Single-cell transcriptomic profiling of the neonatal oviduct and uterus reveals new insights into Mullerian duct regionalization

The upper Mullerian duct (MD) is patterned and specified into two morphologically and functionally distinct organs, the oviduct and uterus. It is known that this regionalization process is instructed by inductive signals from the adjacent mesenchyme. However, the interaction landscape between epithelium and mesenchyme during upper MD development remains largely unknown. Here, we performed single-cell transcriptomic profiling of mouse neonatal oviducts and uteri at the initiation of MD epithelial differentiation (postnatal day 3). We identified major cell types including epithelium, mesenchyme, pericytes, mesothelium, endothelium, and immune cells in both organs with established markers. Moreover, we uncovered region-specific epithelial and mesenchymal subpopulations and then deduced region-specific ligand-receptor pairs mediating mesenchymal-epithelial interactions along the craniocaudal axis. Unexpectedly, we discovered a mesenchymal subpopulation marked by neurofilaments with specific localizations at the mesometrial pole of both the neonatal oviduct and uterus. Lastly, we analyzed and revealed organ-specific signature genes of pericytes and mesothelial cells. Taken together, our study enriches our knowledge of upper Mullerian duct development, and provides a manageable list of potential genes, pathways, and region-specific cell subtypes for future functional studies. Oviducts and uteri were collected from CD1 neonates at PND3 and dissociated to achieve single cell suspension. Two replicates for each organ were pooled from the same 3 and 4 neonates米勒管上段（upper Mullerian duct, MD）可通过模式形成与细胞特化过程，分化为形态与功能均显著不同的两个器官：输卵管（oviduct）与子宫（uterus）。已有研究表明，该区域化过程由邻近间充质（mesenchyme）发出的诱导信号所调控，但目前学界对米勒管上段发育过程中上皮细胞与间充质细胞间的互作调控网络仍知之甚少。本研究针对小鼠米勒管上皮细胞分化起始阶段（出生后第3天，postnatal day 3, PND3）的新生输卵管与子宫组织，开展了单细胞转录组分析（single-cell transcriptomic profiling）。我们通过已验证的分子标记，在两种器官中鉴定出上皮细胞、间充质细胞、周细胞（pericytes）、间皮细胞（mesothelium）、内皮细胞（endothelium）与免疫细胞等主要细胞类群。此外，我们还发现了区域特异性的上皮与间充质细胞亚群，并据此推导了沿头尾轴（craniocaudal axis）介导间充质-上皮互作的区域特异性配体-受体对。出乎意料的是，我们鉴定到一类以神经丝蛋白（neurofilaments）为标记的间充质细胞亚群，该亚群特异性定位于新生输卵管与子宫的系膜侧缘（mesometrial pole）。最后，我们通过分析揭示了周细胞与间皮细胞的器官特异性特征基因。综上，本研究丰富了我们对米勒管上段发育机制的认知，并为后续功能研究提供了一份可操作性强的潜在基因、信号通路及区域特异性细胞亚群列表。本研究的实验材料为CD1品系新生小鼠在PND3时期采集的输卵管与子宫组织，经解离后获得单细胞悬液；每个器官设置两个生物学重复，分别混合3只与4只新生小鼠的组织样本。