精子发生是指精原干细胞自我更新与分化为精子细胞的过程。精原干细胞的命运决定主要受细胞内源性分子的调控。然而，非编码小RNA（miRNA）对人类精原干细胞自我更新与分化的作用及其机理尚未见报道。申请人前期RNA深度测序显示，395种miRNA在正常与非梗阻性无精子症（NOA）病人精原细胞的表达差异显著。因此，本项目拟开展如下重要研究内容：1）发现和鉴定关键miRNA（如miR-122-5p、miR-373-3p、miR-100-5p、miR-145-5p）在正常与NOA病人精原干细胞的差异和定位表达；2）探讨关键miRNA对正常和NOA病人的精原干细胞自我更新与分化为精子细胞的作用；3）揭示关键miRNA对正常和NOA病人的精原干细胞自我更新与分化为精子细胞的调控靶标及其生物学功能。本项目将为阐释人类精子发生及其异常机制提供新的表观遗传调控机理，对治疗男性不育和预防子代出生缺陷有重要意义。

Spermatogenesis is a complex process by which spermatogonial stem cells self-renew and differentiate into mature sperm. Studies on spermatogonial stem cells are of great significance since they are the unique stem cells that transmit genetic information to subsequent generations. The fate determinations of spermatogonial stem cells are mainly regulated by endogenous molecules within the cells. However, it remains unknown about the roles and mechanisms of non-coding small RNA (miRNA) in regulating the self-renewal and differentiation of spermatogonial stem cells. We have explored the function and targets of miRNA-20 and miRNA-106a in mediating the proliferation and maintenance of mouse spermatogonial stem cells (Stem Cells, 2013, 31: 2205-2217). Furthermore, we have compared the global expression profiles of miRNAs among human spermatogonia, pachytene spermatocytes, and round spermatids, and we identified 599 miRNAs that are differentially expressed among these cells (Scientific Reports, 2015, 5: 8084). Notably, we have recently revealed, using RNA deep-sequencing and real time PCR, that 395 miRNA were differentially expressed in human spermatogonia between normal men and non-obstructive azoospermia (NOA) patients. Based upon these findings, it is essential and necessary for us to conduct this important project with the following Aims: i) to identify the differential expression and cellular localization of certain key miRNA, including has-miR-122-5p,has-miR-373-3p,has-miR-100-5p, and has-miR-145-5p, in human spermatogonial stem cells between normal men and NOA patients using Northern blots and the in situ hybridization; ii) to explore the roles of certain key miRNA, including has-miR-122-5p, has-miR-373-3p, has-miR-100-5p, and has-miR-145-5p, in regulating the self-renewal and differentiation of normal and aberrant human spermatogonial stem cells of NOA patients; and iii) to unveil the binding targets of these key miRNA and the function of their targets in mediating the fate determinations of normal and aberrant human spermatogonial stem cells of NOA patients. Significantly, this project could provide novel epigenetic regulatory mechanisms underlying human normal and abnormal spermatogenesis. This project is of particular significance, because it could offer novel endogenous targets for treating male infertility and preventing the birth defect of the offspring.