Wnt9b permits the androgen action in promoting Wolffian duct

The Wolffian or mesonephric duct is the embryonic primordium that gives rise to the epididymis, vas deferens and seminal vesicle. The androgen action in the Wolffian duct mesenchyme during fetal development is the predominant driver for Wolffian duct maintenance which is critical for male fertility. However, the androgens capability of promoting Wolffian duct maintenance was completely lost in the absence of Wnt9b in mice. In this study, we followed up with this interesting phenomenon and elucidated cellular and molecular mechanisms whereby Wnt9b facilitates Wolffian duct maintenance in male embryos. Wnt9b belongs to the WNT family of secreted proteins and is expressed in the Wolffian duct epithelium. We found that the Wolffian duct degeneration in Wnt9b-/- male embryos was accompanied with decreased cell proliferation in the epithelium but not in the mesenchyme during sexual differentiation. Wnt9b deletion did not impair testicular androgen synthesis but altered expression pattern of mesenchymal androgen receptor. The percentage of androgen receptor positive cells in the mesenchyme was significantly reduced, which can be the cause of decreased epithelial proliferation. Wnt9b actions can be transduced by both -catenin-dependent and -catenin-independent pathways in the context of target cells. Transcriptomic analysis of E12.5 Wnt9b+/+ and Wnt9b-/- mesonephroi revealed that expression of multiple WNT/-catenin-target genes was reduced in the absence of Wnt9b. Deletion of mesenchymal -catenin led to caudal Wolffian duct degeneration and cystic formation in the cranial region. Taken together, our study uncovers the important WNT9B-AR signaling axis mediating the epithelial-mesenchymal interaction in Wolffian duct development. To investigate the role of Wnt9b on Wolffian duct development and androgen action. In this model, we used a global knockout of Wnt9b. We prepared 3 Wnt9b+/+ and Wnt9b-/- male mesonephroi from mouse embryos at embryonic day E12.5.午非管（Wolffian duct，又称中肾管mesonephric duct）是胚胎发育的原基，可分化为附睾、输精管与精囊。胎儿发育阶段，午非管间充质所受的雄激素作用是维持午非管稳态的核心驱动因素，这一过程对雄性生育能力至关重要。然而，在缺失Wnt9b的小鼠中，雄激素维持午非管的能力完全丧失。 本研究针对这一有趣现象展开深入探究，阐明了Wnt9b促进雄性胚胎午非管维持的细胞与分子机制。Wnt9b属于分泌型WNT家族蛋白，在午非管上皮细胞中特异性表达。研究发现，在性别分化时期，Wnt9b敲除（Wnt9b-/-）的雄性胚胎中，午非管发生退化，伴随上皮细胞增殖水平显著降低，但间充质细胞增殖无明显变化。Wnt9b缺失并未损伤睾丸的雄激素合成功能，却改变了间充质雄激素受体（androgen receptor, AR）的表达模式：间充质内雄激素受体阳性细胞的占比显著下降，这可能是上皮细胞增殖受抑的关键诱因。 在靶细胞中，Wnt9b的信号转导可同时依赖β-连环蛋白（β-catenin）依赖与非依赖两条通路。对胚胎发育第12.5天（E12.5）的Wnt9b野生型（Wnt9b+/+）与敲除型（Wnt9b-/-）小鼠中肾组织的转录组分析显示，缺失Wnt9b后，多个WNT/β-连环蛋白靶基因的表达水平显著下调。进一步实验发现，间充质β-连环蛋白敲除会引发尾侧午非管退化，并在颅侧区域形成囊性结构。 综上，本研究揭示了调控午非管发育过程中上皮-间充质相互作用的关键WNT9B-AR信号轴。为探究Wnt9b在午非管发育及雄激素作用中的功能，本研究构建了Wnt9b全局基因敲除小鼠模型，并从胚胎发育第12.5天（E12.5）的小鼠胚胎中获取了3份Wnt9b野生型（Wnt9b+/+）与敲除型（Wnt9b-/-）雄性中肾组织。