Sertoli Cell Wt1 Regulates Peritubular Myoid Cell and Fetal Leydig Cell Differentiation during Fetal Testis Development

Sertoli cells play a significant role in regulating fetal testis compartmentalization to generate testis cords and interstitium during development. The Sertoli cell Wilms’ tumor 1 (Wt1) gene, which encodes ~24 zinc finger-containing transcription factors, is known to play a crucial role in fetal testis cord assembly and maintenance. However, whether Wt1 regulates fetal testis compartmentalization by modulating the development of peritubular myoid cells (PMCs) and/or fetal Leydig cells (FLCs) remains unknown. Using a Wt1-/flox; Amh-Cre mouse model by deleting Wt1 in Sertoli cells (Wt1SC-cKO) at embryonic day 14.5 (E14.5), Wt1 was found to regulate PMC and FLC development. Wt1 deletion in fetal testis Sertoli cells caused aberrant differentiation and proliferation of PMCs, FLCs and interstitial progenitor cells from embryo to newborn, leading to abnormal fetal testis interstitial development. Specifically, the expression of PMC marker genes α-Sma, Myh11 and Des, and interstitial progenitor cell marker gene Vcam1 were down-regulated, whereas FLC marker genes StAR, Cyp11a1, Cyp17a1 and Hsd3b1 were up-regulated, in neonatal Wt1SC-cKO testes. The ratio of PMC:FLC were also reduced in Wt1SC-cKO testes, concomitant with a down-regulation of Notch signaling molecules Jag 1, Notch 2, Notch 3, and Hes1 in neonatal Wt1SC-cKO testes, illustrating changes in the differentiation status of FLC from their interstitial progenitor cells during fetal testis development. In summary, Wt1 regulates the development of FLC and interstitial progenitor cell lineages through Notch signaling, and it also plays a role in PMC development. Collectively, these effects confer fetal testis compartmentalization.支持细胞（Sertoli cells）在胚胎发育过程中调控胎儿睾丸分区化，以形成睾丸索与睾丸间质，发挥着关键作用。威尔姆斯瘤1基因（Wilms’ tumor 1，Wt1）可编码约24个含锌指结构的转录因子，既往研究已证实其在胎儿睾丸索组装与维持中具有核心功能。然而，Wt1是否通过调控管周肌样细胞（peritubular myoid cells，PMCs）和/或胎儿莱迪希细胞（fetal Leydig cells，FLCs）的发育来参与胎儿睾丸分区化，目前仍不明确。本研究利用Wt1条件性敲除小鼠模型（Wt1-/flox; Amh-Cre mouse model），在胚胎第14.5天（E14.5）时敲除支持细胞中的Wt1（即Wt1SC-cKO模型），结果发现Wt1可调控PMCs与FLCs的发育。在胚胎发育至新生阶段，敲除胎儿睾丸支持细胞中的Wt1会导致PMCs、FLCs及间质祖细胞出现异常分化与增殖，最终引发胎儿睾丸间质发育异常。具体而言，在新生期Wt1SC-cKO小鼠睾丸中，PMCs标记基因α-Sma、Myh11及Des的表达水平显著下调，间质祖细胞标记基因Vcam1的表达亦出现下调；而FLCs标记基因StAR、Cyp11a1、Cyp17a1及Hsd3b1的表达则显著上调。同时，Wt1SC-cKO小鼠睾丸中PMCs与FLCs的比例降低，且Notch信号分子Jag 1、Notch 2、Notch 3及Hes1的表达水平下调，这表明在胎儿睾丸发育过程中，FLCs由其间质祖细胞分化的状态发生了改变。综上，Wt1可通过Notch信号通路调控FLCs与间质祖细胞谱系的发育，同时也参与PMCs的发育过程。上述效应共同促成了胎儿睾丸的正常分区化。