Key Laboratory of Model Animals and Stem Cell Biology in Hunan Province, Engineering Research Center of Reproduction and Translational Medicine of Hunan Province, Hunan Normal University School of Medicine, Changsha, 410013, Hunan, China.
Department of Pharmacy, The Third Xiangya Hospital, Central South University, Changsha, 410011, Hunan, China.
Cell Mol Life Sci. 2024 Feb 17;81(1):93. doi: 10.1007/s00018-023-05077-z.
Stem Leydig cells (SLCs) are essential for maintaining normal spermatogenesis as the significant component of testis microenvironment and gonadal aging. Although progress has been achieved in the regulation of male germ cells in mammals and humans, it remains unknown about the genes and signaling pathways of human SLCs. Here we have demonstrated, for the first time, that WNT5A (Wnt family member 5a) mediates the proliferation, apoptosis, and stemness of human SLCs, namely NGFR Leydig cells. We revealed that NGFR Leydig cells expressed NGFR, PDGFRA, NES, NR2F2, and THY1, hallmarks for SLCs. RNA-sequencing showed that WNT5A was expressed at a higher level in human SLCs than non-SLCs, while immunohistochemistry and Western blots further illustrated that WNT5A was predominantly expressed in human SLCs. Notably, CCK-8, EdU and Western blots displayed that WNT5A enhanced the proliferation and DNA synthesis and retained stemness of human SLCs, whereas flow cytometry and TUNEL analyses demonstrated that WNT5A inhibited the apoptosis of these cells. WNT5A knockdown caused an increase in LC lineage differentiation of human SLCs and reversed the effect of WNT5A overexpression on fate decisions of human SLCs. In addition, WNT5A silencing resulted in the decreases in nuclear translocation of β-catenin and expression levels of c-Myc, CD44, and Cyclin D1. Collectively, these results implicate that WNT5A regulates the proliferation, apoptosis and stemness of human SLCs through the activation of the β-catenin signaling pathway. This study thus provides a novel molecular mechanism underlying the fate determinations of human SLCs, and it offers a new insight into the niche regulation of human testis.
睾丸间质干细胞（Stem Leydig cells，SLCs）是睾丸微环境和性腺衰老的重要组成部分，对维持正常精子发生至关重要。尽管在哺乳动物和人类中对雄性生殖细胞的调控已经取得了进展，但人类 SLCs 的基因和信号通路仍不清楚。在这里，我们首次证明 WNT5A（Wnt 家族成员 5a）介导了人类 SLCs（即 NGFR 间质细胞）的增殖、凋亡和干性。我们揭示了 NGFR 间质细胞表达 NGFR、PDGFRA、NES、NR2F2 和 THY1，这是 SLCs 的标志。RNA 测序显示，WNT5A 在人类 SLCs 中的表达水平高于非 SLCs，而免疫组织化学和 Western blot 进一步表明 WNT5A 主要在人类 SLCs 中表达。值得注意的是，CCK-8、EdU 和 Western blot 显示 WNT5A 增强了人类 SLCs 的增殖和 DNA 合成，并保留了其干性，而流式细胞术和 TUNEL 分析表明 WNT5A 抑制了这些细胞的凋亡。WNT5A 敲低导致人类 SLCs 的 LC 谱系分化增加，并逆转了 WNT5A 过表达对人类 SLCs 命运决定的影响。此外，WNT5A 沉默导致β-catenin 的核易位和 c-Myc、CD44 和 Cyclin D1 的表达水平降低。总之，这些结果表明 WNT5A 通过激活β-catenin 信号通路调节人类 SLCs 的增殖、凋亡和干性。因此，本研究为人类 SLCs 的命运决定提供了一个新的分子机制，并为人类睾丸龛调控提供了新的见解。
