补肾祛湿法通过调控精原干细胞niche改善大鼠生精功能的研究

Male infertility is a common male disease, the incidence of which increased year by year. Spermatogonial stem cell niche environment is closely related to the male spermatogenesis. The secretion of cytokines in niche has important implications on the proliferation and differentiation of spermatogonial stem cells. The Youjing granule is developed based on the pathogenesis characteristics of "kidney deficiency, dampness" and therapies method of tonifying kidney and dampness removing. Youjing granule has a significant effect on male infertility in clinical application. Previous studies found that Youjing granule could improve the quality of sperm in infertile men, which may be related to spermatogonial stem cells niche. On this study, we use animal experiments and in vitro co-culture experiments, and observe the effects of Youjing granule on spermatogenesis, microvascular density, secretory function of Leydig cell and Sertoli cell of spermatogenic damage rat model. Simultaneously, it was detected that the effects of Youjing granule on pro spermatogonial stem cell proliferation and differentiation cytokines secreted in Sertoli cells in vitro, as well as the proliferation and differentiation of spermatogonial stem cells after cell co-culture. And then its mechanism was clarified the tonifying kidney and dampness removing method improved spermatogenesis through regulation of spermatogonial stem cells niche. In conclusion, this study would contribute to elaborate the scientific connotation of “tonifying kidney and dampness removing method”, and use Chinese medicine to treat male infertility.

男性不育症是临床常见的男性疾病，发病比例逐年增高。精原干细胞龛境(niche)与男性的生精功能密切相关，其分泌的细胞因子对精原干细胞的增殖和分化具有重要影响。基于“肾虚、湿浊”病机特点，以“补肾祛湿”为治法研制的“优精颗粒”，临床应用以来，治疗男性不育症疗效显著。前期研究发现，其改善男性不育症患者的精子质量，可能与调控精原干细胞niche有关。本研究以“优精颗粒”为“补肾祛湿”法的代表方，利用动物实验观察其对生精损伤模型大鼠生精功能、微血管密度、睾丸间质细胞和支持细胞分泌功能的影响；利用体外细胞培养实验，检测其对体外支持细胞分泌促精原干细胞增殖、分化细胞因子，对细胞共培养后精原干细胞的增殖、分化状态的影响，探讨其作用机制。进而明确“补肾祛湿”法通过调控精原干细胞niche改善生精功能的作用机制。这对于指导临床治疗男性不育症，阐明“补肾祛湿”法的理论内涵，具有重要的理论意义和应用价值。

研究表明补肾祛湿中药方剂可以提高大鼠的精子质量。本课题进一步利用生精损伤模型大鼠生精功能下降，以及生精环境损伤的特点，在体研究补肾祛湿方对大鼠生精功能以及睾丸内精原干细胞 niche 组成细胞（Sertoli 细胞、Leydig 细胞）、微血管密度，以及促进 SSCs 增殖分化细胞因子的影响；原代分离大鼠的精原干细胞和小鼠胚胎成纤维（MEFs）细胞，进行体外共培养，观察补肾祛湿方含药血清对其分泌细胞因子的影响，以及 SSCs 增殖、分化基因表达的变化，阐释补肾祛湿中药提高生精功能，促进 SSCs 增殖分化的作用机制。本课题通过研究，较充分地获得了补肾祛湿法通过调控精原干细胞niche改善大鼠生精功能的证据，初步揭示了补肾祛湿法影响精原干细胞增殖能力的机制，获得了较多有用的实验依据，为后续研究提供了新线索、新经验、新依据。