研究者詳細 - 佐藤　伸

FGF and BMP derived from dorsal root ganglia regulate blastema induction in limb regeneration in Ambystoma mexicanum 査読

Akira Satoh, Aki Makanae, Yurie Nishimoto, Kazumasa Mitogawa

DEVELOPMENTAL BIOLOGY 417 ( 1 ) 114 - 125 2016年9月

担当区分：筆頭著者,　責任著者記述言語：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

Urodele amphibians have a remarkable organ regeneration ability that is regulated by neural inputs. The identification of these neural inputs has been a challenge. Recently, Fibroblast growth factor (Fgf) and Bone morphogenic protein (Bmp) were shown to substitute for nerve functions in limb and tail regeneration in urodele amphibians. However, direct evidence of Fgf and Bmp being secreted from nerve endings and regulating regeneration has not yet been shown. Thus, it remained uncertain whether they were the nerve factors responsible for successful limb regeneration. To gather experimental evidence, the technical difficulties involved in the usage of axolotls had to be overcome. We achieved this by modifying the electroporation method. When Fgf8-AcGFP or Bmp7-AcGFP was electroporated into the axolotl dorsal root ganglia (DRG), GFP signals were detectable in the regenerating limb region. This suggested that Fgf8 and Bmp7 synthesized in neural cells in the DRG were delivered to the limbs through the long axons. Further knockdown experiments with double-stranded RNA interference resulted in impaired limb regeneration ability. These results strongly suggest that Fgf and Bmp are the major neural inputs that control the organ regeneration ability. (C) 2016 Elsevier Inc. All rights reserved.

DOI： 10.1016/j.ydbio.2016.07.005

Cooperative inputs of Bmp and Fgf signaling induce tail regeneration in urodele amphibians 査読

Aki Makanae, Kazumasa Mitogawa, Akira Satoh

DEVELOPMENTAL BIOLOGY 410 ( 1 ) 45 - 55 2016年2月

担当区分：最終著者,　責任著者記述言語：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

Urodele amphibians have remarkable organ regeneration ability. They can regenerate not only limbs but also a tail throughout their life. It has been demonstrated that the regeneration of some organs are governed by the presence of neural tissues. For instance, limb regeneration cannot be induced without nerves. Thus, identifying the nerve factors has been the primary focus in amphibian organ regeneration research. Recently, substitute molecules for nerves in limb regeneration, Bmp and Fgfs, were identified. Cooperative inputs of Bmp and Fgfs can induce limb regeneration in the absence of nerves. In the present study, we investigated whether similar or same regeneration mechanisms control another neural tissue governed organ regeneration, i.e., tail regeneration, in Ambystoma mexicanum. Neural tissues in a tail, which is the spinal cord, could transform wound healing responses into organ regeneration responses, similar to nerves in limb regeneration. Furthermore, the identified regeneration inducer Fgf2+Fgf8+Bmp7 showed similar inductive effects. However, further analysis revealed that the blastema cells induced by Fgf2+Fgf8+Bmp7 could participate in the regeneration of several tissues, but could not organize a patterned tail. Regeneration inductive ability of Fgf2+Fgf8+Bmp7 was confirmed in another urodele, Pleurodeles waltl. These results suggest that the organ regeneration ability in urodele amphibians is controlled by a common mechanism. (C) 2015 The Authors. Published by Elsevier Inc.

DOI： 10.1016/j.ydbio.2015.12.012

Skeletal callus formation is a nerve-independent regenerative response to limb amputation in mice and Xenopus. 査読

Miura S, Takahashi Y, Satoh A, Endo T

Regeneration (Oxford, England) 2 ( 4 ) 202 - 216 2015年8月

Regeneration inducers in limb regeneration 査読

Akira Satoh, Kazumasa Mitogawa, Aki Makanae

DEVELOPMENT GROWTH & DIFFERENTIATION 57 ( 6 ) 421 - 429 2015年8月

担当区分：筆頭著者,　責任著者記述言語：英語出版者・発行元：WILEY-BLACKWELL

Limb regeneration ability, which can be observed in amphibians, has been investigated as a representative phenomenon of organ regeneration. Recently, an alternative experimental system called the accessory limb model was developed to investigate early regulation of amphibian limb regeneration. The accessory limb model contributed to identification of limb regeneration inducers in urodele amphibians. Furthermore, the accessory limb model may be applied to other species to explore universality of regeneration mechanisms. This review aims to connect the insights recently gained to emboss universality of regeneration mechanisms among species. The defined molecules (BMP7 (or2)+FGF2+FGF8) can transform skin wound healing to organ (limb) regeneration responses. The same molecules can initiate regeneration responses in some species.

DOI： 10.1111/dgd.12230

Comparative Analysis of Cartilage Marker Gene Expression Patterns during Axolotl and Xenopus Limb Regeneration 査読

Kazumasa Mitogawa, Aki Makanae, Ayano Satoh, Akira Satoh

PLOS ONE 10 ( 7 ) e0133375 2015年7月

担当区分：最終著者,　責任著者記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：PUBLIC LIBRARY SCIENCE

Axolotls (Ambystoma mexicanum) can completely regenerate lost limbs, whereas Xenopus laevis frogs cannot. During limb regeneration, a blastema is first formed at the amputation plane. It is thought that this regeneration blastema forms a limb by mechanisms similar to those of a developing embryonic limb bud. Furthermore, Xenopus laevis frogs can form a blastema after amputation; however, the blastema results in a terminal cone-shaped cartilaginous structure called a "spike." The causes of this patterning defect in Xenopus frog limb regeneration were explored. We hypothesized that differences in chondrogenesis may underlie the patterning defect. Thus, we focused on chondrogenesis. Chondrogenesis marker genes, type I and type II collagen, were compared in regenerative and nonregenerative environments. There were marked differences between axolotls and Xenopus in the expression pattern of these chondrogenesis-associated genes. The relative deficit in the chondrogenic capacity of Xenopus blastema cells may account for the absence of total limb regenerative capacity.

DOI： 10.1371/journal.pone.0133375

Skeletal Callus Formation is a Nerve-independent Regenerative Response to Limb Amputation in Mice and Xenopus 査読

Miura S, Takahashi Y, Satoh A, Endo T

Regeneration 2 ( 4 ) 202 - 216 2015年1月

The accessory limb model: an alternative experimental system of limb regeneration. 査読

Endo T, Gardiner DM, Makanae A, Satoh A

Methods in molecular biology (Clifton, N.J.) 1290 101 - 113 2015年

Identification of a feather β-keratin gene exclusively expressed in pennaceous barbule cells of contour feathers in chicken. 査読

Kowata K, Nakaoka M, Nishio K, Fukao A, Satoh A, Ogoshi M, Takahashi S, Tsudzuki M, Takeuchi S

Gene 542 ( 1 ) 23 - 28 2014年5月

Ectopic blastema induction by nerve deviation and skin wounding: A new regeneration model in Xenopus laevis 査読

Mitogawa K, Hirata A, Moriyasu M, Makanae A, Miura S, Endo T, Satoh A

Regeneration 1 ( 2 ) 26 - 36 2014年1月

Implication of two different regeneration systems in limb regeneration 査読

Makanae A, Mitogawa K, Satoh A

Regeneration 1 ( 3 ) 1 - 9 2014年1月

Conservation of Position-specific Gene Expression in Axolotl Limb Skin 査読

Akira Satoh, Aki Makanae

ZOOLOGICAL SCIENCE 31 ( 1 ) 6 - 13 2014年1月

担当区分：最終著者,　責任著者記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：ZOOLOGICAL SOC JAPAN

Urodele amphibians can regenerate their limbs after amputation. After amputation, undifferentiated cells appear on the amputation plane and form regeneration blastema. A limb blastema recreates a complete replica of the original limb. It is well known that disturbance of the location of limb tissues prior to amputation perturbs limb patterning, suggesting that different intact limb tissues carry different location information despite their identical appearance. The cause of such differences in intact tissues remains unknown. In this study, we found that Lmx1b, Tbx2, and Tbx3 genes, which are expressed in developing limb in a region specific manner, remained detectable in a mature axolotl limb. Furthermore, those position-specific gene expression patterns were conserved in mature limbs. Treatment with retinoic acid ( RA), which is known to have ventralizing activity, changed Lmx1b expression in intact dorsal skin and dorsal character to ventral, indicating that conserved Lmx1b expression was due to the dorsal character and not leaky gene expression. Furthermore, we found that such conserved gene expression was rewritable in regeneration blastemas. These results suggest that axolotl limb cells can recognize their locations and maintain limbness via conserved expression profiles of developmental genes.

DOI： 10.2108/zsj.31.6

Nerve independent limb induction in axolotls 査読

Aki Makanae, Ayako Hirata, Yasuko Honjo, Kazumasa Mitogawa, Akira Satoh

Developmental Biology 381 ( 1 ) 213 - 226 2013年9月

担当区分：最終著者,　責任著者記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：Academic Press Inc.

Urodele amphibians can regenerate their limbs. During limb regeneration, dermal fibroblasts are transformed into undifferentiated cells called blastema cells. These dermis-blastema cells show multipotency. Such so-called endogenous reprogramming of cell differentiation is one of the main targets of amphibian limb regeneration studies. It is well recognized that nerve presence controls the initiation of limb regeneration. Accordingly, nerve factors have been sought in amphibian limb regeneration. To investigate it, a relatively new study system called the accessory limb model (ALM) was developed. Using ALM, two signaling cascades (Fgf and Gdf5 signaling) came under focus. In the present study, Growth and differentiation factor-5 (Gdf5) application to wounded skin initiated limb regeneration responses and resulted in induction of a blastema-like structure in the absence of a nerve. However, the Gdf5-induced structure showed defects as a regeneration blastema, such as absence of detectable Prrx1 expression by in situ hybridization. The defects could be remedied by additional Fibroblasts growth factor (Fgf) inputs. These two inputs (Gdf5 and Fgfs) were sufficient to substitute for the nerve functions in the induction of limb regeneration. Indeed, Fgf2, Fgf8, and Gdf5 applications with the contralateral skin graft resulted in limb formation without nerve supply. Furthermore, acquisition of cartilage differentiation potential of dermal fibroblasts was tested in an in vivo and in vitro combination assay. Dermal fibroblasts cultured with Gdf5 were difficult to participate in cartilage formation when the cultured cells were grafted into cartilage forming region. In contrast, dermal fibroblasts cultured with Fgf2 and Fgf8 became easier to participate into cartilage formation in the same procedure. These results contribute to our understanding of molecular mechanisms of the early phase of amphibian limb regeneration. © 2013 The Authors.

DOI： 10.1016/j.ydbio.2013.05.010

Scopus

Accessory limb induction on flank region and its muscle regulation in axolotl 査読

Ayako Hirata, Aki Makanae, Akira Satoh

Developmental Dynamics 242 ( 8 ) 932 - 940 2013年8月

担当区分：最終著者,　責任著者記述言語：英語掲載種別：研究論文（学術雑誌）

Background: Urodele amphibians have high regeneration capability that has been studied for a long time. Recently, a new experimental system called the accessory limb model was developed and becomes alternative choice for amphibian limb regeneration study. Although the accessory limb model has many advantages, an improvement was needed for some specific analysis, such as studying muscle origin. For that purpose, an accessory limb induction on nonlimb regions was attempted. Results: Accessory limb induction on a nonlimb region (flank) was possible by nerve deviation and limb skin grafting. Retinoic acid injections improved the induction rate. The induced limb possessed the same tissue context as a normal limb. Muscle cells were also abundantly observed. It is speculated that the muscle cells are derived from flank muscle tissues, because limb muscle cells are a migratory cell population and the accessory limb was induced apart from the original limb. We also found that migration of the muscle cells was regulated by Hgf/cMet signaling as in other vertebrates. Conclusions: Accessory limb induction was possible even in the nonlimb flank region. The flank-induced limb would be useful for further analysis of limb regeneration, especially for migratory cell populations such as muscle cells. Developmental Dynamics 242:932-940, 2013. © 2013 Wiley Periodicals, Inc.

DOI： 10.1002/dvdy.23984

Scopus

Spatiotemporal regulation of keratin 5 and 17 in the axolotl limb 査読

Miyuki Moriyasu, Aki Makanae, Akira Satoh

DEVELOPMENTAL DYNAMICS 241 ( 10 ) 1616 - 1624 2012年10月

担当区分：最終著者,　責任著者記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：WILEY-BLACKWELL

Background: Amphibians have greater regeneration capability than higher vertebrates. They can regenerate their limbs after an amputation. As a limb is regenerated, a regeneration-specific epithelium called the apical epithelial cap (AEC) is induced. The AEC is an essential structure for limb regeneration. Despite the importance of the AEC, molecular marker genes have not been well studied at the molecular level. Results: In the present study, keratin5 (KRT5) and KRT17 were investigated in an axolotl-regenerating limb. KRT5 and KRT17 were expressed in a regenerating limb but down-regulated in a differentiating limb. KRT5 showed characteristic regulation in a regenerating blastema. KRT5 was suppressed in the basal layer of the AEC. This KRT5 suppression was correlated to the blastema differentiation and nerve presence. Simple skin wounding could also upregulate both KRT5 and KRT17 gene expression. But these genes were suppressed within a shorter time than in limb regeneration. Conclusions: The KRT5 and KRT17 gene profile can be a useful marker gene to investigate AEC in limb regeneration. Developmental Dynamics 241:16161624, 2012. (c) 2012 Wiley Periodicals, Inc.

DOI： 10.1002/dvdy.23839

Early Regulation of Axolotl Limb Regeneration 査読

Aki Makanae, Akira Satoh

ANATOMICAL RECORD-ADVANCES IN INTEGRATIVE ANATOMY AND EVOLUTIONARY BIOLOGY 295 ( 10 ) 1566 - 1574 2012年10月

担当区分：最終著者,　責任著者記述言語：英語掲載種別：研究論文（学術雑誌）出版者・発行元：WILEY-BLACKWELL

Amphibian limb regeneration has been studied for a long time. In amphibian limb regeneration, an undifferentiated blastema is formed around the region damaged by amputation. The induction process of blastema formation has remained largely unknown because it is difficult to study the induction of limb regeneration. The recently developed accessory limb model (ALM) allows the investigation of limb induction and reveals early events of amphibian limb regeneration. The interaction between nerves and wound epidermis/epithelium is an important aspect of limb regeneration. During early limb regeneration, neurotrophic factors act on wound epithelium, leading to development of a functional epidermis/epithelium called the apical epithelial cap (AEC). AEC and nerves create a specific environment that inhibits wound healing and induces regeneration through blastema formation. It is suggested that FGF-signaling and MMP activities participate in creating a regenerative environment. To understand why urodele amphibians can create such a regenerative environment and humans cannot, it is necessary to identify the similarities and differences between regenerative and nonregenerative animals. Here we focus on ALM to consider limb regeneration from a new perspective and we also reported that focal adhesion kinase (FAK)Src signaling controlled fibroblasts migration in axolotl limb regeneration. Anat Rec, 2012. (c) 2012 Wiley Periodicals, Inc.

DOI： 10.1002/ar.22529

Activation of germline-specific genes is required for limb regeneration in the Mexican axolotl 査読

Wei Zhu, Gerald M. Pao, Akira Satoh, Gillian Cummings, James R. Monaghan, Timothy T. Harkins, Susan V. Bryant, S. Randal Voss, David M. Gardiner, Tony Hunter

DEVELOPMENTAL BIOLOGY 370 ( 1 ) 42 - 51 2012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

The capacity for tissue and organ regeneration in humans is dwarfed by comparison to that of salamanders. Emerging evidence suggests that mechanisms learned from the early phase of salamander limb regeneration-wound healing, cellular dedifferentiation and blastemal formation-will reveal therapeutic approaches for tissue regeneration in humans. Here we describe a unique transcriptional fingerprint of regenerating limb tissue in the Mexican axolotl (Ambystoma mexicanum) that is indicative of cellular reprogramming of differentiated cells to a germline-like state. Two genes that are required for self-renewal of germ cells in mice and flies, Piwi-like 1 (PL1) and Piwi-like 2 (PL2), are expressed in limb blastemal cells, the basal layer keratinocytes and the thickened apical epithelial cap in the wound epidermis in the regenerating limb. Depletion of PL1 and PL2 by morpholino oligonucleotides decreased cell proliferation and increased cell death in the blastema leading to a significant retardation of regeneration. Examination of key molecules that are known to be required for limb development or regeneration further revealed that FGF8 is transcriptionally downregulated in the presence of the morpholino oligos, indicating PL1 and PL2 might participate in FGF signaling during limb regeneration. Given the requirement for FGF signaling in limb development and regeneration, the results suggest that PL1 and PL2 function to establish a unique germline-like state that is associated with successful regeneration. (C) 2012 Elsevier Inc. All rights reserved.

DOI： 10.1016/j.ydbio.2012.07.021

Retrotransposon long interspersed nucleotide element-1 (LINE-1) is activated during salamander limb regeneration 査読

Wei Zhu, Dwight Kuo, Jason Nathanson, Akira Satoh, Gerald M. Pao, Gene W. Yeo, Susan V. Bryant, S. Randal Voss, David M. Gardiner, Tony Hunter

DEVELOPMENT GROWTH & DIFFERENTIATION 54 ( 7 ) 673 - 685 2012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：WILEY-BLACKWELL

Salamanders possess an extraordinary capacity for tissue and organ regeneration when compared to mammals. In our effort to characterize the unique transcriptional fingerprint emerging during the early phase of salamander limb regeneration, we identified transcriptional activation of some germline-specific genes within the Mexican axolotl (Ambystoma mexicanum) that is indicative of cellular reprogramming of differentiated cells into a germline-like state. In this work, we focus on one of these genes, the long interspersed nucleotide element-1 (LINE-1) retrotransposon, which is usually active in germ cells and silent in most of the somatic tissues in other organisms. LINE-1 was found to be dramatically upregulated during regeneration. In addition, higher genomic LINE-1 content was also detected in the limb regenerate when compared to that before amputation indicating that LINE-1 retrotransposition is indeed active during regeneration. Active LINE-1 retrotransposition has been suggested to have a potentially deleterious impact on genomic integrity. Silencing of activated LINE-1 by small RNAs has been reported to be part of the machinery aiming to maintain genomic integrity. Indeed, we were able to identify putative LINE-1-related piRNAs in the limb blastema. Transposable element-related piRNAs have been identified frequently in the germline in other organisms. Thus, we present here a scenario in which a unique germline-like state is established during axolotl limb regeneration, and the re-activation of LINE-1 may serve as a marker for cellular dedifferentiation in the early-stage of limb regeneration.

DOI： 10.1111/j.1440-169X.2012.01368.x

Nerve signaling regulates basal keratinocyte proliferation in the blastema apical epithelial cap in the axolotl (Ambystoma mexicanum) 査読

Akira Satoh, Susan V. Bryant, David M. Gardiner

DEVELOPMENTAL BIOLOGY 366 ( 2 ) 374 - 381 2012年6月

担当区分：筆頭著者記述言語：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

The ability of adult vertebrates to repair tissue damage is widespread and impressive; however, the ability to regenerate structurally complex organs such as the limb is limited largely to the salamanders. The fact that most of the tissues of the limb can regenerate has led investigators to question and identify the barriers to organ regeneration. From studies in the salamander, it is known that one of the earliest steps required for successful regeneration involves signaling between nerves and the wound epithelium/apical epithelial cap (AEC). In this study we confirm an earlier report that the keratinocytes of the AEC acquire their function coincident with exiting the cell cycle. We have discovered that this unique, coordinated behavior is regulated by nerve signaling and is associated with the presence of gap junctions between the basal keratinocytes of the AEC. Disruption of nerve signaling results in a loss of gap junction protein, the reentry of the cells into the cell cycle, and regenerative failure. Finally, coordinated exit from the cell cycle appears to be a conserved behavior of populations of cells that function as signaling centers during both development and regeneration. (C) 2012 Elsevier Inc. All rights reserved.

DOI： 10.1016/j.ydbio.2012.03.022