Identification of Novel Molecular Markers through Transcriptomic Analysis in Human Fetal and Adult Corneal Endothelial Cells. Homo sapiens

Corneal endothelium is composed of a monolayer of corneal endothelial cells (CECs) in the inner layer of cornea, which is essential for maintaining corneal transparency. In order to better characterize CECs in different developmental stages, we profiled mRNA transcriptomes in human fetal and adult corneal endothelium with the goal to identify novel molecular markers in these cells. By comparing CECs with 12 other types of tissues, we identified 245 and 284 signature genes that are highly expressed in fetal and adult CECs, respectively. Functionally, these genes are characteristic of CECs, involving in cell adhesion, proteoglycan and sulfur metabolic process. Importantly, several of these genes are disease target genes in hereditary corneal dystrophies, consistent with their functional significance in CEC physiology. By comparing fetal and adult CECs, we also identified stage-specific markers associated with CEC maturation, such as the activation of the Wnt pathway genes in fetal, but not in adult CECs. Lastly, by immunohistochemistry of ocular tissues, we further confirmed the unique protein expression patterns for Wnt5a, S100A4, S100A6, and IER3, the four novel markers for either fetal or adult CECs. The identification of a new panel of molecular markers for fetal and mature CECs would be very useful for characterizing and quality controlling CECs through ex vivo expansion or stem cell differentiation for cell replacement therapy. Overall design: mRNA profile between adult and fetal CECs by high-throughput sequencing

角膜内皮（corneal endothelium）由角膜内层的单层角膜内皮细胞（corneal endothelial cells, CECs）构成，是维持角膜透明度的关键结构。为更好地表征不同发育阶段的角膜内皮细胞，我们对人类胎儿与成人角膜内皮细胞的mRNA转录组进行了测序分析，以期鉴定这类细胞中的新型分子标志物。通过将角膜内皮细胞与其余12种组织进行对比，我们分别筛选出在胎儿及成人角膜内皮细胞中高表达的245个和284个特征基因。功能层面，这些基因具备角膜内皮细胞的功能特征，参与细胞黏附、蛋白聚糖及硫代谢过程。值得注意的是，其中多个基因是遗传性角膜营养不良的疾病靶标基因，这与其在角膜内皮细胞生理活动中的功能重要性高度契合。通过对比胎儿与成人角膜内皮细胞，我们还鉴定出与角膜内皮细胞成熟相关的阶段特异性标志物，例如胎儿角膜内皮细胞中存在Wnt通路基因的激活，而成人角膜内皮细胞中并无此类激活。最后，我们通过眼部组织免疫组织化学（immunohistochemistry）检测，进一步验证了Wnt5a、S100A4、S100A6及IER3这四种分别针对胎儿或成人角膜内皮细胞的新型标志物的独特蛋白表达模式。鉴定胎儿与成熟角膜内皮细胞的新型分子标志物组合，将为通过体外扩增或干细胞分化获取的角膜内皮细胞的特性鉴定与质量控制提供重要支撑，以应用于细胞替代治疗。总体实验设计：采用高通量测序分析成人与胎儿角膜内皮细胞的mRNA表达谱。