研究者詳細 - 沢　禎彦

論文 - 沢　禎彦

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Quantitative evaluation of myofibroblast apoptosis during wound healing in rat palate after post-operative administration of basic fibroblast growth factor (bFGF). 国際誌

Yuichiro Hata, Hiroyuki Ishikawa, Takeshi Ueki, Takashi S Kajii, Sachio Tamaoki, Eichi Tsuruga, Yoshihiko Sawa, Kunihisa Taniguchi

Acta odontologica Scandinavica 71 ( 6 ) 1501 - 7 2013年11月

記述言語：英語掲載種別：研究論文（学術雑誌）

OBJECTIVE: Excessive wound contraction apparently inhibits maxillary growth; thus, myofibroblast apoptosis needs to be accelerated in mucoperiosteal denudation after palatoplasty. The aim of this study was to evaluate myofibroblast apoptosis during wound healing in mucoperiosteal denudation of rat palates immediately after post-operative administration of basic fibroblast growth factor (bFGF). MATERIALS AND METHODS: A total of 100 male Wistar rats aged 20 days were divided into control, scar, sham and bFGF groups (n = 25 each). In the scar, sham and bFGF groups, mucoperiosteum was removed from the palate and fibrin glue was applied to the exposed bone surface immediately after surgery. In the bFGF group, 10 μL of 2 μg/μL bFGF solution was injected into the operated area beneath the fibrin glue. At 2, 5, 7, 14 and 28 days post-operatively, myofibroblast apoptosis during the wound healing process was investigated by double immunofluorescence staining. The apoptotic area of myofibroblasts was measured using image software. RESULTS: In the bFGF group, at 2 days, apoptosis of myofibroblasts in the lamina propria and submucosa was marked, as compared with the other three groups and apoptosis of myofibroblasts was scarcely seen at 5 days. At 5 and 7 days, the apoptotic area of myofibroblasts in the bFGF group was statistically significantly smaller when compared to the scar and sham groups. CONCLUSION: The results confirmed that bFGF injection immediately after surgery accelerated apoptosis of myofibroblasts in mucoperiosteal denudation of rats. This may reduce maxillary growth retardation due to excessive wound contraction.

DOI： 10.3109/00016357.2013.773370

Altered dynamics in the renal lymphatic circulation of type 1 and type 2 diabetic mice.

Takanobu Uchiyama, Shunsuke Takata, Hiroyuki Ishikawa, Yoshihiko Sawa

Acta histochemica et cytochemica 46 ( 2 ) 97 - 104 2013年4月

記述言語：英語掲載種別：研究論文（学術雑誌）

The dynamics of the renal lymphatic circulation in diabetic nephropathy is not fully elucidated. The present study evaluated the effect of diabetic nephropathy on the renal lymphatic circulation in streptozotocin (STZ)-induced type 1 diabetic mice (ICR-STZ) and in type 2 diabetic KK/Ta mice which were fed a high fat diet (KK/Ta-HF). The diabetic mouse kidneys developed edema because of the nephropathy. In control mice renal lymphatic vessels distributed in the cortex but rarely in the medulla while in ICR-STZ and KK/Ta-HF mice, there were many lymphatic vessels with small lumen in both cortex and medulla. Total numbers and areas of renal blood vessels in the diabetic mice were similar to those in the controls while the total numbers and areas of renal lymphatic vessels were larger in diabetic mice than in the controls. There were statistically significant differences in the numbers of lymphatic vessels with diameters of 50-100 µm between the ICR-STZ and the control ICR mice, and in the numbers of lymphatic capillaries with diameters smaller than 50 µm between the KK/Ta-HF and the control KK/Ta mice. The diabetic nephropathy may induce the lymphangiogenesis or result in at least the renal lymphatic vessel expansion.

DOI： 10.1267/ahc.13006

Immunohistochemical Study of Lysosome-Associated Membrane Proteins During Periodontal Ligament Development 査読

Yuji Hatakeyama, Junko Hatakeyama, Kyoko Oka, Eichi Tsuruga, Tetsuichiro Inai, Yoshihiko Sawa

JOURNAL OF HARD TISSUE BIOLOGY 22 ( 2 ) 233 - 239 2013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：JOURNAL HARD TISSUE BIOLOGY

Amelogenin is an enamel matrix protein also expressed in periodontal tissues. Recent studies suggest that lysosome-associated membrane proteins (LAMPs) can bind cell surface amelogenin and act as signaling receptors for amelogenin in periodontal ligament (PDL) cells and cementoblasts in vitro. However, whether amelogenin and LAMPs are involved in the development of periodontal tissues is unclear. To address this issue, we used immunohistochemistry to profile LAMPs and amelogenin expression during periodontal tissue development. Paraffin sections from mice at embryonic day 15 (E15) and 1, 2, and 4 weeks were made, and serial sections at each age were stained with hematoxylin-eosin, followed by immunostaining with anti-amelogenin, LAMP-1, and LAMP-3. At El 5, dental follicle cells, which are the source of periodontal tissues, were amelogenin-positive and weakly positive for LAMP-1 and LAMP-3. At 1 week, dental follicle cells were immunopositive for amelogenin and LAMP-1 but weakly positive for LAMP-3. At 2 weeks, PDL cells were positive for amelogenin and LAMP-1 but negative for LAMP-3. At 4 weeks, PDL cells were immunopositive for both amelogenin and LAMP-1. These results suggest that LAMP-1 is expressed in both the dental follicle and PDL, and that LAMP-1 and LAMP-3 have temporally distinct roles as signaling receptors for amelogenin.

Case of autoimmune hepatitis with markedly enlarged hepatoduodenal ligament lymph nodes. 国際誌

Hideki Fujii, Naoki Ohnishi, Kazuho Shimura, Masafumi Sakamoto, Tohru Ohkawara, Yoshihiko Sawa, Koichi Nishida, Yasuo Ohkawara, Tatsuro Kobata, Kanji Yamaguchi, Yoshito Itoh

World journal of gastroenterology 19 ( 11 ) 1834 - 40 2013年3月

記述言語：英語掲載種別：研究論文（学術雑誌）

Autoimmune hepatitis (AIH) is a necroinflammatory liver disease of unknown etiology. The disease is characterized histologically by interface hepatitis, biochemically by increased aspartate aminotransferase and alanine aminotransferase levels, and serologically by increased autoantibodies and immunoglobulin G levels. Here we discuss AIH in a previously healthy 37-year-old male with highly elevated serum levels of soluble interleukin-2 receptor and markedly enlarged hepatoduodenal ligament lymph nodes (HLLNs, diameter, 50 mm). Based on these observations, the differential diagnoses were AIH, lymphoma, or Castleman's disease. Liver biopsy revealed the features of interface hepatitis without bridging fibrosis along with plasma cell infiltration which is the typical characteristics of acute AIH. Lymph node biopsy revealed lymphoid follicles with inflammatory lymphocytic infiltration; immunohistochemical examination excluded the presence of lymphoma cells. Thereafter, he was administered corticosteroid therapy: after 2 mo, the enlarged liver reached an almost normal size and the enlarged HLLNs reduced in size. We could not find AIH cases with such enlarged lymph nodes (diameter, 50 mm) in our literature review. Hence, we speculate that markedly enlarged lymph nodes observed in our patient may be caused by a highly activated, humoral immune response in AIH.

DOI： 10.3748/wjg.v19.i11.1834

Expression of Toll-Like Receptor 4 in Glomerular Endothelial Cells under Diabetic Conditions.

Shunsuke Takata, Yoshihiko Sawa, Takanobu Uchiyama, Hiroyuki Ishikawa

Acta histochemica et cytochemica 46 ( 1 ) 35 - 42 2013年2月

記述言語：英語掲載種別：研究論文（学術雑誌）

Diabetic conditions promote glomerulosclerosis by mesangial cells but the mechanisms are not fully elucidated. The present study evaluated the expression of toll-like receptor 4 in glomerular endothelial cells in the streptozotocin (STZ)-induced type 1 diabetic mouse (ICR-STZ) and the type 2 diabetic KK/TaJcl mouse which were fed a high fat diet feed (KK/Ta-HF). In the ICR-STZ and KK/Ta-HF almost glomeruli were immunostained with anti-TLR4 but there was no glomerulus immunostained by ani-TLR4 in the control ICR and KK/Ta. Laser-scanning confocal microscopy showed that the TLR4-positive region did not coincide with the podoplanin-positive region but coincide with the PECAM-1- and VE-cadherin-positive regions in the glomeruli of the ICR-STZ and KK/Ta-HF. The in situ hybridization showed that almost signals for TLR4 mRNA were present in the glomerulus of the ICR-STZ and KK/Ta-HF to a stronger extent than in the control ICR and KK/Ta. These suggest that glomerular endothelial cells usually express the TLR4 gene and hyperglycemia in the diabetic condition induces the TLR4 protein expression in the glomerular capillary endothelial cells. Cytokine productions through the TLR signaling pathway in glomerular endothelial cells may allow mesangial cells to produce extracellular matrix proteins in the diabetic milieu.

DOI： 10.1267/ahc.13002

Matrix Metalloproteinase-2 Degrades Fibrillin-1 and Fibrillin-2 of Oxytalan Fibers in the Human Eye and Periodontal Ligaments In Vitro 査読

Megumi Kawagoe, Eichi Tsuruga, Kyoko Oka, Yoshihiko Sawa, Hiroyuki Ishikawa

ACTA HISTOCHEMICA ET CYTOCHEMICA 46 ( 5 ) 153 - 159 2013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 HISTOCHEMISTRY & CYTOCHEMISTRY

Oxytalan fibers are distributed in the eye and periodontal ligaments (PDL). The ciliary zonule, known as Zinn's zonule, in the eye is composed of oxytalan fibers, which are bundles of microfibrils consisting mainly of fibrillin-1 and fibrillin-2. As turnover of oxytalan fibers is slow during life, their degradation mechanism remains unclarified. This study was performed to examine degradation pattern of fibrillin-1 and fibrillin-2 by experimental MMP activation. We cultured human non-pigmented ciliary epithelial cells (HNPCEC) and PDL fibroblasts for 7 days, then treated them with concanavalin A to activate matrix metalloproteinase (MMP)-2, and examined the degradation of fibrillin-1 and fibrillin-2 for 72 hr using immunofluorescence. At 7 days of HNPCEC culture, fibrillin-1-positive fibers were observed, some of which merged with fibrillin-2. After MMP-2 activation, fibrillin-1-positive fibers became thin and disappeared by 72 hr, while fibrillin-2-positive fibers disappeared almost completely within 24 hr. At 7 days of PDL fibroblast culture, fibrillin-1-positive fibers were mostly merged with fibrillin-2. After MMP-2 activation, fibrillin-1-positive fibers became thin by 24 hr and had almost disappeared by 48 hr, while fibrillin-2-positive fibers decreased constantly after 24 hr. A MMP-2 inhibitor completely suppressed these degradations. These results suggest that the patterns of fibrillin-1 and fibrillin-2 degradation differ between the eye and the PDL, possibly reflecting the sensitivity of fibrillin-1 and fibrillin-2 of each type of oxytalan fiber against MMP-2.

DOI： 10.1267/ahc.13024

Cellular turnover in epithelial rests of Malassez in the periodontal ligament of the mouse molar 査読国際誌

Kyoko Oka, Masakazu Morokuma, Kyoko Imanaka-Yoshida, Yoshihiko Sawa, Keitaro Isokawa, Masaki J Honda

European Journal of Oral Sciences 120 ( 6 ) 484 - 494 2012年12月

記述言語：英語掲載種別：研究論文（学術雑誌）

Fragments of Hertwig's epithelial root sheath persist in the periodontal ligament (PDL) in small clusters known as epithelial rests of Malassez (ERM). It is generally agreed that ERM are maintained as a quiescent and exclusively dental epithelial cluster in PDL. However, we speculate that homeostasis and cellular turnover underlies cluster maintenance. We also hypothesize that the fate of ERM clusters - diminishing or remaining - might be regulated via the presence or absence of epithelial stem cells therein. Histological analysis of aging mouse molar PDL showed that ERM clusters gradually increase in size with increasing age. Immunocytochemistry and cell culture revealed that ERM clusters contained Ki67-positive cells and were able to expand when brought in culture. The TdT-mediated biotin-dUTP nick-end labeling (TUNEL) procedure also detected signs of apoptosis. Finally, we identified putative epithelial stem cells in the clusters by 5-bromo-2′-deoxyuridine (BrdU) pulse-chase experiments and immunohistochemistry, using the stem-cell marker leucine-rich repeat-containing G-protein-coupled receptor 5 (Lgr5). The results suggest that ERM clusters are maintained in the PDL, via cellular turnover, throughout life. © 2012 Eur J Oral Sci.

DOI： 10.1111/eos.12003

Latent transforming growth factor-β binding protein 2 negatively regulates coalescence of oxytalan fibers induced by stretching stress 査読国際誌

Eichi Tsuruga, Kyoko Oka, Yuji Hatakeyama, Keitaro Isokawa, Yoshihiko Sawa

Connective Tissue Research 53 ( 6 ) 521 - 527 2012年12月

記述言語：英語掲載種別：研究論文（学術雑誌）

Oxytalan fibers are extracellular matrix components consisting of pure microfibrils. However, the mechanism whereby oxytalan fibers develop is not fully understood. We have previously reported that in human periodontal ligament (PDL) fibroblasts subjected to stretching stress, bundles of oxytalan fibers coalesce under the control of fibulin-5. Latent transforming growth factor-β binding protein 2 (LTBP-2) is known to bind to fibulin-5. The purpose of this study was to clarify the role of LTBP-2 in the coalescence of oxytalan fibers. We subjected PDL fibroblasts to stretching in order to examine the effects of LTBP-2 on the coalescence of oxytalan fibers in cell/matrix layers. Interaction of LTBP-2 with fibulin-5 was examined by immunoprecipitation assay, and changes in LTBP-2 deposition upon stretching were investigated by Western blotting and immunofluorescence assays. We used small interfering RNA against LTBP-2 in PDL cell culture and examined the appearance of oxytalan fibers on the basis of immunofluorescence. Stretching induced coalescence of oxytalan fibers, but did not affect LTBP-2 expression. The amount of extracellularly deposited LTBP-2 was decreased by about 70% as a result of stretching, compared with the control. LTBP-2 interacted with fibulin-5 on the fibers, and stretching decreased the amount of the LTBP-2 interacted with fibulin-5 by about 60%. Oxytalan fiber coalescence did not occur when LTBP-2 was suppressed by about 95%, whereas it occurred when LTBP-2 was suppressed by about 40%, fibulin-5 being colocalized with oxytalan fibers. These results suggest that LTBP-2, in response to tension stress, may negatively control the function of fibulin-5, thereby modulating the mechanism of oxytalan fiber coalescence. Copyright © Informa Healthcare USA, Inc.

DOI： 10.3109/03008207.2012.702816

Chimeric anti-podoplanin antibody suppresses tumor metastasis through neutralization and antibody-dependent cellular cytotoxicity 査読国際誌

Mika Kato Kaneko, Akiko Kunita, Shinji Abe, Yuta Tsujimoto, Masashi Fukayama, Kaoru Goto, Yoshihiko Sawa, Yasuhiko Nishioka, Yukinari Kato

Cancer Science 103 ( 11 ) 1913 - 1919 2012年11月

記述言語：英語掲載種別：研究論文（学術雑誌）

Podoplanin is a platelet aggregation-inducing factor associated with tumor metastasis, malignant progression, and cancer stem cells. We produced a rat-human chimeric anti-podoplanin mAb, NZ-8, from rat anti-podoplanin mAb (NZ-1). Although both NZ-1 and NZ-8 possess high binding affinities and high neutralizing activities of platelet aggregation, the antibody-dependent cellular cytotoxicity and complement-dependent cytotoxicity of NZ-8 were much higher than NZ-1. Furthermore, both NZ-1 and NZ-8 inhibited the growth of podoplanin-expressing tumors in vivo. Both NZ-1 and NZ-8 also suppressed hematogenous metastasis of podoplanin-expressing tumors. These results suggest that anti-podoplanin mAbs suppressed hematogenous metastasis by both neutralization and antibody-dependent cellular cytotoxicity/complement-dependent cytotoxicity activities. Targeting therapy to podoplanin-expressing tumors should be useful as a novel immunotherapy. © 2012 Japanese Cancer Association.

DOI： 10.1111/j.1349-7006.2012.02385.x

歯周組織においてFibulin-4はLOXL2/エラスチン複合体形成を調節する

山内由宣, 中冨佑香, 中島一記, 敦賀英知, 沢禎彦, 石川博之

福岡歯科大学学会雑誌 38 ( 増補 ) 37 - 37 2012年11月

小児癌の抗癌剤治療が引き起こす永久歯の形成障害について

西村紗和, 沢禎彦, 石川博之

福岡歯科大学学会雑誌 38 ( 増補 ) 29 - 29 2012年11月

Intracellular interaction of EMILIN-1 with fibrillin-1 in human periodontal ligament cells 査読

Yuka Nakatomi, Eichi Tsuruga, Yoshinori Yamauchi, Megumi Kawagoe, Kaori Yamanouchi, Kazuki Nakashima, Yoshihiko Sawa, Hiroyuki Ishikawa

Orthodontic Waves 71 ( 2 ) 66 - 69 2012年6月

記述言語：英語掲載種別：研究論文（学術雑誌）

The elastic system fibers comprise oxytalan, elaunin and elastic fibers, differing in their relative microfibril and elastin contents. Human periodontal ligaments (PDLs) contain oxytalan fibers (pure microfibrils), which are composed mainly of fibrillin-1, the major component of microfibrils. We recently demonstrated that EMILIN-1, located at the interface between elastin and microfibrils, controls the amount of fibrillin-1 assembly in PDL fibroblast cell/matrix layers [8], although the mechanism involved was unclear. We subjected cultured PDL fibroblasts to immunofluorescence and immunoprecipitation assays in order to examine the intracellular interaction of EMILIN-1 with fibrillin-1. Immunofluorescence showed that EMILIN-1 was colocalized with fibrillin-1, both inside and outside the cells. Additionally, EMILIN-1 formed a complex with fibrillin-1 in the intracellular fraction. These results suggest that EMILIN-1 may form complexes with fibrillin-1 in cellular vesicles, thus contributing effectively to the initial assembly of pericellular fibrillin-1 during the process of oxytalan fiber formation. © 2012 Elsevier Ltd and the Japanese Orthodontic Society.

DOI： 10.1016/j.odw.2012.01.002

The expression of podoplanin and classic cadherins in the mouse brain 査読国際誌

Chiaki Kaji, Miwa Tomooka, Yukinari Kato, Hiroshi Kojima, Yoshihiko Sawa

Journal of Anatomy 220 ( 5 ) 435 - 446 2012年5月

記述言語：英語掲載種別：研究論文（学術雑誌）

Podoplanin is a transmembrane glycoprotein indirectly linked to classic cadherins through ezrin-actin networks. Recently, the overexpression of podoplanin in high-grade malignancy brain tumors has been reported. The aim of this study was to investigate the expression of podoplanin and classic cadherins in the mouse brain. Immunohistochemistry showed that podoplanin was expressed on ependymal cells and choroid plexus epithelial cells at the ventricle side of the cell surface and at the cell-cell junctions, and on retinal pigment epithelial cells and in the pia mater
P-cadherin between choroid plexus epithelial cells and endothelial cells at the basement membrane side of cell surface, and between retinal pigment epithelial cells
VE-cadherin on the PECAM-1 positive-choroid plexus endothelial cells of the fibrovascular core
and N-cadherin on the cell surface and at the cell-cell junctions of ependymal cells, and in the pia mater. The regions expressing podoplanin, P-cadherin, and VE-cadherin did not coincide. In real-time PCR analysis, the amounts of podoplanin and P- and N-cadherin mRNA were larger in the ventricular wall with choroid plexus than in the abdominal aorta and cerebrum. In the RT-PCR analysis, the intensities of amplicon for VE-cadherin mRNA were the same for the abdominal aorta, cerebrum, and ventricular wall with the choroid plexus, suggesting that mouse ependymal cells, choroid plexus epithelial cells, and glial cells under the pia mater have the ability to express podoplanin and P- and N-cadherins. Glial cells and retinal pigment epithelial cells may create barriers by podoplanin and classic cadherins as a rate-determining step for transmission of blood components. © 2012 The Authors. Journal of Anatomy © 2012 Anatomical Society.

DOI： 10.1111/j.1469-7580.2012.01484.x

Factors associated with the overall survival of elderly patients with hepatocellular carcinoma. 国際誌

Hideki Fujii, Yoshito Itoh, Naoki Ohnishi, Masafumi Sakamoto, Tohru Ohkawara, Yoshihiko Sawa, Koichi Nishida, Yasuo Ohkawara, Kanji Yamaguchi, Masahito Minami, Takeshi Okanoue

World journal of gastroenterology 18 ( 16 ) 1926 - 32 2012年4月

記述言語：英語掲載種別：研究論文（学術雑誌）

AIM: To identify the factors associated with overall survival of elderly patients with hepatocellular carcinoma (HCC). METHODS: A total of 286 patients with HCC (male/female: 178/108, age: 46-100 years), who were diagnosed and treated by appropriate therapeutic procedures between January 2000 and December 2010, were enrolled in this study. Patients were stratified into two groups on the basis of age: Elderly (≥ 75 years old) and non-elderly (< 75 years old). Baseline clinical characteristics as well as cumulative survival rates were then compared between the two groups. Univariate and multivariate analyses were used to identify the factors associated with prolonged overall survival of patients in each group. Cumulative survival rates in the two groups were calculated separately for each modified Japan Integrated Stage score (mJIS score) category by the Kaplan-Meier method. In addition, we compared the cumulative survival rates of elderly and non-elderly patients with good hepatic reserve capacity (≤ 2 points as per mJIS). RESULTS: In the elderly group, the proportion of female patients, patients with absence of hepatitis B or hepatitis C viral infection, and patients with coexisting extrahepatic comorbid illness was higher (56.8% vs 31.1%, P < 0.001; 27.0% vs 16.0%, P = 0.038; 33.8% vs 22.2%, P = 0.047; respectively) than that in the non-elderly group. In the non-elderly group, the proportion of hepatitis B virus (HBV)-infected patients was higher than that in the elderly group (9.4% vs 0%, P = 0.006). The cumulative survival rates in the elderly group were 53.7% at 3 years and 32.9% at 5 years, which were equivalent to those in the non-elderly group (55.9% and 39.4%, respectively), as shown by a log-rank test (P = 0.601). In multivariate analysis, prolonged survival was significantly associated with the extent of liver damage and stage (P < 0.001 and P < 0.001, respectively), but was not associated with patient age. However, on individual evaluation of factors in both groups, stage was significantly (P < 0.001) associated with prolonged survival. Regarding mJIS scores of ≤ 2, the rate of female patients with this score was higher in the elderly group when compared to that in the non-elderly group (P = 0.012) and patients ≥ 80 years of age tended to demonstrate shortened survival. CONCLUSION: Survival of elderly HCC patients was associated with liver damage and stage, but not age, except for patients ≥ 80 years with mJIS score ≤ 2.

DOI： 10.3748/wjg.v18.i16.1926

Roles of Collagen and Periostin Expression by Cranial Neural Crest Cells during Soft Palate Development 査読国際誌

Kyoko Oka, Masaki J. Honda, Eichi Tsuruga, Yuji Hatakeyama, Keitaro Isokawa, Yoshihiko Sawa

Journal of Histochemistry and Cytochemistry 60 ( 1 ) 57 - 68 2012年1月

記述言語：英語掲載種別：研究論文（学術雑誌）

The tissue in the palatal region can be divided into the hard and the soft palates, each having a specialized function such as occlusion, speech, or swallowing. Therefore, an understanding of the mechanism of palatogenesis in relation to the function of each region is important. However, in comparison with the hard palate, there is still a lack of information about the mechanisms of soft palate development. In this study, the authors investigated the contribution of cranial neural crest (CNC) cells to development of both hard and soft palates. They also demonstrated a unique pattern of periostin expression during soft palate development, which was closely related to that of collagen type I (Col I) in palatine aponeurosis. Furthermore, organ culture analysis showed that exogenous transforming growth factor-β (TGF-β) induced the expression of both periostin and Col I. These novel patterns of expression in the extracellular matrix (ECM) induced by CNC cells suggest that these cells may help to determine the character of both the hard and soft palates through ECM induction. TGF-β signaling appears to be one of the mediators of Col I and periostin expression in the formation of functional structures during soft palate development. © The Histochemical Society 2012.

DOI： 10.1369/0022155411427059

Immunohistochemical examination of novel rat monoclonal antibodies against mouse and human podoplanin 査読

Chiaki Kaji, Yuta Tsujimoto, Mika Kato Kaneko, Yukinari Kato, Yoshihiko Sawa

Acta Histochemica et Cytochemica 45 ( 4 ) 227 - 237 2012年

記述言語：英語掲載種別：研究論文（学術雑誌）

This study aims to develop new monoclonal antibodies (mAbs) against mouse and human podoplanin. Rats were immunized with synthetic peptides, corresponding to amino acids 38-51 of mouse podoplanin or human podoplanin which is 100% homologous to the same site of monkey podoplanin
anti-mouse podoplanin mAb PMab-1 (IgG2a) and anti-human mAb NZ-1.2 (IgG2a) were established. In immunocytochemistry, the mouse melanoma B16-F10 and mouse podoplanin (mPDPN)-expressed CHO transfectant were stained by PMab-1
human lymphatic endothelial cells (LEC) and human podoplanin (hPDPN)-expressed squamous cell carcinoma HSC3 transfectant, were stained by NZ-1.2. Western-blot analysis detected an about 40-kDa protein in CHO-mPDPN and B16-F10 by PMab-1, and in HSC3-hPDPN and LEC by NZ-1.2. In frozen sections, PMab-1 reacted with mouse kidney, pulmonary alveoli, pulmonary pleura, and salivary gland myoepithelial cells while NZ-1.2 reacted to the human salivary gland myoepithelial cells. The immunostaining of paraffin-embedded sections also showed the reaction of PMab-1 or NZ-1.2 to the mouse or monkey kidney glomerulus, pulmonary alveoli, and lung lymphatic vessels. These results indicate that the two novel rat mAbs to the mouse and human/monkey podoplanin are useful for Western-blot and immunostaining of somatic tissues on paraffin-embedded sections as well as frozen sections. © 2012 The Japan Society of Histochemistry and Cytochemistry.

DOI： 10.1267/ahc.12008

Fibrillin-1 and Fibrillin-2 Are Essential for Formation of Thick Oxytalan Fibers in Human Nonpigmented Ciliary Epithelial Cells In Vitro 査読国際誌

Kaori Yamanouchi, Eichi Tsuruga, Kyoko Oka, Yoshihiko Sawa, Hiroyuki Ishikawa

CONNECTIVE TISSUE RESEARCH 53 ( 1 ) 14 - 20 2012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：INFORMA HEALTHCARE

The ciliary zonule, also known as Zinn's zonule, is composed of oxytalan fibers. However, the mechanism by which epithelial cells in the ciliary body form these fibers in not fully understood. We examined human nonpigmented ciliary epithelial cells to determine the appearance and amount of oxytalan fibers in terms of positivity for their major components, fibrillin-1 and fibrillin-2. Examination of fibrillin-1 and fibrillin-2 expression by immunofluorescence revealed that thin fibers positive for fibrillin-1 on Day 2 changed to thick fibers by Day 8. The fibers positive for fibrillin-2 appeared on the thick fibrillin-1-positive fibers after Day 4. Northern blot analysis revealed that the level of fibrillin-1 did not change markedly, while induction of fibrillin-2 gene was evident on Day 5. Western blot analysis showed that fibrillin-1 deposition increased gradually, while that of fibrillin-2 increased markedly from Day 5 to Day 8. Fibrillin-1 suppression did not lead to the formation of fibrillin-2-positive thick fibers, whereas fibrillin-2 suppression led to the formation of fibrillin-1-positive thin fibers, but not thick fibers. These results suggest that both fibrillin-1 and fibrillin-2 are essential for the formation of thick oxytalan fibers in the ciliary zonule and are informative for clarifying the mechanism of homeostasis of the ocular matrix.

DOI： 10.3109/03008207.2011.602767

Immunohistochemical and immunocytochemical localization of amylase in rat parotid glands and von Ebner's glands by ion etching-immunoscanning electron microscopy 査読

Junko Yahiro, Tetsuichiro Inai, Akihito Tsutsui, Atsuko Sato, Toshikazu Nagato, Kunihisa Taniguchi, Eichi Tsuruga, Yoshihiko Sawa

Acta Histochemica et Cytochemica 44 ( 5 ) 201 - 212 2011年8月

記述言語：英語掲載種別：研究論文（学術雑誌）

The distribution of amylase in rat parotid glands and von Ebner's glands was examined using ion etching-immunoscanning electron microscopy, which enables both light and electron microscopic observations of identical semi-thin resin sections immunolabeled with anti-α- amylase and immunogold in association with silver enhancement. At the light microscopic level, most acinar secretory granules (SG) and striated duct secretions of parotid glands were strongly stained dark brown. In von Ebner's glands, acinar SG and duct secretions were weakly to strongly stained light to dark brown. At the electron microscopic level, labeling was observed as bright gold-silver particles. The labeling intensity of acinar SG of parotid glands was higher than that of von Ebner's glands. In parotid glands, weak labeling of SG in transitional cells between acini and intercalated ducts, very weak labeling of SG in intercalated ducts, and strong labeling of striated duct secretions were observed. In von Ebner's glands, the secretions and some SG of interlobular ducts were strongly labeled compared to those of intralobular ducts and SG of acini. Less amylase was synthesized in von Ebner's acini compared to parotid acini, whereas von Ebner's ducts may secrete significantly more amylase to modify saliva than parotid ducts. © 2011 The Japan Society of Histochemistry and Cytochemistry.

DOI： 10.1267/ahc.10039

Establishment of a novel monoclonal antibody SMab-1 specific for IDH1-R132S mutation 査読国際誌

Mika Kato Kaneko, Wei Tian, Shingo Takano, Hiroyuki Suzuki, Yoshihiko Sawa, Yasukazu Hozumi, Kaoru Goto, Kentaro Yamazaki, Chifumi Kitanaka, Yukinari Kato

Biochemical and Biophysical Research Communications 406 ( 4 ) 608 - 613 2011年3月

記述言語：英語掲載種別：研究論文（学術雑誌）

Isocitrate dehydrogenase 1 (IDH1) mutations, which are early and frequent genetic alterations in gliomas, are specific to a single codon in the conserved and functionally important Arginine 132 (R132) in IDH1. We earlier established a monoclonal antibody (mAb), IMab-1, which is specific for R132H-containing IDH1 (IDH1-R132H), the most frequent IDH1 mutation in gliomas. To establish IDH1-R132S-specific mAb, we immunized mice with R132S-containing IDH1 (IDH1-R132S) peptide. After cell fusion using Sendai virus envelope, IDH1-R132S-specific mAbs were screened in ELISA. One mAb, SMab-1, reacted with the IDH1-R132S peptide, but not with other IDH1 mutants. Western-blot analysis showed that SMab-1 reacted only with the IDH1-R132S protein, not with IDH1-WT protein or IDH1 mutants, indicating that SMab-1 is IDH1-R132S-specific. Furthermore, SMab-1 specifically stained the IDH1-R132S-expressing glioblastoma cells in immunocytochemistry and immunohistochemistry, but did not react with IDH1-WT or IDH1-R132H-containing glioblastoma cells. We newly established an anti-IDH1-R132S-specific mAb SMab-1 for use in diagnosis of mutation-bearing gliomas. © 2011 Elsevier Inc.

DOI： 10.1016/j.bbrc.2011.02.102

Stretch stimuli increase fibulin-5/EMILIN-1 complex on oxytalan fibers in human periodontal ligament cells 査読

Kazuki Nakashima, Eichi Tsuruga, Yuka Nakatomi, Yoshinori Yamauchi, Yuichiro Hata, Sachio Tamaoki, Yoshihiko Sawa, Hiroyuki Ishikawa

Orthodontic Waves 70 ( 1 ) 15 - 20 2011年3月

記述言語：英語掲載種別：研究論文（学術雑誌）

Periodontal ligaments (PDLs) are continuously exposed to various functional forces, such as tooth movement and occlusal loading. Human PDLs comprise elastic system fibers as well as collagen fibers. The elastic system fibers in turn comprise oxytalan, elaunin and elastic fibers, which differ in their relative microfibril and elastin contents. Human PDLs contain oxytalan fibers (pure microfibrils), which are composed mainly of fibrillin-1 (the major component of microfibrils). We have previously reported that bundles of oxytalan fibers in cultured PDL cells coalesce in response to mechanical strain, and that this coalescence is controlled by fibulin-5. However, the relationship between fibulin-5 and other fibrillin-1-binding molecules is unclear. In the present study we investigated whether fibulin-5 and EMILIN-1 (both of which are fibrillin-1-binding molecules) contribute to the formation of oxytalan fibers upon exposure to mechanical strain. We subjected PDL cells to stretching in order to examine the role of fibulin-5 and EMILIN-1 in the formation of oxytalan fibers in cell/matrix layers. We examined the relationship between fibulin-5 and EMILIN-1 in PDL cell cultures using immunofluorescence and immunoprecipitation assay. Immunofluorescence showed that fibulin-5 and EMILIN-1 were colocalized on fibrillin-1-positive oxytalan fibers. Fibulin-5 formed a complex with EMILIN-1, and stretching increased the amount of this complex relative to cells that were not subjected to stretching. These results suggest that the expression of the fibulin-5/EMILIN-1 complex is upregulated in response to tension strain, and may control the formation of oxytalan fibers in PDLs. © 2010 Japanese Orthodontic Society.

DOI： 10.1016/j.odw.2010.09.001

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