A cellular and spatial map of the choroid plexus across brain ventricles and ages

The choroid plexus (ChP) in each brain ventricle produces cerebrospinal fluid (CSF) and forms the blood-CSF barrier. We apply single cell and single nuclei sequencing to identify region and age specific shifts in gene expression of the constituent cell types of the choroid plexus. First, we sequenced whole cells (15,620) from each ventricle (lateral, third and fourth) of the embryonic mouse brain (Embryonic day (E) 16.5). Our analyses and validation of gene (smFISH) and protein (immunohistochemistry) expression combined with spatial mapping (confocal imaging) revealed the identity and location of major cell types, subtypes, proliferating cells, progenitor populations and regionalized gene expression, across each ventricle in the developing brain. Next, to track age-dependent shifts in the choroid plexus properties, we performed single nuclei sequencing (83,040) across each ventricle of the embryonic (E16.5), adult (4 months) and aging (20 months) mouse brain. Epithelial and mesenchymal cells showed regionalized gene expression patterns by ventricle, starting at embryonic stages and persisting with age. Dramatic transcriptional shifts were found with maturation (embryonic to adult) and a smaller shift within each aged cell type. With aging, epithelial cells upregulated host defense programs and resident macrophages enhanced expression of IL-1b signaling genes. Our atlas revealed ChP brain barrier cellular diversity, architecture and signaling across ventricles during development, maturation and aging. We sequenced whole cells isolated from lateral, third and fourth ventricle of embryonic brains (E16.5, CD-1) in triplicate. We sequenced whole nuclei isolated from each ventricle of embryonic (E16.5), adult (4 months post natal) and aging (20 months post natal) mouse (C57BL/6J) brains.每侧脑室中的脉络丛（Choroid Plexus, ChP）可生成脑脊液（cerebrospinal fluid, CSF），并构成血-脑脊液屏障。本研究采用单细胞与单细胞核测序技术，解析脉络丛组成细胞类型的区域特异性与年龄特异性基因表达变化。 首先，我们对胚胎小鼠脑（胚胎第16.5天，E16.5）的侧脑室、第三脑室与第四脑室的全细胞（共计15620个）进行了测序。通过对基因表达（单分子荧光原位杂交，smFISH）与蛋白表达（免疫组织化学，immunohistochemistry）的分析与验证，结合空间定位成像（共聚焦成像，confocal imaging），我们揭示了发育中小鼠脑各脑室中主要细胞类型、亚型、增殖细胞、祖细胞群的身份与定位，以及区域特异性基因表达模式。 其次，为追踪脉络丛特性的年龄依赖性变化，我们对胚胎期（E16.5）、成年期（4月龄）与衰老期（20月龄）小鼠脑的各脑室进行了单细胞核测序，共获取83040个细胞核。研究发现，上皮细胞与间充质细胞呈现出依脑室区分的区域特异性基因表达模式，该模式自胚胎阶段便已确立，并随年龄增长持续存在。在发育成熟阶段（胚胎期至成年期），细胞转录组发生显著改变；而各衰老细胞类型的转录组仅出现小幅变化。衰老过程中，上皮细胞上调了宿主防御相关通路的表达，驻留巨噬细胞则增强了IL-1β信号通路相关基因的转录水平。 本数据集图谱揭示了发育、成熟与衰老阶段中，各脑室脉络丛脑屏障的细胞多样性、组织结构与信号通路特征。我们对CD-1品系胚胎小鼠脑（E16.5）的侧脑室、第三脑室与第四脑室分离得到的全细胞进行了三次重复测序。同时，我们对C57BL/6J品系小鼠胚胎期（E16.5）、成年期（出生后4月龄）与衰老期（出生后20月龄）的各脑室分离得到的全细胞核进行了测序。