Hedgehog-Wnt信号通路参与特发性肺纤维化发病的分子机制探讨及疾病干预的基础研究

Idiopathic pulmonary fibrosis (IPF) is a devastating lung disease of unknown cause, characterized by pulmonary interstitial fibrosis. There was no significant benefit in clinical treatment, and patients have a median survival of less than 3 years after diagnosis. In our previous study, we found that excessive activation of the Wnt signaling pathway can promote fibroblast differentiation of lung mesenchymal stem cells and facilitate the development of pulmonary fibrosis. The further study showed that featured Wnt proteins were significantly overexpressed during the differentiation of lung mesenchymal stem cells to fibroblast. As known that the expression of Wnt proteins are regulated by the Hedgehog signaling pathway, we speculated that ascertain its regulatory mechanism may help us to find specific targets that inhibit IPF effectively and provide new ideas for clinical treatment. On the basis of preliminary work, the project is proposed to carry out the following research: ①Ascertain the molecular mechanism that the overexpression of Wnt proteins were activated by Hedgehog signaling pathway in lung mesenchymal stem cells. ②Screen the overexpressed featured GLI proteins and key Wnt proteins during the differentiation of human lung mesenchymal stem cells to fibroblast and search specific targets that inhibit IPF effectively. ③Seek specific targeting inhibitors to prove that targeting block Hedgehog signaling pathway can inhibit the differentiation of lung mesenchymal stem cells to fibroblast, as well as the occurrence and development of IPF.

特发性肺纤维化（IPF）是一种病因不明，以肺间质纤维化为特征的疾病，其治疗效果差，确诊后患者中位生存期不到3年。我们在前期研究中发现，Wnt信号通路过度激活可促进肺间质干细胞向成纤维细胞分化，加快IPF进程。进一步研究表明，在肺间质干细胞向成纤维细胞分化过程中特定Wnt蛋白高表达。已知Wnt蛋白表达受Hedgehog信号通路调控，提示探明该通路调控机制，可能有利于找到特异性抑制IPF发生的有效靶点，为临床治疗提供新思路。本项目拟在前期工作基础上，开展以下研究：①探明肺间质干细胞内Hedgehog信号通路激活特定Wnt蛋白高表达导致肺纤维化发生的分子机制；②筛选人肺间质干细胞向成纤维细胞分化过程中高表达的特定GLI蛋白和关键Wnt蛋白，寻找合理有效的IPF干预靶点；③寻找特异性靶向抑制剂，证明靶向阻断Hedgehog-Wnt通路，可特异性抑制肺间质干细胞向成纤维细胞分化，抑制IPF发生发展。

特发性肺纤维化（IPF）是一种病因不明的老年性肺间质疾病，目前该病的发病率呈明显上升趋势，而临床上尚无有效的治疗手段，患者确诊后的五年生存率不足50%。成纤维细胞的大量聚集是导致肺纤维化发生的直接原因，我们的前期研究发现Wnt信号通路激活能够诱导肺间质干细胞向成纤维细胞分化，加速肺纤维化发生；进一步研究表明Wnt信号通路的激活可能受到Hedgehog信号通路的调控。本项目在前期工作基础上，开展了以下研究：①探明肺间质干细胞内Hedgehog信号通路激活特定Wnt蛋白高表达，导致肺纤维化发生的分子机制；②筛选在人肺间质干细胞向成纤维细胞分化过程中高表达的特定GLI蛋白和Wnt蛋白，确定合理有效的IPF干预靶点；③通过多位点筛选，寻找特异性靶向抑制剂，验证靶向阻断Hedgehog-Wnt信号通路能够特异性抑制肺间质干细胞向成纤维细胞分化，抑制肺纤维化发生发展。研究表明，在肺间质干细胞向成纤维细胞分化及肺纤维化发生过程中Hedgehog-Wnt信号通路处于过度激活状态，其中Wnt信号通路上游配体Wnt7b、Wnt10a及其受体Fzd9和Fzd10特异性高表达，Hedgehog信号通路下游关键因子Gli1和Gli2的表达特异性升高；进一步实验确认Gli1能够通过与Wnt7b和Wnt10a基因的启动子区结合促Wnt7b和Wnt10a的表达，高表达的Wnt7b和Wnt10a则能够与下游的Fzd10受体特异性结合，介导Wnt信号通路的活化，进而促进肺间质干细胞向成纤维细胞分化；通过建立人肺间质干细胞-成纤维细胞诱导分化体系，筛选发现Wnt10a和Gli1分别是肺间质干细胞向成纤维细胞分化过程中人与小鼠共同高表达的特异蛋白，且上述实验结果在IPF病人肺组织中得到验证；最后我们以Gli1、Wnt10a及其相应的Fzd10受体为靶点进行干预，证实阻断Hedgehog-Wnt信号通路能够特异性抑制肺间质干细胞向成纤维细胞分化，抑制IPF发生发展。研究成果以学术论文形式发表于SCI收录期刊11篇，为进一步阐明IPF的发病机制及临床治疗IPF、发展抗IPF生物药物提供了新的思路和理论支撑。