Corneal Nonmyelinating Schwann Cells Illuminated by Single Cell Transcriptomics and Visualized by Protein Biomarkers

The cornea is the most innervated tissue in the human body. Myelinated axons upon inserting into the peripheral corneal stroma lose their myelin sheaths and continue into the central cornea wrapped by only nonmyelinating corneal SCs (nm-cSCs). This anatomical organization is believed to be important for central vision. Here we employed single-cell RNA sequencing (scRNA-seq), microscopy, and transgenics to characterize these nm-cSCs of the central cornea. Using principal component analysis, uniform manifold approximation and projection, and unsupervised hierarchal cell clustering of scRNA-seq data derived from central corneal cells of male rabbits, we successfully identified several clusters representing different corneal cell types, including a unique cell cluster representing nm-cSCs. To confirm protein expression of cSC genes, we performed cross-species validation, employing corneal whole mount immunostaining with confocal microscopy in mouse corneas. We expect that our results will advance the future study of nm-cSCs in applications of nerve repair, and provide a resource for the study of corneal sensory function. Overall design: Examination of non-myelinating Coreneal Schwann Cells through single cell RNA sequencing

角膜是人体中神经支配最为丰富的组织。有髓轴突插入外周角膜基质后会丧失髓鞘，随后仅由非髓鞘化角膜施万细胞（nonmyelinating corneal SCs, nm-cSCs）包裹，并继续向中央角膜延伸。这种解剖结构被认为对中央视觉功能具有重要意义。本研究采用单细胞RNA测序（single-cell RNA sequencing, scRNA-seq）、显微成像及转基因技术，对中央角膜的nm-cSCs进行了系统表征。通过对雄性家兔中央角膜细胞的scRNA-seq数据开展主成分分析、均匀流形近似与投影（uniform manifold approximation and projection, UMAP）以及无监督分层细胞聚类，我们成功鉴定出多个代表不同角膜细胞类型的细胞簇，其中包含一个独特的、对应nm-cSCs的细胞簇。为验证角膜施万细胞相关基因的蛋白表达情况，我们开展了跨物种验证实验，对小鼠角膜进行整体铺片免疫染色结合共聚焦显微镜观察。我们预期本研究结果将推动nm-cSCs在神经修复领域的后续研究，并为角膜感觉功能的相关研究提供宝贵的资源。整体实验设计：通过单细胞RNA测序技术对非髓鞘化角膜施万细胞进行分析鉴定。