角膜缘干细胞移植是角膜缘干细胞缺乏有效的治疗方法，但供体角膜缘干细胞来源不足限制了其临床应用。胚胎干细胞来源的角膜上皮干细胞可作为捐献角膜缘组织的有效补充。胚胎时期角膜上皮起源于眼表外胚层。前期研究中我们已探明胚胎干细胞向眼表外胚层分化的调控机制，但眼表外胚层向角膜上皮分化这一关键环节的分子机制尚不清楚。预实验中我们通过单细胞测序及差异基因功能验证发现DKK2和BMP4可促进眼表外胚层向角膜上皮分化。DKK2是Wnt经典通路抑制剂，而Wnt经典通路可负性调控BMP4通路。预实验中我们还发现DKK2可使BMP4下游效应分子Smad1/5磷酸化激活。据此推测：DKK2通过阻断Wnt经典通路对BMP4通路的负性调控促进眼表外胚层向角膜上皮分化。本项目拟采用多组学分析和差异基因功能验证等方法对DKK2促进眼表外胚层向角膜上皮分化的作用及分子机制进行阐明，为角膜上皮发育和体外定向分化等研究提供线索。

Limbal stem cell transplantation is an effective treatment for limbal stem cell deficiency (LSCD), but the shortage of donor limbal stem cells limits its clinical application. Corneal epithelial stem cells derived from embryonic stem cells (ES) can be used as an alternative substitute of donated limbal tissue for the therapy of LSCD. Corneal epithelium originates from the ocular surface ectoderm (OSE) of the embryo. In previous studies, we have clarified the regulatory mechanism involved in the differentiation of OSE cells from ES. However, the molecular mechanism inducing differentiation of corneal epithelial cell from ocular surface ectoderm cells is still unclear, which has become the main limitation for treating LSCD with ES-derived corneal epithelial stem cells. In our pre-experiment, single cell sequencing and differential gene function test were performed, and the results showed that DKK2 and BMP4 could promote the differentiation of OSE cells into corneal epithelial cells. DKK2 is a canonical Wnt signaling pathway inhibitor, and BMP4 signaling pathway is negatively regulated by canonical Wnt signaling pathway in corneal epithelial cell. We also found that DKK2 can activate BMP4 signaling pathway by increasing Smad1/5 phosphorylation in our pre-experiment. We hypothesize that DKK2 promotes the differentiation of OSE cells into corneal epithelial cells by blocking the negative regulation of canonical Wnt signaling pathway on BMP4 signaling pathway. The purpose of this study is to investigate the role of DKK2 in the differentiation of OSE cells into corneal epithelial cells and involved molecular mechanisms, and to provide clues for the studies on corneal epithelial development and directed differentiation of corneal epithelial cells from pluripotent stem cells in vitro.