基于“肾藏精”理论研究补肾填精法通过miRNA-20a调控小鼠精原干细胞增殖的作用机制

Spermatogonial stem cells (SSCs) have a high degree of self-renewal and differentiation potential, which plays an irreplaceable role in the maintenance of male reproductive capacity. In recent years, it has been found that overexpression of miRNA-20a can downregulate the expression of target gene STAT3 and Bim, and then promote the proliferation of SSCs. Preliminary studies show that Yangjing capsule (TCM) can promote the proliferation of SSCs, and prove its mechanism is partly through PI3K/Akt signaling pathway. In the preliminary experiment, Yangjing capsule extract can increase the content of miRNA-20a in cells, and cell proliferation activity decreased after downregulating miRNA-20a. Therefore, the study group speculated that the Chinese medicine of enhancing kidney essence can increase miRNA-20a, and then downregulate the expression of STAT3 and Bim, which promote the proliferation of SSCs. This study will do animal and cell experiments , using RT-PCR, Western blot, fluorescence in situ hybridization (FISH), siRNA technology,to confirm whether medicine of enhancing kidney essence can promote SSCs proliferation by miRNA and the specific molecular mechanism. This will provide a new scientific basis for the treatment of male infertility by tonifying the kidney essence, which will contribute to the sublimation of the theory of kidney storing essence.

精原干细胞（SSCs）具有高度自我更新和分化潜能，在维持男性生殖能力方面发挥着不可替代的作用。近年研究发现，过表达miRNA-20a可下调靶基因STAT3、Bim的表达，进而促进SSCs的增殖。前期研究表明，养精胶囊（补肾填精中药）可促进SSCs的增殖，且其部分机制是通过PI3K/Akt通路实现的。预实验中，养精胶囊提取液可增加细胞中miRNA-20a的含量，且下调miRNA-20a后，细胞增殖活性降低。故课题组推测补肾填精中药可通过上调miRNA-20a，进而下调STAT3及Bim的表达，进而促进SSCs的增殖。本研究拟通过动物及细胞实验，运用RT-PCR、Western blot、荧光原位杂交（FISH）、小干扰等技术，进一步研究补肾填精中药通过miRNA促进SSCs的增殖及其具体的分子机制。这将为补肾填精治疗男性不育症提供新的科学依据，并有利于升华“肾藏精，主生殖”理论。

近年来男性精液质量呈逐渐下降的趋势，但由于造成男性生育能力下降的因素尚未完全明确，故现代医学在临床治疗上以经验性治疗为主，但临床疗效不确切。祖国传统医学以“肾藏精，主生殖”为基本指导理论在治疗男性不育症方面有着良好的疗效。本研究则证实补肾填精中药可以通过提高minRNA-20a的表达，一方面降调STAT3的表达，从而增加Plzf的表达，促进细胞增殖；另一方面，降调Bim，继而减少Casepase-3的表达，减少细胞凋亡。这对于阐明补肾填精法治疗男性不育提供现代医学证据，对于揭示“肾主生殖”的科学内涵及提高男性的生殖健康具有重要的理论意义和临床价值。