筛选小分子诱导角质细胞逆分化为ESCs促进糖尿病难愈创面的愈合作用和机制研究

研究证明表皮干细胞（ESCs）可以促进糖尿病难愈创面（DW）修复，可是其在创面的数量极少。因此如何获取足够数量的ESCs是治疗DW的关键。既往研究发现许多分化成熟的细胞在特定的诱导条件下可以通过逆分化形成干细胞，提示分化细胞具有很强的可塑性。我们前期研究也发现多个可以体外诱导角质细胞（Krt10+）向ESCs（Krt14+）逆分化的功能性小分子，但其作用及机制均需要进一步明确。本项目将在此基础上应用体外细胞逆分化筛选模型完成多个小分子库的高通量筛选，并通过我们自主建立的双CRE细胞体内追踪体系进行体内逆分化效果的验证。同时我们将采用RNA-Seq测序比对分析寻找逆分化过程中的差异基因和信号通路，随后利用siRNA和cDNA文库验证，最后建立表皮类器官模型体系及转基因小鼠进一步体内验证这些核心调控基因的作用，藉此阐明角质细胞体内逆分化的调控机制，从而为DW的治疗提供新的理论依据和治疗靶点。

Previous studies have shown that epidermal stem cells (ESCs) can enhance the repair of refractory Diabetic wound （DW）, but only limited numbers of ESCs are found in the wounds. And thus, the key for treating DW is to get enough ESCs. It has been reported that many mature cells can be induced to dedifferentiate into stem cells under specific conditions, suggesting that differentiated cells have strong plasticity. In the preliminary study of this project, several functional small molecules that can induce keratinocytes (Krt10+) to dedifferentiate into ESCs (Krt14+) have been found. However, its effect and mechanism need to be further studied. Based on our previous findings, this project will apply the in vitro dedifferentiation screening model to find out the functional small molecule and use the double CRE cells intracellular tracking system which was previously established by the research team to verify the effect of dedifferentiation. Meanwhile, we adopt RNA-Seq method to identify differential genes and signaling pathways in the process of dedifferentiation, then we use siRNA and cDNA library to validate, and finally establish the epidermis organ model system and genetically engineered mice to verify the functions of these possible regulatory gene in vivo, thus explain the regulatory mechanism of the dedifferentiation in keratinocyte, and provide new theoretical basis and therapeutic targets for the treatment of DW.