POLO-like1在精母细胞减数分裂过程的机制研究

Non-obstructive azoospermia is troubled by couples of childbearing age who can not get their offsprings. Spermatogenesis disorder is an important cause of male spermatogenesis, however, its pathogenesis is not clear. Our results of phosphorylated proteomics in mice testis showed that POLO-like1 (PLK1) was highly actived in mouse testes, regulated the cell cycle of spermatocytes and affected the spermatogenesis. It suggested that PLK1 regulated meiosis. Through the preliminary work, we propose that PLK1 may regulate the meiosis by regulating the substrate Racgap1, REC8 and BRCA1. In order to verify this hypothesis, we use mouse germ cell lines, mouse model, human testicular tissue samples, to discuss the important role of PLK1 in spermatogenesis and meiosis and certify the regulate and control mechanism by proteomics, flow cytometry, spermatogonial stem cell gene editing and transplantation. In this study, we show a new viewpoint to human sperm abnormalities, infertility diagnosis and infertility treatment. Furthermore, it provides an important theoretical support for non-obstructive azoospermia patients and the clinical treatment program.

非梗阻性无精子症（NOA）是困扰育龄夫妇不能获得后代的难题，生精障碍是导致NOA的重要原因，其发病机制尚不明确。我们前期小鼠睾丸组织磷酸化蛋白质组学结果显示POLO-like1（PLK1）在小鼠睾丸中具有高活性，在精母细胞系中抑制PLK1活性导致细胞周期阻滞，在小鼠中抑制其活性导致精母细胞减少；通过构建PLK1与精子发生相关底物蛋白质网络，筛选出Racgap1、REC8、BRCA1为PLK1在精子发生调控过程中的底物蛋白质。据此我们提出假说：PLK1可能通过调节底物Racgap1、REC8、BRCA1调控精母细胞减数分裂过程。本项目拟通过小鼠生殖细胞系、小鼠造模模型、人睾丸组织样本，运用蛋白质组学、流式细胞检测、精原干细胞基因编辑及移植等研究方法和手段，深入探讨PLK1在精母细胞减数分裂过程中的作用，为进一步研究人精子发生异常机制及不育症的诊疗提供新的线索和切入点。

非梗阻性无精子症（NOA）是困扰育龄夫妇不能获得后代的难题，生精障碍是导致NOA的重要原因，其发病机制尚不明确。通过辅助生殖技术从睾丸中获得功能性精子可以使部分NOA患者拥有生物学后代。我们通过临床数据分析，发现血清FSH水平可以预测NOA患者取精的结果，对指导临床取精方式及减少患者心理、生理及经济的负担有重要作用。但仍未解决NOA患者无精子的困扰，探寻生精障碍的发生机制是亟需解决的问题。在前期小鼠睾丸组织磷酸化蛋白质组学结果的基础上，对POLO-like1（PLK1）的功能在小鼠模型上进行验证。通过构建PLK1磷酸化活性位点失活小鼠模型，运用HE、IF、western blot等技术手段，深入探讨PLK1在精母细胞减数分裂过程中的作用，为进一步研究人精子发生异常机制及不育症的诊疗提供新的线索及切入点。