R-spondin1分子内Furin样序列调节肠隐窝干细胞自更新及其增殖的分子机制研究

R-spondin1蛋白作为一种重要配体与Frizzled/LRP6、Lgr5和Lgr6受体结合并激活经典Wnt信号通路，它的激活能有效促进肠隐窝干细胞（CBC细胞）的自更新及增殖。异体CBC细胞移植治疗可以修复损伤的肠粘膜。但目前CBC细胞培养效率低下且耗费昂贵。Furin样序列作为R-spondin1的保守功能活性区域，其调节CBC细胞增殖的作用及机制不清。本课题拟通过比较Furin样序列的功能片段、Furin样序列删除突变的R-spondin1和完整的R-spondin1对Wnt信号的激活而验证Furin样序列的生物学功能；生物合成含Furin样序列的多肽与R-spondin1比较它们调节CBC细胞的自更新及增殖能力；并对其合成肽进行四分枝修饰，以期取代R-spondin 1优化CBC细胞培养体系。研究为CBC细胞扩增及培养条件的优化提供实验依据，为肠粘膜损伤性疾病的治疗提供新思路。

As an important ligand binding with Frizzled/LRP6 Lgr5 and Lgr6 receptor, The R-spondin1 protein activates the canonical Wnt signaling pathway that is effective in promoting self-renewal and proliferation of intestinal crypt stem cells (the CBC cells). Allogeneic CBC cell transplant therapy can repair the damaged intestinal mucosa, But the CBC cell culture is inefficient and expensive. As the conserved active region of the R-spondin1 protein, Furin-like sequence regulates the CBC cell proliferation, but the mechanism is unclear. This subject contemplated to comparise the Wnt signal activation of Furin-like sequence functional fragment, Furin-like sequence deleting mutation of the R-spondin1 and the complete R-spondin1, and verify on the biological function of Furin-like sequence. Whatmore, biosynthesis peptide of Furin-like sequence regulates self-updating and proliferation capacity of CBC cell. And four branches modification peptides of Furin-like sequence are synthesised trying to replace R-spondin 1 on CBC cell culture system. Our study will provide experimental evidence for optimization of CBC cell culture conditions and new ideas for the treatment of diseases of the intestinal mucosal injury.