Characterization of the dedifferentiation process induced by newt regeneration derived extracts

蝾螈，Notophthalmus viridescens，有能力再生结构（如。四肢、尾巴、脊髓、透镜），其因损伤或截肢而丧失。 损伤部位的细胞恢复为胚胎类型的细胞，然后可以增殖并改变命运以重建丢失的结构。 这个过程被称为去分化，是再生过程的关键之一，了解去分化背后的分子基础有助于增强人类的再生潜力。 研究表明，哺乳动物的肌肉细胞，当用蝾螈再生提取物处理时，重新进入细胞周期，并从其多核结构中分裂出来，产生增殖的单核细胞。然而，这个过程不是很有效，许多细胞在这个过程中经历细胞死亡。 目前研究计划的目的是研究蝾螈细胞的去分化过程，以便更好地了解如何改善人类的去分化和再生。 我们将研究蝾螈A1肌细胞在培养中已被诱导去分化，通过添加血清到培养基中或通过处理与蝾螈衍生的提取物。 我们将使用一种称为代表性差异分析（RDA）的技术来分离和表征蝾螈去分化过程中具有潜在调控作用的基因。 我们提出了一个全面的战略，用于分离，优先考虑和分析的候选基因与调节作用的去分化过程。 我们将1）对细胞系进行RDA以鉴定去分化起始候选基因，2）优先考虑cDNA以鉴定相关候选基因，3）对再生中的候选基因进行功能分析。 了解参与去分化的机制将在退行性疾病和神经元损伤中具有巨大的治疗应用。

The newt, Notophthalmus viridescens, has the ability to regenerate structures (eg. limbs, tail, spinal cord, lens) which are lost to injury or amputation. Cells at the site of the injury revert back to an embryonic type of cell that can then proliferate and switch fate to rebuild the lost structure. This process, called dedifferentiation, is one of the keys to the regeneration process, and understanding the molecular basis behind dedifferentiation could help to enhance regenerative potential in humans. Studies have shown that mammalian muscle cells, when treated with newt regeneration extracts, re-enter the cell cycle and break-up from their multinucleated structures to yield proliferating mononuclear cells. This process is not very efficient, however, and many of the cells undergo cell death in the process. The aim of the current research proposal is to study the dedifferentiation process in newt cells in order to better understand how to improve dedifferentiation and regeneration in humans. We will study newt A1 muscle cells in culture that have been induced to dedifferentiate by adding serum to the medium or by treatment with newt-derived extracts. We will use a technique called Representational Difference Analysis (RDA) to isolate and characterize genes with potential regulatory roles during the newt dedifferentiation process. We present a comprehensive strategy for the isolation, prioritization and analysis of candidate genes with regulatory roles in the dedifferentiation process. We will 1) Conduct RDA on cell lines to identify dedifferentiation-initiating candidate genes, 2) Prioritize cDNAs to identify relevant candidates and 3) Perform functional analysis of candidate genes in regeneration. An understanding of the mechanisms involved in dedifferentiation will have tremendous therapeutic applications in degenerative disease and in neuronal injury.