single cell RNA sequencing of sham and 3 dpi spinal cords in mice

We report the application of single cell RNA sequencing technology for high-throughput profiling of sham and 3 dpi spinal cords in mice. By obtaining a total of 16051 cells from sham and 14295 cells from 3 dpi tissue, we identified 26 distinct cell clusters and 11 major cell types expressing known markers. These clusters comprised oligodendrocytes (Mbp, Mobp, Mog, Plp1), microglia (Gpr84, Cx3cr1, Csf1r), astrocytes (Gpr37l1, Bcan, Gfap), neutrophils (S100a9, Mmp9, S100a8), fibroblasts (Col1a1, Col1a2, Dcn), pericytes (Pdgfrb, Rgs5), endothelial cells (Flt1, Pecam1, Egfl7), macrophages (Ms4a7), monocytes (Ccr2, Cxcl3), b cells (CD19) and t cells (CD3e). neurons were not included because they were not expected to fit the sequencing protocol. We found that the most abundant cell type in the sham group was oligodendrocytes, accounting for 46.12%, while it decreased to 15.9% in the 3 dpi group. The second was microglia, accounting for 22.41%, which increased in number in the 3 dpi group, making it the most abundant cell in the SCI 3 dpi tissue (26.65%). The number of astrocytes also increased to 9.24% after SCI. Foreign immune cells such as neutrophils, macrophages, and monocytes accounted for 11.15%, 17.74%, and 9.41% of all cells after SCI, respectively. Among these, the percentage of pericytes decreased from 8.64% in the sham group to 1.28% in the 3 dpi group. The number of fibroblasts varied dramatically, rising rapidly from 0.33% in the sham group to 5.65% in the SCI 3 dpi group (Figure 2B). This study provides a framework for the application of single cell sequencing towards characterization of diverse cell populations in the sham and 3 dpi spinal cord. Single cell mRNA profiles of sham and 3dpi spinal cords (five male and five female C57/BL6 mice aged 8w in each group)

本研究报道了单细胞RNA测序（single cell RNA sequencing）技术在小鼠假手术组与脊髓损伤后3天（3 dpi）脊髓组织高通量转录组谱分析中的应用。本研究共从假手术组脊髓组织获取16051个细胞，从3 dpi损伤脊髓组织获取14295个细胞，通过已知标记基因鉴定出26个独立细胞簇与11种主要细胞类型。这些细胞簇包括：少突胶质细胞（oligodendrocytes，标记基因为Mbp、Mobp、Mog、Plp1）、小胶质细胞（microglia，标记基因为Gpr84、Cx3cr1、Csf1r）、星形胶质细胞（astrocytes，标记基因为Gpr37l1、Bcan、Gfap）、中性粒细胞（neutrophils，标记基因为S100a9、Mmp9、S100a8）、成纤维细胞（fibroblasts，标记基因为Col1a1、Col1a2、Dcn）、周细胞（pericytes，标记基因为Pdgfrb、Rgs5）、内皮细胞（endothelial cells，标记基因为Flt1、Pecam1、Egfl7）、巨噬细胞（macrophages，标记基因为Ms4a7）、单核细胞（monocytes，标记基因为Ccr2、Cxcl3）、B细胞（b cells，标记基因为CD19）以及T细胞（t cells，标记基因为CD3e）。神经元未纳入本次分析，因其不符合本测序实验的实验流程要求。

研究发现，假手术组中丰度最高的细胞类型为少突胶质细胞，占比达46.12%；而在3 dpi损伤组中其占比降至15.9%。其次为小胶质细胞，占假手术组细胞总数的22.41%；在3 dpi损伤组中其数量显著增加，占比达26.65%，成为脊髓损伤（Spinal Cord Injury, SCI）3 dpi损伤组织中丰度最高的细胞类型。脊髓损伤后，星形胶质细胞的占比亦升至9.24%。中性粒细胞、巨噬细胞与单核细胞等外来免疫细胞在脊髓损伤后分别占所有细胞的11.15%、17.74%与9.41%。其中，周细胞的占比从假手术组的8.64%降至3 dpi损伤组的1.28%。成纤维细胞的占比则出现显著变化：从假手术组的0.33%快速升高至SCI 3 dpi损伤组的5.65%（图2B）。

本研究为利用单细胞测序技术表征假手术组与3 dpi损伤组脊髓组织中的各类细胞群提供了研究框架。本数据集包含假手术组与3 dpi脊髓组织的单细胞mRNA转录组谱，每组均使用8周龄的C57/BL6小鼠各5只，雌雄各半。