近年来男性不育发生率日益增加，已成为影响社会发展的潜在问题之一，研究精原干细胞干性维持的机制对理解精子发生和男性不育的治疗具有十分重要意义。各种因素导致的DNA损伤可能是精原干细胞干性丧失的重要原因之一，我们前期的研究表明miR-499在小鼠精原干细胞DNA双链损伤应答过程中与53BP1表达密切相关，初步数据分析也提示53BP1可能是miR-499的靶基因之一。故本研究拟采免疫组化和流式细胞仪检测技术，探讨miR-499对电离辐射导致的精原干细胞DNA双链损伤应答反应的关键作用，同时采用基因芯片技术和生物信息学等手段检测分析miR-499对53BP1的相互作用，并验证miR-499对DNA损伤应答的分子机制。本研究将有助于阐明miR-499-53BP1信号通路在精原干细胞DNA双链损伤和修复过程中的具体途径，将很有可能为临床上男性不育的防治提供新策略。

The incidence of male infertility is dramatically increasing in recent years, which has become one of the potential problems affecting the development of society. Unraveling the mechanisms of the maintenance of stemness of spermatagonial stem cells will definitely benefit the understanding of spermatogenesis and the therapyfor male infertility.DNA damage caused by various reasons may lead to the loss of stemness of spermatagonial stem cells.Our current results indicate that miR-499 may play an important role in regulation of mouse spermatogonial stem cellsto the response tothe double-stranded DNA damage. Our preliminary datashowed that53BP1 expression level is closely controlled by miR-499, which suggests that53BP1 is one of the potentialtargets of miR-499.Therefore, this study is designed to understand the roles of miR-499 in double-stranded DNA damagecaused by ionizing radiationin spermatogonial stem cellsby taking advantages of immunohistochemistry and flow cytometry instrument testing technology. Meanwhile, we will analyze the interaction between miR-499 and 53BP1 by using gene chip technology and bioinformatics method. Further rescue experimentsby rebuilding seminiferous tubule with potential targets for miR-499 reposing to DNA damage will also be performed to verify the potential targets. This study will unravel the signaling pathways of miR-499-53BP1 in spermatogonial stem cells in the process of double-stranded DNA damage. It will also open a new avenue to develop novel strategies for prevention and control of male infertility in clinic.

精原干细胞（spermatogonial stem cell，SSC）是雄性哺乳动物体内唯一的生殖干细胞，DNA损伤，尤其是双链DNA损伤（DNA double strand break, DSB）对于SSC的自我更新和分化有重要的影响。因此，SSC对DSB的修复对于维持生殖细胞的遗传稳定性发挥重要的作用。本课题研究小鼠SSC经低剂量电离辐射照射后产生DNA损伤应答，检验53BP1及其它相关分子在照射后的动态变化，发现在SSC中53BP1蛋白的表达量显著增高，而传统的DSB修复相关分子γH2AX以及凋亡相关的分子Caspase3表达量并未见改变。不同的是其它的生精细胞及体细胞γH2AX表达量增多。我们的研究表明在SSC中，经低剂量的电离辐射照射后没有引起SSC凋亡，触发了不同于其它细胞对DSB损伤修复机制的，由53BP1参与介导SSC特有的DSB损伤修复通路。我们的研究对于揭示SSC的DNA损伤修复机制有重要的贡献，对于由环境因素，尤其是电离辐射造成的男性不育症及男性生育力下降的诊断和治疗有重要意义。