Hirofumi Noguchi, Takeshi Hirata, Masakiyo Sakaguchi, Nam-ho Huh, Yuji Kashiwakura, Haruki Kaku, Hiromi Kumon

BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS 412 ( 2 ) 391 - 395 2011年8月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：ACADEMIC PRESS INC ELSEVIER SCIENCE

REIC/Dkk-3 is a member of the Dickkopf family proteins known as Wnt-antagonists, and REIC/Dkk-3 expression is downregulated in a broad range of cancer types. REIC/Dkk-3 acts as a tumor suppressor in multiple cancer cell lines by inducing apoptosis through endoplasmic reticulum (ER) stress signaling. However, the intracellular interaction partners of REIC/Dkk-3 have not been fully elucidated. By employing yeast two-hybrid screening, we identified the human dynein light chain, Tctex-1, as a novel interaction partner of REIC/Dkk-3. We further disclosed that the interaction involves the 136-157 amino acid region of REIC/Dkk-3 by using the mammalian two-hybrid system. Interestingly, this binding region of REIC/Dkk-3 with Tctex-1 contains an amino acid sequence motif [-(E) under bar -X-(G) under bar-(R) under bar-(R) under bar -X-(H) under bar-] which was previously reported as the Tctex-1 binding domain of dynein intermediate chain (DIC). Immunocytochemistry demonstrated that both REIC/Dkk-3 and Tctex-1 were localized around the ER of human fibroblasts, and the similar distribution pattern of the proteins suggests that their interaction occurs around the ER. This is the first study showing the interaction of a Dickkopf family protein with a dynein motor complex protein. The link between REIC/Dkk-3 and Tctex-1 may be of significance for understanding the molecular functions of the proteins in ER stress signaling and intracellular dynein motor dynamics, respectively. (C) 2011 Elsevier Inc. All rights reserved.

DOI： 10.1016/j.bbrc.2011.07.109

[REIC/Dkk-3 gene therapy]. 査読

Masami Watanabe, Haruki Kaku, Peng Huang, Hiromi Kumon, Yasutomo Nasu

Nihon rinsho. Japanese journal of clinical medicine 69 Suppl 5 559 - 63 2011年6月

A PHASE I/II STUDY OF ADENOVIRUS-MEDIATED INTERLEUKIN-12 GENE THERAPY FOR HORMONE REFRACTORY PROSTATE CANCER; A REPORT OF INITIAL 6 CASES 査読

Sasaki Katsumi, Nasu Yasutomo, Kaku Haruki, Watanabe Masami, Edamura Kohei, Saika Takashi, Kumon Hiromi, Brenner Malcolm K

JOURNAL OF GENE MEDICINE 12 ( 12 ) 1025 2010年12月

Expression pattern of REIC/Dkk-3 in various cell types and the implications of the soluble form in prostatic acinar development 査読

Kai Zhang, Masami Watanabe, Yuji Kashiwakura, Shun-Al Li, Kohei Edamura, Peng Huang, Ken Yamaguchi, Yasutomo Nasu, Yasuyuki Kobayashi, Masakiyo Sakaguchi, Kazuhiko Ochiai, Hiroshi Yamada, Kohji Takei, Hideo Ueki, Nam-Ho Huh, Ming Li, Haruki Kaku, Yanqun Na, Hiromi Kumon

INTERNATIONAL JOURNAL OF ONCOLOGY 37 ( 6 ) 1495 - 1501 2010年12月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：SPANDIDOS PUBL LTD

The tumor suppressor REIC/Dkk-3 is a secretory protein which was originally identified to be downregulated in human immortalized cells In the present study, we investigated the expression pattern of REIC/Dkk-3 in various cell types to characterize its physiological functions We first examined the expression level of REIC/Dkk-3 in a broad range of cancer cell types and confirmed that it was significantly downregulated in all of the cell types We also examined the tissue distribution pattern in a variety of normal mouse organs Ubiquitous REIC/Dkk-3 protein expression was observed in the organs The expression was abundant in the liver, heart and brain tissue, but was absent in the spleen and peripheral blood mononuclear cells The immunohistochemical analyses revealed that the subcellular localization of REIC/Dkk-3 had a punctate pattern around the nucleus, indicating its association with secretory vesicles In cancer cells stably transfected with REIC/Dkk-3 the protein was predominantly localized to the endoplasmic reticulum (ER) under observation with confocal microscopy Because REIC/ Dkk-3 was found to be abundantly expressed in the acinar epithelial cells of the mouse prostate, we analyzed the effects of recombinant REIC/Dkk-3 protein on the acinar morphogenesis of RWPE-1 cells, which are derived from human normal prostate epithelium Statistically significant acinar growth was observed in the culture condition with 10 mu g/m1 REIC/Dkk-3 protein, implicating the soluble form m prostatic acinar development Current results suggest that REIC/Dkk-3 may play a role in regulating the morphological process of normal tissue architecture through an autocrine and/or paracrine manner

DOI： 10.3892/ijo_00000802

Potent antitumor effects of combined therapy with a telomerase-specific, replication-competent adenovirus (OBP-301) and IL-2 in a mouse model of renal cell carcinoma 査読

P. Huang, H. Kaku, J. Chen, Y. Kashiwakura, T. Saika, Y. Nasu, Y. Urata, T. Fujiwara, M. Watanabe, H. Kumon

CANCER GENE THERAPY 17 ( 7 ) 484 - 491 2010年7月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：NATURE PUBLISHING GROUP

OBP-301 (a telomerase-specific, replication-competent adenovirus with hTERT promoter) was constructed in a previous study and it showed a strong anticancer effect by inducing cell lysis in human lung and prostate cancer cells. This study investigated the effectiveness of a combination therapy of OBP-301 and interleukin-2 (IL-2) in a mouse model of renal cell carcinoma (RCC). The cell-killing effect of OBP-301 was confirmed in vitro in the RENCA cancer cells. In in vivo experiment, luciferase-expressing RENCA cells were implanted in the left kidney and lung of BALB/c mice to prepare the RCC metastatic model. The animals were randomly divided into four treatment groups: PBS, IL-2 alone, OBP-301 alone and the combination. The analyses of orthotopic tumor weight, lung metastasis and luciferin-stained tumor images 14 days after each treatment showed significant tumor growth inhibition in the combination group in comparison with that in the OBP-301- or IL-2-treated groups. In addition, the percentage of regulatory T-cells (Tregs) in the combination group was significantly suppressed in comparison with that in the PBS and single-agent treatment groups. The outcomes of this study suggest that tumor-specific oncolytic immunovirotherapy may become an attractive strategy for the treatment of human RCC. Cancer Gene Therapy (2010) 17, 484-491; doi:10.1038/cgt.2010.5; published online 19 February 2010

DOI： 10.1038/cgt.2010.5

The role of caveolin-1 in prostate cancer: clinical implications 査読

T. C. Thompson, S. A. Tahir, L. Li, M. Watanabe, K. Naruishi, G. Yang, D. Kadmon, C. J. Logothetis, P. Troncoso, C. Ren, A. Goltsov, S. Park

PROSTATE CANCER AND PROSTATIC DISEASES 13 ( 1 ) 6 - 11 2010年3月

記述言語：英語出版者・発行元：NATURE PUBLISHING GROUP

Caveolin-1 (cav-1) is reportedly overexpressed in prostate cancer cells and is associated with disease progression. Specific oncogenic activities of cav-1 associated with Akt activation also occur in prostate cancer. A membrane-associated protein, cav-1, is nonetheless secreted by prostate cancer cells; results of recent studies showed that secreted cav-1 can stimulate cell survival and angiogenic activities, defining a role for cav-1 in the prostate cancer microenvironment. Serum cav-1 levels were also higher in prostate cancer patients than in control men without prostate cancer, and the preoperative serum cav-1 concentration had prognostic potential in men undergoing radical prostatectomy. Secreted cav-1 is therefore a potential biomarker and therapeutic target for prostate cancer. Prostate Cancer and Prostatic Diseases (2010) 13, 6-11; doi:10.1038/pcan.2009.29; published online 7 July 2009

DOI： 10.1038/pcan.2009.29

PP-470 低ゴナドトロピン性男子性腺機能低下症に対するhCG、hMG療法の臨床的検討(発表・討論,第98回日本泌尿器科学会総会)

石井和史, 大石智子, 渡部昌実, 公文裕巳, 永井敦

日本泌尿器科学会雑誌 101 ( 2 ) 486 - 486 2010年

Dynasore, a dynamin inhibitor, suppresses lamellipodia formation and cancer cell invasion by destabilizing actin filaments 査読

Hiroshi Yamada, Tadashi Abe, Shun-Ai Li, Yuki Masuoka, Mihoko Isoda, Masami Watanabe, Yasutomo Nasu, Hiromi Kumon, Akira Asai, Kohji Takei

BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS 390 ( 4 ) 1142 - 1148 2009年12月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：ACADEMIC PRESS INC ELSEVIER SCIENCE

Dynamic remodeling of actin filaments are bases for a variety of cellular events including cell motility and cancer invasion, and the regulation of actin dynamics implies dynamin, well characterized endocytotic protein. Here we report that dynasore, a inhibitor of dynamin GTPase, potently destabilizes F-actin in vitro, and it severely inhibits the formation of pseudopodia and cancer cell invasion, both of which are supported by active F-actin formation. Dynasore rapidly disrupted F-actin formed in brain cytosol in vitro, and the dynasore's effect on F-actin was indirect. Dynasore significantly suppressed serum-induced lamellipodia formation in U2OS cell. Dynasore also destabilized F-actin in resting cells, which caused the retraction of the plasma membrane. A certain amount of dynamin 2 in U2OS cells localized along F-actin, and co-localized with cortactin, a physiological binding partner of dynamin and F-actin. However. these associations of dynamin were partially disrupted by dynasore treatment. Furthermore, invasion activity of H1080 cell, a lung cancer cell line, was suppressed by approximately 40% with dynasore treatment. These results strongly suggest that dynasore potently destabilizes F-actin, and the effect implies dynamin. Dynasore or its derivative would be suitable candidates as potent anti-cancer drugs. (C) 2009 Elsevier Inc. All rights reserved.

DOI： 10.1016/j.bbrc.2009.10.105

REIC/Dkk-3 stable transfection reduces the malignant phenotype of mouse prostate cancer RM9 cells 査読

Jie Chen, Masami Watanabe, Peng Huang, Masakiyo Sakaguchi, Kazuhiko Ochiai, Yasutomo Nasu, Mamoru Ouchida, Nam-Ho Huh, Kenji Shimizu, Yuji Kashiwakura, Haruki Kaku, Hiromi Kumon

INTERNATIONAL JOURNAL OF MOLECULAR MEDICINE 24 ( 6 ) 789 - 794 2009年12月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：SPANDIDOS PUBL LTD

The reduced expression in immortalized cells (REIC)/Dickkopf (Dkk)-3. a member of the Dkk gene family, is a tumor suppressor in a broad range of cancers. REIC/Dkk-3 transfected stable clones of mouse prostate cancer RM9 cells (RM9-REIC) and the empty vector-transfected control clone cells (RM9-EV) were established. Clones were used to evaluate the anti-cancer effects and a proteomics analysis of REIC/Dkk-3 continuous expression was performed. The RM9-REIC cells show a feeble appearance and the cell membrane shows irregular buds known as blebs. In vitro cell proliferation was significantly suppressed in RM9-REIC clones in comparison to the control. The apoptosis assay was done under standard culture conditions and RM9-REIC showed a higher incidence of apoptosis. The RM9-EV and RM9-REIC cells were orthotopically implanted into a C57BL/6 mouse prostate. After 2 weeks, the tumor growth was significantly inhibited in RM9-REIC cells in comparison to the control. Two-dimensional gel electrophoresis was used to examine the modification of protein expression by the gene transfection. The analysis with mass spectrometry disclosed that expression of peroxiredoxin-l, GST-P1. transgelin-2, MRP-L12, ARD, GRP78 and Sorcin were increased and eEF1A-1 and cyclophilin-40 protein were decreased in RM9-REIC cells. Therefore, REIC/Dkk-3 stable transfectants show a reduction of malignancy in mouse prostate cancer RM9 cells in vitro and in vivo. The result of the proteomics analysis might provide important clues to clarify the anti-cancer molecular mechanism of REIC/Dkk-3 gene transfer.

DOI： 10.3892/ijmm_00000293

Functional Analysis of Secreted Caveolin-1 in Mouse Models of Prostate Cancer Progression 査読

Masami Watanabe, Guang Yang, Guangwen Cao, Salahaldin A. Tahir, Koji Naruishi, Ken-ichi Tabata, Elmoataz Abdel Fattah, Kartik Rajagopalan, Terry L. Timme, Sanghee Park, Shinji Kurosaka, Kohei Edamura, Ryuta Tanimoto, Francesco J. Demayo, Alexei A. Goltsov, Timothy C. Thompson

MOLECULAR CANCER RESEARCH 7 ( 9 ) 1446 - 1455 2009年9月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：AMER ASSOC CANCER RESEARCH

Previously, we reported that caveolin-1 (cav-1) is overexpressed in metastatic prostate cancer and that virulent prostate cancer cells secrete biologically active cav-1. We also showed that cav-1 expression leads to prosurvival activities through maintenance of activated Akt and that cav-1 is taken up by other cav-1-negative tumor cells and/or endothelial cells, leading to stimulation of angiogenic activities through PI-3-K-Akt-eNOS signaling. To analyze the functional consequences of cav-1 overexpression on the development and progression of prostate cancer in vivo, we generated PBcav-1 transgenic mice. Adult male PBcav-1 mice showed significantly increased prostatic wet weight and higher incidence of epithelial hyperplasia compared with nontransgenic littermates. Increased immunostaining for cav-1, proliferative cell nuclear antigen, P-Akt, and reduced nuclear p27(KiP1) staining occurred in PBcav-1 hyperplastic prostatic lesions. PBcav-1 mice showed increased resistance to castration-induced prostatic regression and elevated serum cav-1 levels compared with nontransgenic littermates. Intraprostatic injection of androgen-sensitive, cav-1-secreting RM-9 mouse prostate cancer cells resulted in tumors that were larger in PBcav-1 mice than in nontransgenic littermates (P = 0.04). Tall vein inoculation of RM-9 cells produced significantly more experimental lung metastases in PBcav-1 males than in nontransgenic male littermates (P = 0.001), and in cav-1(+/+) mice than in cav-1(-/-) mice (P = 0.041). Combination treatment with surgical castration and systemic cav-1 antibody dramatically reduced the number of experimental metastases. These experimental data suggest a causal association of secreted cav-1 and prostate cancer growth and progression. (Mol Cancer Res 2009;7(9):1446-55)

DOI： 10.1158/1541-7786.MCR-09-0071

Therapeutic effects of gelatin matrix-embedded IL-12 gene-modified macrophages in a mouse model of residual prostate cancer 査読

K. Tabata, M. Watanabe, K. Naruishi, K. Edamura, T. Satoh, G. Yang, E. Abdel Fattah, J. Wang, A. Goltsov, D. Floryk, S. D. Soni, D. Kadmon, T. C. Thompson

PROSTATE CANCER AND PROSTATIC DISEASES 12 ( 3 ) 301 - 309 2009年9月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：NATURE PUBLISHING GROUP

We evaluated the potential use of intraoperative gelatin matrix hemostatic sealant (GMHS; FloSeal; Baxter Healthcare) embedded with macrophages (M phi) transduced with murine interleukin (IL)-12 recombinant adenoviral vector (G/M phi/AdmIL-12) for prevention of recurrence of prostate cancer following radical prostatectomy. Application of G/M phi/AdmIL-12 resulted in significant suppression of tumor growth and spontaneous lung metastases, a statistically significant survival advantage of the G/M phi/AdmIL-12-treated animals, more efficient trafficking of M phi to lymph nodes draining from the prostate and generation of systemic natural killer cell activity and tumor-specific cytolytic T lymphocyte responses compared to the controls in a preclinical mouse model of residual prostate cancer. Our data recommend this treatment as a novel adjuvant for prevention of local recurrence of prostate cancer following radical prostatectomy. Prostate Cancer and Prostatic Diseases (2009) 12, 301-309; doi: 10.1038/pcan.2008.57; published online 23 December 2008

DOI： 10.1038/pcan.2008.57

Overexpression of REIC/Dkk-3 in Normal Fibroblasts Suppresses Tumor Growth via Induction of Interleukin-7 査読

Masakiyo Sakaguchi, Ken Kataoka, Fernando Abarzua, Ryuta Tanimoto, Masami Watanabe, Hitoshi Murata, Swe Swe Than, Kaoru Kurose, Yuji Kashiwakura, Kazuhiko Ochiai, Yasutomo Nasu, Hiromi Kumon, Nam-ho Huh

JOURNAL OF BIOLOGICAL CHEMISTRY 284 ( 21 ) 14236 - 14244 2009年5月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC

We previously showed that the tumor suppressor gene REIC/Dkk-3, when overexpressed by an adenovirus (Ad-REIC), exhibited a dramatic therapeutic effect on human cancers through a mechanism triggered by endoplasmic reticulum stress. Adenovirus vectors show no target cell specificity and thus may elicit unfavorable side effects through infection of normal cells even upon intra-tumoral injection. In this study, we examined possible effects of Ad-REIC on normal cells. We found that infection of normal human fibroblasts (NHF) did not cause apoptosis but induced production of interleukin (IL)-7. The induction was triggered by endoplasmic reticulum stress and mediated through IRE1 alpha, ASK1, p38, and IRF-1. When Ad-REIC-infected NHF were transplanted in a mixture with untreated human prostate cancer cells, the growth of the cancer cells was significantly suppressed. Injection of an IL-7 antibody partially abrogated the suppressive effect of Ad-REIC-infected NHF. These results indicate that Ad-REIC has another arm against human cancer, an indirect host-mediated effect because of overproduction of IL-7 by mis-targeted NHF, in addition to its direct effect on cancer cells.

DOI： 10.1074/jbc.M808002200

Immunological aspects of REIC/Dkk-3 in monocyte differentiation and tumor regression 査読

Masami Watanabe, Yuji Kashiwakura, Peng Huang, Kazuhiko Ochiai, Junichiro Futami, Shun-Ai Li, Munenori Takaoka, Yasutomo Nasu, Masakiyo Sakaguchi, Nam-Ho Huh, Hiromi Kumon

INTERNATIONAL JOURNAL OF ONCOLOGY 34 ( 3 ) 657 - 663 2009年3月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：SPANDIDOS PUBL LTD

The REIC/Dkk-3 gene has been reported to be a tumor suppressor and the expression is significantly down-regulated in a broad range of cancer cell types. The protein is secretory, but the physiological function remains unclear. This study demonstrated that recombinant REIC/Dkk-3 protein induced the differentiation of human CD14(+) monocytes into a novel cell type ((REIC/Dkx-3)Mo). (REIC/Dkk-3)Mo resembles immature dendritic cells generated with IL-4 and GM-CSF. Both these cell populations exhibit similar proportions of CD11c(+), CD40(+), CD86(+) and HLA-DR(+) cells and endocytic capacity, but (REIC/Dkk-3)Mo is negative for CD1a antigen. An analysis of the signal transducers and activators of transcription (STAT) pathways revealed that REIC/Dkk-3 induces phosphorylation of STAT 1 and STAT 3. Furthermore, intratumoral administration of REIC/Dkk-3 protein significantly suppressed tumor growth with CD11c(+) and CD8(+) (dendritic and killer T cell marker, respectively) cell accumulation and enhanced anticancer cytolytic activity of splenocytes. These data indicated a cytokine-like role of REIC/Dkk-3 protein in monocyte differentiation that might be exploited therapeutically.

DOI： 10.3892/ijo_00000191

REIC/Dkk-3 overexpression downregulates P-glycoprotein in multidrug-resistant MCF7/ADR cells and induces apoptosis in breast cancer 査読

K. Kawasaki, M. Watanabe, M. Sakaguchi, Y. Ogasawara, K. Ochiai, Y. Nasu, H. Doihara, Y. Kashiwakura, N-h Huh, H. Kumon, H. Date

CANCER GENE THERAPY 16 ( 1 ) 65 - 72 2009年1月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：NATURE PUBLISHING GROUP

The overexpression of reduced expression in immortalized cells (REIC)/Dickkopf-3 (Dkk-3), a tumor suppressor gene, induced apoptosis in human prostatic and testicular cancer cells. The aim of this study is to examine the potential of REIC/Dkk-3 as a therapeutic target against breast cancer. First, the in vitro apoptotic effect of Ad-REIC treatment was investigated in breast cancer cell lines and the adenovirus-mediated overexpression of REIC/Dkk-3 was thus found to lead to apoptotic cell death in a c-Jun-NH(2)-kinase (JNK) phosphorylaion-dependent manner. Moreover, an in vivo apoptotic effect and MCF/Wt tumor growth inhibition were observed in the mouse model after intratumoral Ad-REIC injection. As multidrug resistance (MDR) is a major problem in the chemotherapy of progressive breast cancer, the in vitro effects of Ad-REIC treatment were investigated in terms of the sensitivity of multidrug-resistant MCF7/ADR cells to doxorubicin and of the P-glycoprotein expression. Ad-REIC treatment in MCF7/ADR cells also downregulated P-glycoprotein expresssion through JNK activation, and sensitized its drug resistance against doxorubicin. Therefore, not only apoptosis induction but also the reversal of anticancer drug resistance was achieved using Ad-REIC. We suggest that REIC/Dkk-3 is a novel target for breast cancer treatment and that Ad-REIC might be an attractive agent against drug-resistant cancer in combination with conventional antineoplastic agents.

DOI： 10.1038/cgt.2008.58

Dynamin 2 is required for actin assembly in phagocytosis in Sertoli cells 査読

Atsushi Otsuka, Tadashi Abe, Masami Watanabe, Hitoshi Yagisawa, Kohji Takei, Hiroshi Yamada

BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS 378 ( 3 ) 478 - 482 2009年1月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：ACADEMIC PRESS INC ELSEVIER SCIENCE

Dynamin 2 has been reported to be implicated in phagocytosis. However, the mode of action of dynamin is Poorly understood. In this Study, we examined whether dynamin 2 participates in actin assembly during phagocytosis in Sertoli cells. In the presence of dynasore, a dynamin inhibitor, phagocytosis was reduced by 60-70% in Sertoli cells and macrophages. Scanning electron microscopy revealed that Sertoli cells treated with dynasore were unable to form phagocytic cups. In addition, dysfunction of dynamin 2 reduced both actin polymerization and recruitment of actin and dynamin 2 to phosphatidylinositol (4,5) bisphosphate [ PI(4,5)P(2)]-containing liposomes. The formation of dynamin 2-positive ruffles of Sertoli cells was decreased by 60-70% by sequestering PI(4,5)P2 either by expression of PH domain of PLC delta or treatment with neomycin. These results strongly Suggest that dynamin 2 is involved in actin dynamics and the formation of dynamin 2-positive ruffles during phagocytosis. (C) 2008 Elsevier Inc. All rights reserved.

DOI： 10.1016/j.bbrc.2008.11.066

A comparison of proteomic profiles changes during 17β-estradiol treatment in human prostate cancer PC-3 cell line 査読

Jie Chen, Peng Huang, Haruki Kaku, Kai Zhang, Masami Watanabe, Takashi Saika, Yasutomo Nasu, Hiromi Kumon

Cancer Genomics and Proteomics 6 ( 6 ) 331 - 335 2009年

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：International Institute of Anticancer Research

Human telomerase reverse transcriptase (hTERT) is overexpressed in prostate cancer. Estrogen plays a central role in the development of prostate cancer. hTERT activity has been shown to be increased after estrogen treatment. Although significant efforts have been made to understand the role of estrogen, the telomerase connection with estrogen is poorly understood. In this report, we describe a proteomics approach for investigating the global changes in protein expression in estrogen-treated human prostate cancer PC-3 cells. PC-3 cells were seeded in medium and then treated with estrogen
the protein extract from these cells was used for two-dimensional (2D) gel electrophoresis. The protein spots were subjected to comparative analysis by liquid chromatography/mass spectrometry (LC/MS). We observed that the expression of 17 proteins, including stress-induced phosphoprotein 1 and lamin-A/C was down-regulated, and that the expression of proteins such as subunit a of T-complex protein 1, tubulin alpha-1B, and other 13 proteins was up-regulated. These proteins may have been closely associated with estrogen-induced hTERT activity. The expression level of these proteins could be a useful parameter for evaluating the estrogen-induced hTERT activity in clinical specimens of human prostate cancer.

PP-125 性同一性障害患者(FTM)に対するテストステロン補充量の検討(性機能障害,一般演題ポスター,第97回日本泌尿器科学会総会)

大石智子, 石井和史, 渡部昌実, 公文裕巳, 松本裕子

日本泌尿器科学会雑誌 100 ( 2 ) 348 - 348 2009年

Dynamin 2 Cooperates with Amphiphysin 1 in Phagocytosis in Sertoli Cells 査読

Akira Nakanishi, Tadashi Abe, Masami Watanabe, Kohji Takei, Hiroshi Yamada

ACTA MEDICA OKAYAMA 62 ( 6 ) 385 - 391 2008年12月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：OKAYAMA UNIV MED SCHOOL

Testicular Sertoli cells highly express dynamin 2 and amphiphysin 1. Here we demonstrate that dynamin 2 is implicated in phosphatidylserine (PS)-dependent phagocytosis in Sertoli cells. Immunofluorescence and dual-live imaging revealed that dynamin 2 and amphiphysin 1 accumulate simultaneously at ruffles. These proteins are specifically bound in vitro. Over-expression of dominant negative dynamin 2 (K44A) inhibits liposome-uptake and leads to the mis-localization of amphiphysin 1. Thus, the cooperative function of dynamin 2 and amphiphysin 1 in PS-dependent phagocytosis is strongly suggested.

DOI： 10.18926/AMO/30954

Down-regulation of Inhibition of Differentiation-1 via Activation of Activating Transcription Factor 3 and Smad Regulates REIC/Dickkopf-3-Induced Apoptosis 査読

Yuji Kashiwakura, Kazuhiko Ochiai, Masami Watanabe, Fernando Abarzua, Masakiyo Sakaguchi, Munenori Takaoka, Ryuta Tanimoto, Yasutomo Nasu, Nam-ho Hub, Hiromi Kumon

CANCER RESEARCH 68 ( 20 ) 8333 - 8341 2008年10月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：AMER ASSOC CANCER RESEARCH

REIC/Dickkopf-3 (Dkk-3), a tumor suppressor gene, has been investigated in gene therapy studies. Our previous study suggested that REIC/Dkk-3-induced apoptosis mainly resulted from phosphorylation of c-Jun-NH2 kinase (JNK) in prostate cancer cells. However, the precise mechanisms, especially the molecular mechanisms regulating JNK phosphorylation, remain unclear. In this study, we investigated the mechanisms participating in JNK phosphorylation in the context of a refractory cancer disease, malignant mesothelioma (MM). Adenovirus-mediated overexpression of REIC/Dkk-3 induced apoptosis mainly through JNK activation in immortalized MM cells (211H cells). Interestingly, transcriptional down-regulation of inhibition of differentiation-1 (Id-1) was detected in REIC/Dkk-3-overexpressed 211H cells. Moreover, restoration of Id-1 expression antagonized REIC/Dkk-3-induced JNK phosphorylation and apoptosis. Mutagenesis experiments with the 2.1-kb human Id-1 promoter revealed that activating transcription factor 3 (ATF3) and Smad interaction, with their respective binding motifs, was essential for REIC/Dkk-3-mediated suppression of Id-1 promoter activity. ATF3 activation was probably induced by endoplasmic reticulum stress. Finally, we showed strong antitumor effects from REIC/Dkk-3 gene transfer into the pleural cavity in an orthotopic MM mouse model. Relative to control tumor tissue, REIC/Dkk-3-treated tumor tissue showed down-regulated expression of Id-1 mRNA, enhanced expression of phosphorylated JNK, and an increased number of apoptotic cells. In summary, we first showed that both ATF3 and Smad were crucially and synergistically involved in down-regulation of Id-1, which regulated JNK phosphorylation in REIC/Dkk-3-induced apoptosis. Thus, gene therapy with REIC/Dkk-3 may be a promising therapeutic tool for MM. [Cancer Res 2008;68(20):8333-41]

DOI： 10.1158/0008-5472.CAN-08-0080

An N-terminal 78 amino acid truncation of REIC/Dkk-3 effectively induces apoptosis 査読

Fernando Abarzua, Yuji Kashiwakura, Munenori Takaoka, Masami Watanabe, Kazuhiko Ochiai, Masakiyo Sakaguchi, Takao Iwawaki, Ryuta Tanimoto, Yasutomo Nasu, Nam-ho Huh, Hiromi Kumon

BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS 375 ( 4 ) 614 - 618 2008年10月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：ACADEMIC PRESS INC ELSEVIER SCIENCE

Overexpression of REIC/Dkk-3 (a tumor suppressor gene) induces cancer cell apoptosis through endoplasmic reticulum (ER) stress. Therefore, the identification of the portion of REIC/Dkk-3 that causes ER stress may be essential for the development of cancer treatment based on REIC/Dkk-3. Here, we made several truncated forms of REIC/Dkk-3 and investigated their therapeutic potentials against prostate cancer. Among three truncated forms, a variant comprising the N-terminal 78 amino acid region of REIC/Dkk-3 ((1-78) REIC/Dkk-3) most strongly induced ER stress and apoptosis in human prostate cancer cells (PC3). For in vivo gene expression, we coupled a biodegradable polymer with naked DNA, which attained robust trans-gene expression in PC3-derived subcutaneous tumor. In therapeutic experiments, we demonstrated that multiple direct injections of polymer-conjugated (1-78)REIC/Dkk-3 plasmid provoke ER stress and significantly reduced the subcutaneous tumor volume compared with the control group. We suggest this non-viral strategy may be an effective alternative to viral gene therapy. (C) 2008 Elsevier Inc. All rights reserved.

DOI： 10.1016/j.bbrc.2008.08.079

Direct and distant antitumor effects of a telomerase-selective oncolytic adenoviral agent, OBP-301, in a mouse prostate cancer model 査読

P. Huang, M. Watanabe, H. Kaku, Y. Kashiwakura, J. Chen, T. Saika, Y. Nasu, T. Fujiwara, Y. Urata, H. Kumon

CANCER GENE THERAPY 15 ( 5 ) 315 - 322 2008年5月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：NATURE PUBLISHING GROUP

We previously constructed OBP-301 ( Telomelysin, a telomerase-specific replication-competent adenovirus with human telomerase reverse transcriptase ( hTERT) promoter), which showed a strong anticancer effect by inducing cell lysis of human non-small cell lung cancer and colorectal cancer cells. To investigate the utility of OBP-301 for prostate cancer treatment, we herein evaluate the cell killing and antitumor effects. First, in vitro hTERT-specific adenovirus transduction in human prostate cancer cells ( LNCaP, PC3, DU145) was confirmed using OBP-401 ( Telomelysin-green fluorescent protein ( GFP)). There was no detectable GFP transduction in the human prostate normal cells ( PrEC, PrSC). Consistently, the cell-killing effect of OBP-301 was observed only in the cancer cells. Second, using an in vivo subcutaneous LNCaP tumor model in nude mice, we demonstrated that three intratumoral OBP-301 injections ( 10(7) PFU per tumor x 3 days) were sufficient to eradicate the detectable LNCaP prostate tumor. We also demonstrated that the ispilateral treatment with OBP-301 significantly suppressed contralateral LNCaP tumor growth in both sides of the tumor model. Histological and immunohistochemical analyses revealed diffuse oncolytic degeneration and adenoviral E1A protein expression in both sides of the tumors. Therefore, in situ OBP-301 administration could be a promising therapeutic strategy against prostate cancer and its metastatic lesions.

DOI： 10.1038/cgt.2008.3

Adenovirus-mediated REIC/DKK-3 gene transfer prevented mesothelioma tumor progressions in orthotopic mice model 査読

Kashiwakura Yuji, Watanabe Masami, Abarzua Fernando, Sakaguchi Masakiyo, Takaoka Munenori, Tanimoto Ryuta, Nasu Yasutomo, Huh Nam-ho, Kumon Hiromi

JOURNAL OF GENE MEDICINE 10 ( 4 ) 469 - 470 2008年4月

Tumor cell-secreted caveolin-1 has proangiogenic activities in prostate cancer 査読

Salahaldin A. Tahir, Guang Yang, Alexei A. Goltsov, Masami Watanabe, Ken-ichi Tabata, Josephine Addai, El Moataz Abdel Fattah, Dov Kadmon, Timothy C. Thompson

CANCER RESEARCH 68 ( 3 ) 731 - 739 2008年2月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：AMER ASSOC CANCER RESEARCH

Caveolin, a major structural component of specialized plasma membrane invaginations (caveolae) that participate in diverse cellular activities, has been implicated in the pathogenesis of several human diseases, including cancer. We showed in earlier studies that caveolin-1 (cav-1) is consistently and strongly overexpressed in metastatic prostate cancer and is secreted in a biologically active form by virulent prostate cancer cells. Using both in vitro and in vivo model systems, we now present evidence supporting a proangiogenic role for cav-1 in prostate cancer development and progression. Recombinant cav-1 (rcav-1) was taken up by cav-1(-/-) endothelial cells through either a lipid raft/caveolae- or clathrin-dependent mechanism, leading to specific angiogenic activities (tubule formation, cell migration, and nitric oxide production) that were mediated by rcav-1 stimulation of the PI3K-Akt-eNOS signaling module. Pathologic angiogenesis induced by cav-1 in prostate cancer-bearing mice correlated with an increased frequency, number, and size of lung metastases. We propose that in addition to its antiapoptotic role, cav-1 secreted by prostate cancer cells functions critically as a proangiogenic factor in metastatic progression of this tumor. These new insights into cav-1 function in prostate cancer may provide a base for the design of clinically applicable therapeutic strategies.

DOI： 10.1158/0008-5472.CAN-07-2668

Amphiphysin 1 is important for actin polymerization During phagocytosis 査読

Hiroshi Yamada, Emiko Ohashi, Tadashi Abe, Norihiro Kusumi, Shun-AI Li, Yumi Yoshida, Masami Watanabe, Kazuhito Tomizawa, Yuji Kashiwakura, Hiromi Kumon, Hideki Matsui, Kohji Takei

MOLECULAR BIOLOGY OF THE CELL 18 ( 11 ) 4669 - 4680 2007年11月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：AMER SOC CELL BIOLOGY

Amphiphysin 1 is involved in clathrin-mediated endocytosis. In this study, we demonstrate that amphiphysin 1 is essential for cellular phagocytosis and that it is critical for actin polymerization. Phagocytosis in Sertoli cells was induced by stimulating phosphatidylserine receptors. This stimulation led to the formation of actin-rich structures, including ruffles, phagocytic cups, and phagosomes, all of which showed an accumulation of amphiphysin 1. Knocking out amphiphysin 1 by RNA interference in the cells resulted in the reduction of ruffle formation, actin polymerization, and phagocytosis. Phagocytosis was also drastically decreased in amph 1(-/-) Sertoli cells. In addition, phosphatidylinositol-4,5-bisphosphate-induced actin polymerization was decreased in the knockout testis cytosol. The addition of recombinant amphiphysin 1 to the cytosol restored the polymerization process. Ruffle formation in small interfering RNA-treated cells was recovered by the expression of constitutively active Rac1, suggesting that amphiphysin 1 functions upstream of the protein. These findings support that amphiphysin 1 is important in the regulation of actin dynamics and that it is required for phagocytosis.

DOI： 10.1091/mbc.E07-04-0296

Adenovirus-mediated REIC/Dkk-3 gene transfer inhibits tumor growth and metastasis in an orthotopic prostate cancer model 査読

K. Edamura, Y. Nasu, M. Takaishi, T. Kobayashi, F. Abarzua, M. Sakaguchi, Y. Kashiwakura, S. Ebara, T. Saika, M. Watanabe, N-H Huh, H. Kumon

CANCER GENE THERAPY 14 ( 9 ) 765 - 772 2007年9月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：NATURE PUBLISHING GROUP

We had previously reported that REIC/Dkk-3, a member of the Dickkopf ( Dkk) gene family, works as a tumor suppressor. In this study, we evaluated the therapeutic effects of an intratumoral injection with adenoviral vector encoding REIC/Dkk-3 gene ( Ad-REIC) using an orthotopic mouse prostate cancer model of RM-9 cells. We also investigated the in vivo anti-metastatic effect and in vitro anti-invasion effect of Ad-REIC gene delivery. We demonstrated that the Ad-REIC treatment inhibited prostate cancer growth and lymph node metastasis, and prolonged mice survival in the model. These therapeutic responses were consistent with the intratumoral apoptosis induction and in vitro suppression of cell invasion/migration with reduced matrix metalloprotease-2 activity. We thus concluded that in situ Ad-REIC/Dkk-3 gene transfer may be a promising therapeutic intervention modality for the treatment of prostate cancer.

DOI： 10.1038/sj.cgt.7701071

Heat shock proteins play a crucial role in tumor-specific apoptosis by REIC/Dkk-3 査読

Fernando Abarzua, Masakiyo Sakaguchi, Ryuta Tanimoto, Hiroyuki Sonegawa, Dai-Wei L, Kohei Edamura, Tomoko Kobayashi, Masami Watanabe, Yuji Kashiwakura, Haruki Kaku, Takashi Saika, Keiichiro Nakamura, Yasutomo Nasu, Hiromi Kumon, Nam-Ho Huh

INTERNATIONAL JOURNAL OF MOLECULAR MEDICINE 20 ( 1 ) 37 - 43 2007年7月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：PROFESSOR D A SPANDIDOS

We recently showed that overexpression of REIC/Dickkopf-3 (Dkk-3), a tumor suppressor gene, induced apoptosis in a tumor cell-specific manner. The aim of the present study was to determine the mechanisms underlying the selective induction of apoptosis. At first, we found a mouse renal carcinoma cell line, RENCA, to be extremely sensitive to an adenovirus carrying REIC/Dkk-3 (Ad-REIC), and we showed that activation of c-Jun N-terminal kinase (JNK) was a critical step in cell death, i.e. a process similar to that in human prostate and testicular cancer observed in our previous studies. Among the proteins interfering with the activation of JNK, heat shock protein (Hsp)70/72 was reduced in expression in RENCA cells compared with that in NIH3T3 cells. An Hsp70/72 inducer protected RENCA cells from Ad-REIC-induced apoptosis, while an Hsp70/72 inhibitor sensitized NIH3T3 cells for apoptosis induction. These results indicate that functionally active Hsp70/72 is a key factor in tumor cell-specific induction of apoptotic cell death and that analyses of the expression levels of Hsp70/72 may be essential in determining the significance of Ad-REIC-based gene therapy against human cancer.

テロメラーゼ特異的複製:前立腺癌に対する選択的ウイルス療法(Telomerase-Specific Replication: Selective Virotherapy for Prostate Cancer) 査読

黄鵬, 渡部昌実, 陳潔, 賀来春紀, 那須保友, 雑賀隆史, 藤原俊義, 浦田泰生, 公文裕巳

日本泌尿器科学会雑誌 98 ( 2 ) 458 - 458 2007年2月

Cooperative effects of adenoviral vector-mediated interleukin 12 gene therapy with radiotherapy in a preclinical model of metastatic prostate cancer 査読

T. Fujita, T. L. Timme, K. Tabata, K. Naruishi, N. Kusaka, M. Watanabe, E. Abdelfattah, J. X. Zhu, C. Ren, C. Ren, G. Yang, A. Goltsov, H. Wang, M. T. Vlachaki, B. S. Teh, E. B. Butler, T. C. Thompson

GENE THERAPY 14 ( 3 ) 227 - 236 2007年2月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：NATURE PUBLISHING GROUP

We investigated the potential benefits of combining adenoviral vector mediated in situ interleukin-12 (AdmIL-12) gene therapy with radiation therapy (XRT) to enhance therapeutic efficacy. In a metastatic mouse prostate cancer cell line, 1782 BMA, AdmIL-12+XRT demonstrated enhanced therapeutic activities in vitro as determined by clonogenic survival, apoptosis, and mIL-12 levels. At the molecular level, increased expression of tumor necrosis factor-alpha mRNA was specific for the combined therapy. In a subcutaneous 178-2 BMA in vivo model, the combination of AdmIL-12+XRT produced statistically significant tumor growth suppression compared to control vector Ad beta gal, Ad beta gal XRT, or AdmIL-12 as monotherapy. In addition, significant prolongation of survival was demonstrated for the combination of AdmIL-12+XRT. The combination of AdmIL-12+XRT significantly suppressed both spontaneous and pre-established lung metastases, and led to a prolonged elevation of serum IL-12 and significantly increased natural killer (NK) activities. Importantly, in vivo depletion of NK cells resulted in significant attenuation of the antimetastatic activities of AdmIL-12 alone or AdmIL-12+XRT. These combined effects suggest that AdIL-12 gene therapy together with radiotherapy may achieve maximal tumor control (both local and systemic) in selected prostate cancer patients via radio-gene therapy induced local cytotoxicity and local and systemic antitumor immunity.

DOI： 10.1038/sj.gt.3302788

Implication of amphiphysin 1 and dynamin 2 in tubulobulbar complex formation and spermatid release 査読

Norihiro Kusumi, Masami Watanabe, Hiroshi Yamada, Shun-Ai Li, Yuji Kashiwakura, Takashi Matsukawa, Atsushi Nagai, Yasutomo Nasu, Hiromi Kumon, Kohji Takei

CELL STRUCTURE AND FUNCTION 32 ( 2 ) 101 - 113 2007年

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：JAPAN SOC CELL BIOLOGY

Tubulobulbar complexes (TBCs) are composed of several tubular invaginations formed at the plasma membrane of testicular Sertoli cells. TBCs are transiently formed at the contact region with spermatids at spermatogenic stage VII in rat and mouse, and such TBC formation is prerequisite for spermatid release. Since the characteristic structure of TBCs suggests that the molecules implicated in endocytosis could be involved in TBC formation, we here investigated the localization and physiological roles of endocytic proteins, amphiphysin 1 and dynamin 2, at TBCs. We demonstrated by immunofluorescence that the endocytic proteins were concentrated at TBCs, where they colocalized with cytoskeletal proteins, such as actin and vinculin. Immunoelectron microscopy disclosed that both amphiphysin 1 and dynamin 2 were localized on TBC membrane. Next, we histologically examined the testis from amphiphysin 1 deficient {Amph(-/-)} mice. Morphometric analysis revealed that the number of TBCs was significantly reduced in Amph(-/-). The ratio of stage VIII seminiferous tubules was increased, and the ratio of stage IX was conversely decreased in Amph(-/-). Moreover, unreleased spermatids in stage VIII seminiferous tubules were increased in Amph(-/-), indicating that spermatid release and the following transition from stage VIII to IX was prolonged in Amph(-/-) mice. These results suggest that amphiphysin 1 and dynamin 2 are involved in TBC formation and spermatid release at Sertoli cells.

DOI： 10.1247/csf.07024

Adenoviral vector-mediated RTVP-1 gene-modified tumor cell-based vaccine suppresses the development of experimental prostate cancer 査読

K. Naruishi, T. L. Timme, N. Kusaka, T. Fujita, G. Yang, A. Goltsov, T. Satoh, X. Ji, W. Tian, E. Abdelfattah, T. Men, M. Watanabe, K. Tabata, T. C. Thompson

CANCER GENE THERAPY 13 ( 7 ) 658 - 663 2006年7月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：NATURE PUBLISHING GROUP

We previously identified a novel p53 target gene, RTVP-1, that possesses unique cytotoxic and immunostimulatory activities which make it potentially useful for cancer gene therapy. To test the therapeutic potential of RTVP-1 in a gene- modified tumor cell-based vaccine model, we used an adenoviral vector capable of efficient transduction and expression of RTVP-1 (AdRTVP-1), together with a highly metastatic mouse prostate cancer cell line (178-2 BMA). A vaccine was prepared with 178-2 BMA cells transduced with AdRTVP-1 or a control adenoviral vector expressing beta-galactosidase (Ad beta gal). After irradiation of the cells, syngeneic 129/Sv mice were vaccinated three times at weekly intervals. After 3 weeks, they were challenged with orthotopic 178-2 BMA cells. After 21 days, fewer than 60% of the RTVP-1-cell-vaccinated mice developed tumors compared to 100% of the control mice. The RTVP-1-cell vaccine significantly reduced primary tumor wet weight compared with control Ad beta gal-cell vaccine (P < 0.0001 at 7 and 14 days). Experimental metastasis to lung was also significantly reduced (P = 0.0377), and survival significantly increased (P = 0.0002). In addition, significantly increased NK and CTL activities were demonstrated in the AdRTVP-1-cell-vaccinated mice. These findings indicate that RTVP-1 gene-modified cell-based vaccines may be useful in the prevention of recurrent prostate cancer.

DOI： 10.1038/sj.cgt.7700919

Extensive biopsy using a combined transperineal and transrectal approach to improve prostate cancer detection 査読

M Watanabe, T Hayashi, T Tsushima, S Irie, T Kaneshige, H Kumon

INTERNATIONAL JOURNAL OF UROLOGY 12 ( 11 ) 959 - 963 2005年11月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：BLACKWELL PUBLISHING

Purpose: Previous studies have indicated that 6-core transrectal prostate biopsy misses a considerable number of cancers. We performed an extensive biopsy protocol of 12-core sampling using both transperineal and transrectal approaches to determine the impact on the cancer detection rate.
Materials and methods: We prospectively evaluated 402 men who underwent 6-core transperineal and 6-core transrectal biopsies simultaneously due to abnormal digital rectal examination (DRE) and/or elevated prostate-specic antigen (PSA) levels of 4.0 ng/mL or greater. Using the transperineal approach we obtained four cores from the bilateral peripheral zone targeting the lateral and parasagittal areas and two cores from the bilateral transition zone. The following transrectal biopsy was performed traditionally. We compared cancer detection rate between the extended 12-core procedure and conventional 6-core transperineal and transrectal groups in terms of total PSA and DRE ndings.
Results: Using the extensive combined method, prostate cancer was detected in 195 cases (48.5%) and the detection rate signicantly increased 7.2% and 8.5% compared to the transperineal and transrectal groups, respectively. According to PSA levels and DRE ndings, the cancer detection rate by the combined method was signicantly improved in patients with PSA levels of 4-10 ng/mL and negative DRE: 10.3% and 11.6% compared to the transperineal and transrectal groups, respectively.
Conclusions: The extensive 12-core method signicantly improved the overall cancer detection rate and was especially efcient for men with PSA levels of 4-10 ng/mL accompanied by a negative DRE nding.

DOI： 10.1111/j.1442-2042.2005.01186.x

Minimal invasiveness and effectivity of subinguinal microscopic varicocelectomy: a comparative study with retroperitoneal high and laparoscopic approaches 査読

M Watanabe, A Nagai, N Kusumi, H Tsuboi, Y Nasu, H Kumon

INTERNATIONAL JOURNAL OF UROLOGY 12 ( 10 ) 892 - 898 2005年10月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：BLACKWELL PUBLISHING

Aim: The standard management of varicocele repair is the subject of ongoing controversy. We retrospectively evaluated three surgical methods of varicocele treatment to determine the minimally invasive and most effective procedure.
Methods: We performed 144 varicocelectomies on infertile patients with left clinical varicocele. Of the patients, 50 were treated with retroperitoneal high ligation under lumbar anesthesia, 33 with laparoscopic ligation under general anesthesia, and 61 with subinguinal microscopic ligation under local anesthesia. Operative time, hospital days, and clinical outcomes were compared between these techniques.
Results: The operating time and hospitalization period required for subinguinal microscopic ligation was signicantly shorter compared to those for the other procedures. All patients treated with subinguinal microscopic ligation could achieve normal activity as soon as they returned to their rooms. Postoperative complications were observed in ve (10.0%) cases treated with high ligation and three (9.1%) laparoscopic cases, but were not observed after the subinguinal procedure. There were six cases (12.0%) of recurrence in the high ligation group and six (6.1%) in the laparoscopic group, but none in the subinguinal group. Sperm density was signicantly improved in all procedures postoperatively, but sperm motility was not improved. The two-year pregnancy rate calculated by the Kaplan-Meier method was 35.8% for high ligation, 40.4% for laparoscopic ligation and 50.9% for subinguinal microscopic ligation, although there were no statistical differences between the three groups.
Conclusion: We concluded that subinguinal microscopic varicocelectomy could be a minimally invasive procedure compared to the other two techniques and a worthy method for treating male infertility due to clinical varicocele.

DOI： 10.1111/j.1442-2042.2005.01142.x

Adeno-associated virus-mediated human IL-10 gene transfer suppresses the development of experimental autoimmune orchitis 査読

M Watanabe, Y Kashiwakura, N Kusumi, K Tamayose, Y Nasu, A Nagai, T Shimada, H Daida, H Kumon

GENE THERAPY 12 ( 14 ) 1126 - 1132 2005年7月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：NATURE PUBLISHING GROUP

Testicular germ cell-induced autoimmune orchitis is characterized by inflammatory cell infiltration followed by disturbance of spermatogenesis. Experimental autoimmune orchitis (EAO) is an animal model for human immunological male infertility; delayed-type hypersensitivity (DTH) response plays a key role in its induction. Interleukin-10 (IL-10) is a regulatory cytokine that is critical in preventing organ-specific autoimmune inflammation. To determine the effects on EAO of human IL-10 (hIL-10) gene transfer, C3H/He mice immunized by unilateral testicular injury were administered intramuscular ( i.m.) injections of adeno-associated viral (AAV) vector-encoding hIL-10 on the day of immunization. Serum hIL-10 was detected beginning at 1 week postinjection, and peaked at 3 weeks. Histological examinations showed a significantly low incidence of orchitis and disturbance of spermatogenesis in AAV hIL-10-treated mice, and the DTH response to autologous testicular cells was significantly suppressed. Immunohistochemical analysis of IFN-gamma and IL-2, T-cell-associated cytokines, in the spleen and testes revealed significantly fewer cytokine-expressing cells after treatment. We conclude that a single i.m. administration of AAV hIL-10 significantly suppresses EAO and hypospermatogenesis by regulating cell-mediated immunity in the testes.

DOI： 10.1038/sj.gt.3302463

Adeno-associated virus 2-mediated intratumoral prostate cancer gene therapy: Long-term maspin expression efficiently suppresses tumor growth 査読

M Watanabe, Y Nasu, Y Kashiwakura, N Kusumi, K Tamayose, A Nagai, T Sasano, T Shimada, H Daida, H Kumon

HUMAN GENE THERAPY 16 ( 6 ) 699 - 710 2005年6月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：MARY ANN LIEBERT INC

Maspin is a member of the serine protease inhibitors and the maspin gene, a tumor suppressor gene, is down-regulated in a large fraction of prostate cancers. We evaluated the use of adeno-associated virus (AAV, serotype 2) vector encoding maspin as a means for in vivo gene therapy for human prostate cancer. TUNEL assay of subcutaneously formed LNCaP or DU145 tumors in nude mice showed that intratumoral AAV-mediated maspin expression significantly upregulated the number of apoptotic cells compared with AAV-LacZ treatment. Immunofluorescence double staining for maspin protein and apoptosis in LNCaP tumors showed that the percentage of apoptotic cells in AAV-maspin-mediated maspin-expressing cells was significantly high compared with that in AAV-GFP-mediated GFP-expressing cells. Moreover, significantly fewer CD31-positive microvessels were observed in AAV-maspin-treated tumors compared with the control tumors. These therapeutic responses were highly correlated to persistent maspin expression in tumors, confirmed by Western blot analysis until at least day 56 after treatment. Finally, intratumoral delivery of AAV-maspin significantly suppressed growth of LNCaP and DU145 tumors and improved survival of mice. We conclude that AAV-mediated prolonged maspin expression efficiently suppresses human prostate tumor growth in vivo by apoptosis induction and inhibition of angiogenesis.

DOI： 10.1089/hum.2005.16.699

Varicocele rupture due to sexual intercourse 査読

Y Nishiyama, A Nagai, Y Nasu, M Watanabe, N Kusumi, K Monden, H Kumon

INTERNATIONAL JOURNAL OF UROLOGY 12 ( 6 ) 585 - 587 2005年6月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：BLACKWELL PUBLISHING ASIA

Varicocele rupture was diagnosed in a 23-year-old man who presented with swelling and pain in the left scrotum after sexual intercourse. Color Doppler ultrasonography revealed blood flowing into the space surrounding the left testis, a hematoma and reflux of blood in the left spermatic vein. Varicocele rupture is a very rare condition and there have been only five reported cases.

DOI： 10.1111/j.1442-2042.2005.01096.x

Clinical results of one-stage urethroplasty with parameatal foreskin flap for hypospadias 査読

A Nagai, Y Nasu, M Watanabe, N Kusumi, H Tsuboi, H Kumon

ACTA MEDICA OKAYAMA 59 ( 2 ) 45 - 48 2005年4月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：OKAYAMA UNIV MED SCHOOL

We investigated the usefulness of one-stage urethroplasty by the parameatal foreskin flap method (OUPF procedure), which is useful for repairing all types of hypospadias. Between June 1992 and March 2001, the OUPF procedure was performed on 18 patients with hypospadias: 10 patients with distal and 8 with proximal hypospadias. The follow-up periods ranged from 33-75 months, with an average of 52 months. The duration of surgery, the catheter indwelling period, and the postoperative complications of each patient were analyzed. The median age of the patients at the time of surgery was 3 years and 8 months. The length of surgery for OUPF II ranged from 150-230 min (average 186 min), and from 190-365 min (average 267 min) for OUPF IV. Postoperative complications were confirmed in 3 of the 18 patients (16.6%). Two patients had fistulas, and one had a meatal regression. The fistulas were successfully closed by the simple multilayered closure method. After adopting DuoDerm dressings instead of elastic bandages for protection of the wound, no fistulization occurred. DuoDerm dressings are useful in the healing of wounds without complications. To date, the longest follow-up period has been 75 months, and during that time there have been no late complications such as urethral stenosis or penile curvature. OUPF is a useful method in the treatment of hypospadias with a low incidence of early and late complications.

DOI： 10.18926/AMO/31964

Analysis of human ejaculation using color Doppler ultrasonography: A comparison between antegrade and retrograde ejaculation 査読

A Nagai, M Watanabe, Y Nasu, H Iguchi, N Kusumi, H Kumon

UROLOGY 65 ( 2 ) 365 - 368 2005年2月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：ELSEVIER SCIENCE INC

Objectives. To observe the phenomenon of human ejaculation dynamically using color Doppler ultrasonography.
Methods. Human ejaculation was observed using transrectal color Doppler ultrasonography in a healthy man and a patient with retrograde ejaculation. Ejaculation was induced manually with audiovisual sexual stimulation. The ejaculatory phenomenon was analyzed and compared with that of retrograde ejaculation.
Results. In the healthy man, the prostatic urethra flattened slightly and the bladder neck contracted just before expulsion. The ejaculatory stream spurted from the seminal vesicles to the bulbous urethra through the ejaculatory duct. In the patient with retrograde ejaculation, the ejaculatory stream from the seminal vesicles and inframontanal and distal prostatic urethras distended into a globular-shaped sac filled with semen. No seminal flow toward the bulbous urethra occurred. The semen remaining in the prostatic urethra began flowing slowly into the bladder.
Conclusions. Differences between antegrade and retrograde ejaculation can be clearly detected by color Doppler ultrasonography, providing a noninvasive method to diagnose ejaculatory disorders. UROLOGY 65: 365-368, 2005. (C) 2005 Elsevier Inc.

DOI： 10.1016/j.urology.2004.09.016

Hepatocyte growth factor receptor is a coreceptor for adeno-associated virus type 2 infection 査読

Y Kashiwakura, K Tamayose, K Iwabuchi, Y Hirai, T Shimada, K Matsumoto, T Nakamura, M Watanabe, K Oshimi, H Daida

JOURNAL OF VIROLOGY 79 ( 1 ) 609 - 614 2005年1月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：AMER SOC MICROBIOLOGY

After the first attachment of virus to the cell surface through a primary receptor, efficient entry of virus requires the presence of a coreceptor. For adeno-associated virus type 2 (AAV2) infection, heparan sulfate proteoglycan is supposed as the primary receptor, and alphavbeta5 integrin and FGFR1 are reported to act as coreceptors. In this study, we were able to demonstrate that hepatocyte growth factor receptor, e-Met, is also a coreceptor for AAV2 infection. AAV2-mediated transgene analyses revealed that c-Met expression significantly up-regulated transgene expression without increasing AAV2 cell binding. Moreover, a viral overlay assay elucidated the physical interaction between AAV2 and the beta subunit of c-Met. These data suggest that c-Met plays the role of coreceptor for AAV2 infection by facilitating AAV2 internalization into the cytoplasm.

DOI： 10.1128/JVI.79.1.609-614.2005

マウス精巣針生検後の造精機能および精細胞に対する遅延型過敏反応の解析(第93回日本泌尿器科学会総会)

坪井啓, 渡部昌実, 久住倫宏, 永井敦, 公文裕巳

日本泌尿器科学会雑誌 96 ( 2 ) 186 - 186 2005年

Cytosolic Ca2+ responses to sub-picomolar and nanomolar PACAP in pancreatic beta-cells are mediated by VPAC2 and PAC1 receptors 査読

H Yamada, M Watanabe, T Yada

REGULATORY PEPTIDES 123 ( 1-3 ) 147 - 153 2004年12月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：ELSEVIER SCIENCE BV

Pituitary adenylate cyclase-activating polypeptide (PACAP) potentiates glucose-induced insulin release and increases cytosolic Ca2+ concentration ([Ca2+](i)) in islet beta-cells in a concentration-dependent manner with two peaks at 10(-13) and 10(-9) M. PAC1 receptor (PAC1-R) and VPAC2 receptor (VPAC2-R) are expressed in pancreatic beta-cells and thought to be involved in insulin release. We aimed to determine the receptor types involved in the [Ca-2](i) responses to 10(-13) and 10(-9) M PACAP. We measured [Ca2+](i) in beta-cells and examined comparative effects of PAC1-R-selective agonist maxadilan, its antagonist M65, VPAC2-R-selective agonist Ro25-1553, and native ligands of PACAP and VIP. In the presence of 8.3 mM glucose, maxadilan, Ro25-1553, PACAP, and VIP at 10(-13) and 10(-9) M all increased [Ca2+](i). PACAP and maxadilan elicited greater effects at 10(-9) M than at 10(-13) M both in the incidence and amplitude of [Ca2+](i) responses. For VIP and Ro25-1553, in contrast, the effects at 10(-9) and 10(-13) M were comparable. Furthermore, the amplitude of [Ca2+](i) responses to 10(-9) M PACAP, but not 10(-13) M PACAP, was suppressed by M65. The results suggest that VPAC2-R and PAC1-R contribute equally to [Ca2+](i) responses to sub-picomolar concentrations of PACAP, while PAC1-R has greater contribution to [Ca2+](i) responses to nanomolar concentrations of this peptide. (C) 2004 Elsevier B.V. All rights reserved.

DOI： 10.1016/j.regpep.2004.03.020

Immunohistochemical localization of Klotho protein in brain, kidney, and reproductive organs of mice 査読

SA Li, M Watanabe, H Yamada, A Nagai, M Kinuta, K Takei

CELL STRUCTURE AND FUNCTION 29 ( 4 ) 91 - 99 2004年12月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：JAPAN SOC CELL BIOLOGY

Klotho mutant mouse (kl-/-), a mouse model for human aging. exhibits various phenotypes in a wide range of organs including arteriosclerosis, neural degeneration. skin and gonadal atrophy, pulmonary emphysema, calcification of soft tissues, and cognition impairment. Klotho mRNA. however. is expressed only in brain, kidney, reproductive organs, pituitary gland, and parathyroid gland. Therefore it remains to be elucidated how lack of Klotho protein in these limited organs leads to the variery of phenotypes. To shed light on mechanisms by which Klotho protein acts on distant targets, we examined localization of Klotho protein in brain. kidney. and reproductive organs, and analyzed brain and kidney in kl-/- mice searching for changes in target regions in these organs. In brain, Klotho proteins were localized at choroid plexus, where the proteins were dominantly localized at the apical plasma membrane of ependymal cells. In kl-/- brain, reduction of synapses was evident in hippocampus, suggesting a role of Klotho as a humoral factor in cerebrospinal fluid. Klotho proteins in kidney localized at distal renal tubules. Interestingly, in kl-/- mice, type IIa Na/phosphate (Pi) cotransporters, which function at the proximal renal tubules in reabsorption of phosphate ions. were translocated. This suggests that Klotho protein in kidney is implicated in calcium homeostasis which regulates localization of type IIa Na/Pi cotransporters via parathyroid hormone (PTH). Klotho proteins in reproductive organs were expressed only in mature germ cells, although in kl-/- mice germ cell maturation was arrested at earlier stages. Thus, Klotho proteins not only function as a humoral factor, but also are implicated in hormonal regulation. which may explain why mutation of klotho gene results in a variety of phenotypes.

DOI： 10.1247/csf.29.91

Dynamin-2 regulates oxidized low-density lipoprotein-induced apoptosis of vascular smooth muscle cell 査読

Y Kashiwakura, M Watanabe, N Kusumi, K Sumiyoshi, Y Nasu, H Yamada, T Sawamura, H Kumon, K Takei, H Daida

CIRCULATION 110 ( 21 ) 3329 - 3334 2004年11月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：LIPPINCOTT WILLIAMS & WILKINS

Background-On exposure to oxidized low-density lipoprotein (oxLDL), vascular cells generally undergo apoptosis, which is one of the major pathogenic factors of atherosclerosis. In this study, we examined the role of dynamin ( a crucial GTPase protein in endocytosis) in oxLDL-induced apoptosis of vascular smooth muscle cells (VSMC).
Methods and Results-After oxLDL stimulation, dynamin-2 colocalized with LOX-1 around the cell surface, as well as oxLDL in the cytoplasm, suggesting that dynamin-2 was involved in scavenger receptor-mediated oxLDL endocytosis. Downregulation of dynamin-2 induced by dynamin-2 dominant negative plasmid (K44A) resulted in a decrease of oxLDL uptake and thereby in a reduction of apoptosis. These data demonstrated that dynamin-2 was involved in oxLDL-induced apoptosis via the oxLDL endocytotic pathway. On the other hand, dynamin-2 wild-type plasmid transfection promoted oxLDL-induced apoptosis without increasing oxLDL uptake. Interestingly, the p53 inhibitor pifithrin-alpha (PFT) significantly reduced apoptosis promoted by wild-type dynamin-2 (78% reduction compared with the PFT[-] condition). These results indicated that dynamin-2 enhanced oxLDL-induced apoptosis of VSMC by participating in the p53 pathway, probably as a signal transducer. Moreover, we demonstrated that, in advanced plaques of apolipoprotein E-/- mice, dynamin-2 expression was often enhanced in apoptotic VSMC, suggesting that dynamin-2 might participate in apoptosis of VSMC even in vivo.
Conclusions - Our data demonstrated that dynamin-2 at least partially regulated oxLDL-induced apoptosis of VSMC by participating in 2 independent pathways: the oxLDL endocytotic pathway and the p53 pathway. These findings suggest that dynamin-2 may serve as a new research or therapeutic target in vascular disease.

DOI： 10.1161/01.CIR.0000147828.86593.85

Analysis of retrograde ejaculation using color Doppler ultrasonography before and after transurethral collagen injection 査読

A Nagai, Y Nasu, M Watanabe, M Tsugawa, H Iguchi, H Kumon

INTERNATIONAL JOURNAL OF IMPOTENCE RESEARCH 16 ( 5 ) 456 - 458 2004年10月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：NATURE PUBLISHING GROUP

Transurethral bladder neck collagen injection therapy was performed in a patient with retrograde ejaculation. The phenomenon of retrograde ejaculation and its correction after the therapy were clearly demonstrated by color Doppler ultrasonography. To our knowledge this is the first report showing successful observation of retrograde ejaculation using color Doppler ultrasonography.

DOI： 10.1038/sj.ijir.3901202

PP3-132 経会陰的および経直腸的前立腺12ヶ所生検の有用性 : 再生検施行症例の検討(一般演題(ポスター))

秋山博伸, 林俊秀, 枝村康平, 渡部昌実, 牧佳男, 入江伸, 金重哲三

日本泌尿器科学会雑誌 95 ( 2 ) 506 - 506 2004年

経会陰的および経直腸的前立腺12ヵ所生検の有用性 : 前立腺全摘標本での癌mappingをふまえて

渡部昌実, 林俊秀, 枝村康平, 森末浩一, 入江伸, 金重哲三

日本泌尿器科学会雑誌 94 ( 2 ) 145 - 145 2003年

上部尿路結石に対するSiemens Lithostar Multilineを用いたESWLの治療経験

枝村康平, 林俊秀, 渡部昌実, 森末浩一, 入江伸, 金重哲三

西日本泌尿器科 64 ( 増刊 ) 190 - 190 2002年11月

Localization of dynamin 2 in rat seminiferous tubules during the spermatogenic cycle 査読

H Iguchi, M Watanabe, A Kamitani, A Nagai, O Hosoya, K Tsutsui, H Kumon

ACTA MEDICA OKAYAMA 56 ( 4 ) 205 - 209 2002年8月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：OKAYAMA UNIV MED SCHOOL

Dynamin is a protein essential to endocytosis. Dynamin 2, a dynamin isoform, is expressed most intensely in testicular tissue; however, precise localization has never been studied. Therefore, we investigated the expression of dynamin 2 in rat testicular tissue using immunohistochemical methods, and discuss here the physiological function of this protein. Testicular tissues were obtained from Wistar rats at 10, 21 and 63 days of age. Immunohistochemistrical examination and Western blot analysis were conducted using dynamin 2 specific antibody. Western blot analysis showed that expression in 21- and 63-day-old rats was more intense than that in 10-day-old rats. Dynamin 2 expression was observed using immunohistochemical method in the seminiferous tubules of all rats. In the 63-day-old rats, the expression was intense, especially in spermatids in the earlier maturation stages and in spermatocytes, and was observed in Sertoli cells. However, in spermatids, the expression gradually declined as spermatids matured to spermatozoa. In the 21-day-old rats, the expression was evident in spermatocytes and Sertoli cells, but that in the 10-day-old rats was weak. Intense expression of dynamin 2 during spermatogenesis suggests that this protein plays an important role in this process.

DOI： 10.18926/AMO/31681

Distribution of dynamins in testis and their possible relation to spermatogenesis 査読

A Kamitani, H Yamada, M Kinuta, M Watanabe, SA Li, T Matsukawa, M McNiven, H Kumon, K Takei

BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS 294 ( 2 ) 261 - 267 2002年6月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：ACADEMIC PRESS INC ELSEVIER SCIENCE

Dynamin 2 and dynamin 3 are highly expressed in testis. However, their functions in the tissue remain unclear. Considering that dynamin 1, neuron-specific isoform of dynamin, plays a pivotal role in endocytosis, functions of dynamin 2 and dynamin 3 in testis must be essential. Cellular expression and subcellular localization of dynamin 2 and dynamin 3 in testis were investigated. Dynamin 2 and dynainin 3 were highly expressed in germ cells and Sertoli cells, constituents of seminiferous tubules. By immunofluorescence it was revealed that dynamin 2 colocalizes with clathrin both at the plasmamembrane and at Golgi in a cell line of Sertoli cells. Immuno reactivity for dynamin 3, on the other hand, appeared as finer puncta, which did not colocalize with clathrin, suggesting that these two dynamins have distinct functions in Sertoli cells. In the klotho deficient mouse testis, which demonstrates disorder in spermatogenesis, expression of dynamin 2 and dynamin 3 was drastically reduced indicating possible association of these proteins with spermatogenesis. (C) 2002 Elsevier Science (USA). All rights reserved.

DOI： 10.1016/S0006-291X(02)00470-9

Norepinephrine triggers Ca2+-dependent exocytosis of 5-hydroxytryptamine from rat pinealocytes in culture 査読

H Yamada, M Hayashi, S Uehara, M Kinoshita, A Muroyama, M Watanabe, K Takei, Y Moriyama

JOURNAL OF NEUROCHEMISTRY 81 ( 3 ) 533 - 540 2002年5月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：BLACKWELL PUBLISHING LTD

5-Hydroxytryptamine (5-HT) is a precursor and a putative modulator for melatonin synthesis in mammalian pinealocytes. 5-HT is present in organelles distinct from L-glutamate-containing synaptic-like microvesicles as well as in the cytoplasm of pinealocytes, and is secreted upon stimulation by norepinephrine (NE) to enhance serotonin N-acetyltransferase activity via the 5-HT2 receptor. However, the mechanism underlying the secretion of 5-HT from pinealocytes is unknown. In this study, we show that NE-evoked release of 5-HT is largely dependent on Ca2+ in rat pinealocytes in culture. Omission of Ca2+ from the medium and incubation of pineal cells with EGTA-tetraacetoxymethyl-ester inhibited by 59 and 97% the NE-evoked 5-HT release, respectively. Phenylephrine also triggered the Ca2+-dependent release of 5-HT, which was blocked by phentolamine, an alpha antagonist, but not by propranolol, a beta antagonist. Botulinum neurotoxin type E cleaved 25 kDa synaptosomal-associated protein and inhibited by 50% of the NE-evoked 5-HT release. Bafilomycin A1, an inhibitor of vacuolar H+-ATPase, and reserpine and tetrabenazine, inhibitors of vesicular monoamine transporter, all decreased the storage of vesicular 5-HT followed by inhibition of the NE-evoked 5-HT release. Agents that trigger L-glutamte exocytosis such as acetylcholine did not trigger any Ca2+-dependent 5-HT release. Vice versa neither NE nor phenylephrine caused synaptic-like microvesicle-mediated L-glutamate exocytosis. These results indicated that upon stimulation of a adrenoceptors pinealocytes secrete 5-HT through a Ca-2-dependent exocytotic mechanism, which is distinct from the exocytosis of synaptic-like microvesicles.

DOI： 10.1046/j.1471-4159.2002.00839.x

Phosphatidylinositol 4,5-bisphosphate stimulates vesicle formation from liposomes by brain cytosol 査読

M Kinuta, H Yamada, T Abe, M Watanabe, SA Li, A Kamitani, T Yasuda, T Matsukawa, H Kumon, K Takei

PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA 99 ( 5 ) 2842 - 2847 2002年3月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：NATL ACAD SCIENCES

As a step toward the elucidation of mechanisms in vesicle budding, a cell-free assay that measures cytosol-induced vesicle generation from liposomes was established. This assay then was used to explore the role of phosphoinositides in vesicle formation. Liposomes incubated with brain cytosol in the presence of ATP and GTP massively generated small vesicles, as assessed both quantitatively and qualitatively by a dynamic light-scattering assay. Both ATP and GTP were required. Vesicle formation was inhibited greatly by the immunodepletion of dynamin 1 from the cytosol, indicating a major contribution of this GTPase in this reaction and suggesting that it mimics endocytic vesicle fission. Increasing the concentration Of L-alpha-phosphatidylinositol 4,5-bisphosphate [PtdIns(4,5)P-2] but not of L-alpha-phosphatidylinositol 4-monophosphate or L-alpha-phosphatidylinositol in the lipid membranes enhanced vesicle formation. Lipid analysis revealed rapid degradation of PtdIns(4,5)P-2 to L-alpha-phosphatidylinositol during the incubation with the reaction reaching a maximum within 5 sec, whereas vesicle formation proceeded with a longer time course. PtdIns(4,5)P-2 degradation was independent of vesicle formation and occurred also in the presence of guanosine 5'-O-(thiotriphosphate), where few vesicle formations occurred. These results suggest that PtdIns(4,5)P-2 plays a critical role in the early step of vesicle formation, possibly in the recruitment of coats and fission factors to membranes.

DOI： 10.1073/pnas.261715599

Primary gastric T-cell lymphoma without human T-lymphotropic virus type 1: Report of a case 査読

M Watanabe, Y Moriyama

SURGERY TODAY 32 ( 6 ) 525 - 530 2002年

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：SPRINGER-VERLAG

Although primary gastric malignant lymphoma accounts for slightly more than 10% of all lymphomas at extranodular sites, it is relatively rare clinically, representing only 1% of all malignant diseases of the stomach. In addition, most such diseases tend to be B-cell lymphoma, while T-cell lymphoma is extremely rare. We encountered a patient with primary gastric T-cell malignant lymphoma who, although demonstrating a very rare phenomenon, was negative for antihuman T-lymphotropic virus type 1 antibody. A 73-year-old man was admitted to the hospital with the chief complaint of upper abdominal pain. The primary lesion was a type 3 tumor located at the cardia to the posterior wall of the upper body of the stomach, which had invaded the tail of the pancreas and a part of the transverse colon. A total gastrectomy, pancreatosplenectomy, and partial resection of the transverse colon were performed. The surgical section contained a giant ulcerative lesion with its bank cleaved, and a histological examination revealed a diffuse, small cell (Lymphoma Study Group classification) malignant lymphoma. An immunohistochemical analysis of the surgical specimen was positive for LCA/CD45, UCLH-1/CD45RO, and Leu-4/CD3, and negative for L-26/CD20, and it was diagnosed to be primary gastric T-cell malignant lymphoma.

ヒト射精現象の超音波カラードプラによる解析

永井敦, 井口裕樹, 紙谷章弘, 渡部昌実, 公文裕巳

日本泌尿器科学会雑誌 93 ( 2 ) 414 - 414 2002年

Identification and Characterization of a Synaptojanin 2 Splice Isoform Predominantly Expressed in Nerve Terminals 査読

Yasuo Nemoto, Markus R. Wenk, Masami Watanabe, Laurie Daniell, Tomoe Murakami, Niels Ringstad, Hiroshi Yamada, Kohji Takei, Pietro De Camilli

Journal of Biological Chemistry 276 ( 44 ) 41133 - 41142 2001年11月

記述言語：英語掲載種別：研究論文（学術雑誌）

We have previously identified synaptojanin 1, a phosphoinositide phosphatase predominantly expressed in the nervous system, and synaptojanin 2, a broadly expressed isoform. Synaptojanin 1 is concentrated in nerve terminals, where it has been implicated in synaptic vesicle recycling and actin function. Synaptojanin 2A is targeted to mitochondria via a PDZ domain-mediated interaction. We have now characterized an alternatively spliced form of synaptojanin 2 that shares several properties with synaptojanin 1. This isoform, synaptojanin 2B, undergoes further alternative splicing to generate synaptojanin 2B1 and 2B2. Both amphiphysin and endophilin, two partners synaptojanin 1, bind synaptojanin 2B2, whereas only amphiphysin binds synaptojanin 2B1. Sequence similar to the endophilin-binding site in synaptojanin 1 is present only in synaptojanin 2B2, and this sequence was capable of affinity purifying endophilin from rat brain. The Sac1 domain of synaptojanin 2 exhibited phosphoinositide phosphatase activity very similar to that of the Sac1 domain of synaptojanin 1. Site-directed mutagenesis further illustrated its functional similarity to the catalytic domain of Sac1 proteins. Antibodies raised against the synaptojanin 2B-specific carboxyl-terminal region identified a 160-kDa protein in brain and testis. Immunofluorescence showed that synaptojanin 2B is localized at nerve terminals in brain and at the spermatid manchette in testis. Active Rac1 GTPase affects the intracellular localization of synaptojanin 2, but not of synaptojanin 1. These results suggest that synaptojanin 2B has a partially overlapping function with synaptojanin 1 in nerve terminals, with additional roles in neurons and other cells including spermatids.

DOI： 10.1074/jbc.M106404200

Expression of amphiphysin I in sertoli cells and its implication in spermatogenesis 査読

M Watanabe, K Tsutsui, O Hosoya, K Tsutsui, H Kumon, A Tokunaga

BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS 287 ( 3 ) 739 - 745 2001年9月

記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：ACADEMIC PRESS INC ELSEVIER SCIENCE

Amphiphysin I is a protein concentrated in nerve terminals and involved in the endocytosis of synaptic vesicle membrane. We show here that amphiphysin I is expressed in the rat testis, localized exclusively in the Sertoli cells. In the postnatal testicular development, expression of amphiphysin I was not evident at birth, but became significant at postnatal day 15 (P15), coinciding with the onset of spermatogenesis. The expression level of amphiphysin I increased 10-fold between P15 and P25 to reach the adult level. In adult testes reversibly damaged by ethane dimethane sulphonate administration, expression of amphiphysin I did not change following the damage, whereas the protein was transiently converted into its phosphorylated form. The increase in levels of phosphorylated amphiphysin I was closely associated with the severe histological damage to germ cells. The present findings suggest that amphiphysin I in Sertoli cells is involved in spermatogenesis, probably through endocytic processes. (C) 2001 Academic Press.

DOI： 10.1006/bbrc.2001.5650

Sertoli cellで発現するシナプスタンパク質Amphiphysin Iと精子形成

渡部昌実, 永井敦, 井口裕樹, 公文裕巳, 筒井公子, 細谷修, 徳永叡, 筒井研

日本泌尿器科学会雑誌 91 ( 3 ) 372 - 372 2000年

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