Tet1/Tet2敲除影响小鼠精原干细胞自我更新和多能性及其分子机制

Tet(ten-eleven translocation) family proteins include Tet1, Tet2 and Tet3. Tets regulate DNA demethylation by epigenetic modification and play important roles in many biological processes, including embryonic development, embryonic stem cell pluripotency and differentiation, and tumorigenesis. Recently we and others found that Tet1 and Tet2 deficiency leads to telomere shortening and reduced differentiation competency of embryonic stem cells. Moreover, our preliminary observations indicate that Tet1 and Tet2 knockout mice exhibit premature reproductive aging symptoms with age. Spermatogonial stem cells can self-renewal and have differentiation potential when cultured in vitro. Therefore spermatogonial stem cells have application potential in regenerative medicine. This proposed study is to investigate whether Tets deficiency leads to spermatogonial stem cell pluripotency and self-renewal defect. We propose the following three aims to be tested in in vitro study: (1) Tet1 and Tet2 deficiency causes spermatogonial stem cell self-renew and pluripotency defect. We anticipate that the data obtained from this study will promote the basic theory for the clinical diagnosis and therapy of male sterile.

Tet家族蛋白包括Tet1，Tet2和Tet3，调控DNA去甲基化，在胚胎发育、胚胎干细胞多能性及肿瘤发生中起重要作用。我们发现Tet1、Tet2缺陷会导致端粒缩短，影响胚胎干细胞分化发育能力，并且Tet1、Tet2缺陷小鼠随年龄增长表现出生殖早衰症状。精原干细胞在体外培养条件下也具有自我更新和多向分化潜能，在再生医学领域具有应用价值。本课题试图研究Tets是否通过DNA去甲基化修饰，影响精原干细胞多能性和自我更新，并揭示其分子机制。提出并验证以下假设：Tet1、Tet2缺陷引起精原干细胞自我更新和多能性降低。期望该研究结果为临床诊断与治疗雄性不育提供理论基础。

统计数据显示，人类不孕不育的发病率逐年上升，已高达12%-20%，部分国家甚至高达25%。不孕不育症将被列为21世纪人类三大疾病之一，仅次于肿瘤和心脑血管疾病。影响不孕不育的因素有环境污染、食品安全、药物毒性、生活压力增加等原因。棒曲霉素是一种由霉菌产生的代谢产物。这类霉菌可以感染不同的水果和蔬菜，主要偏好于苹果，导致其腐烂，因此，以苹果和苹果为原料的副产品是人类接触棒曲霉素的主要来源。多个研究表明棒曲霉素的摄入会对多个器官产生毒性，但生殖毒性研究较少，本研究以雌性生殖细胞和雄性生殖细胞为研究对象，探究棒曲霉素引起的生殖毒性及相关的机制。结果显示，棒曲霉素暴露可导致卵巢颗粒细胞内ROS水平明显升高，高水平ROS导致DNA损伤，继而激活了细胞内的ATM-p53信号通路，最终导致细胞凋亡；卵母细胞暴露棒曲霉素后显著降低了卵母细胞成熟，深入探究发现卵母细胞纺锤体组装异常，中期染色体排列错乱，胞内ROS水平升高。雄性生殖细胞GC1和TM4暴露棒曲霉素后也出现细胞凋亡增加等现象。以上结果显示，棒曲霉素在雄性和雌性中均具有明显的生殖毒性，是不育不孕潜在的原因之一。