基于“肾藏精”理论研究补肾填精法调控小鼠精原干细胞增殖及改善少弱精子症小鼠生精功能的作用及机制

Oligoasthenozoospermia is a main cause of male infertility with an ascending trend year by year, but its pathogenesis is still unclear and no effective modern medicine can be chosen. "Kidney storing essence, governing reproduction" is a core theory of Traditional Chinese Medicine; it plays a key role in understanding male infertility with oligoasthenozoospermia. In our previous study, kidney reinforcing drug, Yangjing capsule, was used to treat oligoasthenozoospermia infertility and the therapeutic effect is obvious. In this study, an oligoasthenozoospermia mouse model, resulted by tripterygium glycosides feeding, was used to investigate the treatment mechanism of Yangjing capsule in reproduction endocrine, gene regulation and micro-environment of spermatogenesis. Sperm stem cell (SSC) is a base and guarantee of male procreation. Recently, GDNF, as well as GDNF signal pathway (GDNF-GFRa1-PI3K/AKT-POU3F1), is considered to be the most important regulator. Considering that Yangjing capsule can cooperate with GDNF to promote SSCs proliferation, we present a hypothesis that Yangjing capsule is though the same GDNF pathway to stimulate SSCs generation. We next blocked the pathway using siRNA for GFRa1 or POU3F1 and specific inhibitor of PI3K/AKT to elucidate the truth. In conclusion, the study contributes to understand "kidney storing essence" of "ZANG-XIANG THEORY" in Traditional Chinese Medicine (i.e. SSCs is the material base of congenital essence), to elucidate its nature and to guide the treatment of male infertility in clinic.

"肾藏精,主生殖"理论与男性不育关系密切。服用补肾填精中药-养精胶囊，可以显著提高男性少、弱精子症患者精子质量，但其现代生物学机制依然不清。本研究利用雷公藤多苷灌胃法建立少、弱精子症小鼠模型，给予养精胶囊治疗，从内分泌、基因和睾丸生精微环境等层面，研究其作用途径。精原干细胞(SSCs)是雄性生育能力的基础和保障，近年来认为GDNF及其通路(GDNF-GFRa1-PI3K/AKT-POU3F1)是维持SSCs自我增殖的主要因子和通路。研究发现养精胶囊可以与GDNF协同促进小鼠精原干细胞增殖，因此我们推测养精胶囊是通过GDNF通路促进SSCs增殖。本研究利用RNA干扰技术(siRNA)及特异性阻断剂阻断上述通路，研究其作用机制。这对于完善与创新"肾藏精"脏象理论即精原干细胞是男子先天之"精"的物质基础，揭示"肾藏精，主生殖"的科学本质，以及指导临床治疗男性不育症，具有重要的理论和现实意义。

“肾藏精，主生殖”理论与男性不育关系密切，课题组前期的研究表明，补肾填精中药—养精胶囊，可以显著提高男性少、弱精子症患者精子质量，但其现代生物学机制依然不清。本研究利用环磷酰胺制造小鼠生精损伤模型，养精胶囊灌胃后，从内分泌、基因和睾丸生精微环境等层面，研究其作用途径和机制。利用小鼠精原细胞系GC-1细胞为精原干细胞的体外模型，检测养精胶囊水提物对其增殖凋亡及相关基因通路的影响。结果表明，养精胶囊能改善环磷酰胺所致小鼠的生精功能损伤，一方面与降低凋亡调控基因Bax/Bcl的表达比值而抑制生精细胞凋亡有关，另一方面与其显著上调AR的表达，增强T在精子发生中的作用相关。养精胶囊水提物可以促进GC-1细胞的增殖，这可能是通过激活GFRα1受体，提高PI3K/AKT信号通路的表达，上调POU3F1基因表达有关，并可以GC-1细胞的凋亡。这对于完善“肾藏精”脏象理论即精原干细胞是男子先天之“精”的物质基础，揭示“肾藏精，主生殖”的科学本质，以及指导临床治疗男性不育症，具有重要的理论和现实意义。