The leptomeninges play a pivotal role in the central nervous system (CNS), serving both as a barrier and as a conduit for fluid and cellular transport. Despite their critical functions, our understanding of leptomeningeal development and maturation during human embryogenesis remains limited. This study seeks to bridge this gap. We conducted single-nucleus RNA sequencing on leptomeningeal tissues from eight human embryos, capturing developmental stages from early fetal to late mid-fetal phases. Our bioinformatic analyses encompassed cell type classification, identification of layer-specific markers, mapping of the arachnoid barrier maturation trajectory, and joint analyses with mouse and aged human leptomeningeal scRNA datasets. Key bioinformatic findings were validated through immunostaining in selected samples. Our study revealed a complex cellular heterogeneity within the developing leptomeninges, identifying distinct subpopulations of fibroblasts, immune cells, and vascular cells. We mapped the transcriptomic dynamics of fibroblast cell types throughout fetal brain development, highlighting a clear maturation process from early fetal to late mid-fetal stages. Comparative analysis with mouse data allowed us to distinguish human-specific layer markers while confirming several conserved markers shared between humans and mice. Joint analysis with aged human datasets identified two unique arachnoid clusters specific to aging leptomeninges. Moreover, we traced the developmental trajectory of the arachnoid barrier, detailing the transcriptomic shifts associated with its gradual formation. Notably, immune cells in early fetal stages were predominantly M2-type macrophages, underscoring a distinctive immune environment. Finally, we explored the molecular interactions between fibroblasts and other cell types, highlighting their coordinated roles in orchestrating leptomeningeal development. Together, our findings provide a comprehensive overview of the cellular and molecular landscape of the developing human leptomeninges.

中文翻译：

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破译人类胎儿软脑膜的时间转录景观


软脑膜在中枢神经系统 （CNS） 中起着关键作用，既是屏障，也是液体和细胞运输的管道。尽管它们具有关键功能，但我们对人类胚胎发生过程中软脑膜发育和成熟的理解仍然有限。本研究旨在弥合这一差距。我们对 8 个人类胚胎的软脑膜组织进行了单核 RNA 测序，捕获了从胎儿早期到胎儿中期晚期的发育阶段。我们的生物信息学分析包括细胞类型分类、层特异性标记物的鉴定、蛛网膜屏障成熟轨迹的映射以及与小鼠和老年人类软脑膜 scRNA 数据集的联合分析。通过对选定样本进行免疫染色来验证关键的生物信息学发现。我们的研究揭示了发育中的软脑膜内复杂的细胞异质性，确定了成纤维细胞、免疫细胞和血管细胞的不同亚群。我们绘制了整个胎儿大脑发育过程中成纤维细胞类型的转录组动力学，突出了从胎儿早期到胎儿中期晚期的明确成熟过程。与小鼠数据的比较分析使我们能够区分人类特异性层标记，同时确认人类和小鼠之间共享的几个保守标记。与老年人类数据集的联合分析确定了两个独特的蛛网膜簇，特定于衰老的软脑膜。此外，我们追踪了蛛网膜屏障的发育轨迹，详细说明了与其逐渐形成相关的转录组学变化。值得注意的是，早期胎儿阶段的免疫细胞主要是 M2 型巨噬细胞，强调了独特的免疫环境。 最后，我们探讨了成纤维细胞与其他细胞类型之间的分子相互作用，强调了它们在协调软脑膜发育中的协调作用。总之，我们的研究结果全面概述了发育中的人类软脑膜的细胞和分子景观。