Dedifferentiation induced by newt extract

点击翻译按钮获取中文摘要

The newt, Notophthalmus viridescens, is an amphibian that can regenerate appendages, eye structures and many vital organs. During the regeneration process, tissues at the injury site revert from their differentiated form into an embryonic-like state. This dedifferentiation process generates a population of proliferating progenitor cells that can then redifferentiate into the multiple tissues of the regenerated structure. A better understanding of the dedifferentiation process would have tremendous implications for our understanding of regenerative potential in both newts and higher vertebrates. Dedifferentiation capabilities in both newts and mammals have been extensively studied in vitro using muscle-derived cell lines. Studies with the newt A1 muscle cell line have shown that mature differentiated A1 cells can re-enter the cell cycle following treatment with foetal bovine serum (FBS). Mammalian differentiated muscle cells do not possess this capability and remain post-mitotic. However, mouse C2C12 myotubes can re-enter the cell cycle following treatment with extract derived from a regenerating newt limb, suggesting that mammalian cells still possess dedifferentiation capabilities but may not have the appropriate triggers to initiate the response. We have recently shown that primary mouse myotubes do not re-enter the cell cycle following extract treatment. Thus it appears that C2C12 cells possess less dedifferentiation capability than newt cells but more than primary muscle cells. The aim of this proposal is to study the dedifferentiation process in newt and mammalian muscle cells, to determine the genetic differences that govern their abilities to dedifferentiate, and to ultimately improve the dedifferentiation process in mammalian cells. We have isolated another newt muscle cell line (called A2). Unlike A1 cells, A2 myotubes do not respond to FBS treatment by re-entering the cell cycle. In Aim 1, we will characterize the dedifferentiation process in newt A1 and A2 cells. By comparing the transcriptomes of A1 and A2 cells, we will determine which mutations allow cell cycle re-entry. We will also further characterize the dedifferentiation process and determine if cell death (caspase activation) and p53 activation are involved. In Aim 2, we will characterize the dedifferentiation process in mammalian C2C12 and primary muscle cells. Since C2C12 cells respond to extracts by fragmenting and re-entering the cell cycle, whereas primary muscle cells only fragment, we will determine the differences between the two cell types and assess whether cell cycle genes and apoptotic genes are involved in the responses of the cells to extract. Ultimately, a better understanding of the differences in dedifferentiation capability between newt cells and mammalian cells may help us to better understand the regenerative capability of newts and to improve regenerative ability in mammalian cells.