Exploring a role for regulation of TGFbeta signaling during photoreceptor regener

描述（由申请人提供）：随着最近对再生医学和干细胞疗法的兴趣激增，生物学家重新投资于使用现代分子技术的再生研究，这些技术在该领域的早期是不可用的。条件突变体和转基因斑马鱼是硬骨鱼模型研究体系，是研究人类遗传疾病、发育、成体干细胞活化和再生机制的重要工具。事实上，斑马鱼被用于再生研究，因为它们具有强大的再生能力，可以再生鳍、部分心脏、肾脏组织，重要的是再生神经视网膜。遗传工具促进了这项研究，因为控制再生的信号机制可以在体内模型系统的背景下进行探索，然后应用于哺乳动物模型系统的生物医学研究。在哺乳动物系统中，当视网膜受损时，小胶质细胞（MG）对损伤的反应是形成胶质疤痕，不能恢复视力。相比之下，斑马鱼MG作为视网膜干细胞，在光损伤后2周内再生失去的光感受器并恢复视力。对光损伤后36小时内再生视网膜中分离MG细胞的微阵列分析发现，与先前发表的再生心脏和鳍的研究相比，MG细胞的基因表达变化相似，包括转录共抑制因子tgf2相互作用因子（tgif1）和sin眼相关同源盒3b （six3b）。这表明tgif1和six3b可能参与了非组织特异性的一般再生程序。这两种基因都抑制TGF2信号，并与人类先天性前脑和面部缺陷有关。由于tgif1和six3b具有共同的作用，并且都在受损视网膜MG的早期增加，因此tgif1和six3b表达的增加可能通过动态调节TGF2信号传导，在视网膜再生的早期阶段允许MG干细胞去分化和/或有丝分裂激活。本研究将全面分析tgif1和six3b在早期视网膜再生中的作用，并开发新的转基因鱼线和视网膜外植体系统，用于未来视网膜再生的研究。本研究将通过识别诱导MG增殖和随后的再生所必需的基因，为视网膜损伤后哺乳动物MG的研究提供信息，从而为再生医学提供潜在的治疗靶点。

DESCRIPTION (provided by applicant): With the recent explosion of interest in regenerative medicine and stem cell therapies, biologists have reinvested in the study of regeneration with modern molecular techniques that were not available in the field's early years. Conditional mutants and transgenic zebrafish, a teleost fish model research system, are useful tools to study human genetic diseases, development, and mechanisms of adult stem cell activation and regeneration. In fact, zebrafish are used in regeneration research because they have a robust capacity to regenerate the fins, portions of the heart, renal tissue, and importantly the neural retina. Genetic tools facilitate this research because signaling mechanisms controlling regeneration can be explored in the context of an in vivo model system and then be applied to biomedical research in mammalian model systems. In mammalian systems, when the retina is damaged, M|ller glial (MG) cells react to the damage by forming a glial scar and do not restore vision. In contrast, the zebrafish MG act as retinal stem cells to regenerate the lost photoreceptors and restore vision within 2 weeks after the light injury. A microarray analysis of isolated MG from regenerating retinas in the first 36 hours after light damage found similar gene expression changes in MG cells compared with previously published studies of the regenerating heart and fin, including the transcriptional co-repressors TGF2-interacting factor (tgif1) and sine oculis-related homeobox 3b (six3b). This suggests that tgif1 and six3b may be involved in a general regeneration program that is not tissue-specific. Both of these genes repress TGF2 signaling and are associated with congenital forebrain and facial defects in humans. Because tgif1 and six3b have common roles and both increase early in the MG of damaged retina, increased tgif1 and six3b expression may allow for the dedifferentiation and/or mitotic activation of MG stem cells during the early stages of retina regeneration by dynamically regulating TGF2 signaling. The proposed research will provide a comprehensive analysis of the role tgif1 and six3b have in early retina regeneration and develop a new transgenic fish line and retinal explant system to use in future studies of retinal regeneration. This research will inform studies of the mammalian MG following retinal damage by identifying genes that are necessary for induction of MG proliferation and subsequent regeneration, thus providing potential therapeutic targets for regenerative medicine. PUBLIC HEALTH RELEVANCE: The proposed research will help define what changes in gene expression are necessary in the stem cells of the retina, M|ller glia, to regenerate lost photoreceptors after damage and restore eye sight. The results of this study will help determine gene targets for developing therapeutic treatments for blindness and low vision in humans.