Caveolin-1/γ-catenin/GSK-3β通路调控巨噬细胞极化在腹膜透析相关腹膜纤维化的作用及机制研究

Peritoneal fibrosis is a crucial complication of long-term peritoneal dialysis, contributes to the development of peritoneal membrane failure, which leads to withdrawal from peritoneal dialysis. On the basis of our previous work that tamoxifen can effectively inhibit peritoneal fibrosis, this project is intended to be used in vitro and in vivo on the model of peritoneal fibrosis in mice. To observe the changes of endogenous Caveolin-1 protein expression and its effect on macrophage polarization related pathways. The proliferation and phenotypic transformation of human peritoneal mesothelial cells by macrophages and the clearance efficiency of mesothelial cells were used as the main indexes. To investigate the regulatory mechanism of Caveolin-1/γ-catenin/GSK-3β signaling pathway in peritoneal macrophages and peritoneal mesothelial cells on peritoneal fibrosis induced by high glucose peritoneal dialysate. The effect of this pathway on the occurrence and progression of the disease was verified in common disease models. To explore the regulatory mechanism of macrophage polarization in peritoneal fibrosis, in order to demonstrate the hypothesis that Caveolin-1 may be used as a target to reverse the process of macrophage polarization. It provides preliminary guidance for the mechanism of the treatment of peritoneal fibrosis and the selection and development of modified peritoneal dialysate for anti-fibrosis drugs such as Tamoxifen and its derivatives. 

腹膜纤维化是腹膜透析患者退出腹膜透析治疗的重要原因。本项目在我们前期工作观察到他莫昔芬能有效抑制腹膜纤维化的基础上，拟于体外细胞和体内小鼠腹膜纤维化模型上，观察内源性的Caveolin-1蛋白表达变化及其对巨噬细胞极化的影响。以巨噬细胞对人腹膜间皮细胞的增殖与表型转变和间皮细胞的清除效率为主要指标，明确腹膜Caveolin-1/γ-catenin/GSK-3β信号通路调控巨噬细胞极化在腹膜透析相关腹膜纤维化的调控机制；并在常见疾病模型中验证该通路对疾病发生及进展的作用，探索腹膜纤维化中巨噬细胞极化的相关调控机制，以论证Caveolin-1可能作为逆转巨噬细胞引起腹膜纤维化过程靶点的假说，对治疗腹膜纤维化的机制及抗纤维化药物如他莫昔芬及其衍生物的改良型腹膜透析液的选择和开发研究提供初步指导。

超滤衰竭是腹膜透析（PD）患者退出治疗的最主要原因，与透析液中高浓度葡萄糖、透析过程大量葡萄糖降解产物等引起腹膜纤维化（PF）有关。腹膜间皮细胞的丧失是腹膜功能衰竭组织病理学的重要特征表现。基于前期发现他莫昔芬（TAM）对上皮间充质转化过程中的作用，本项目通过小鼠PF模型和高糖诱导的间质上皮转化（EMT）细胞模型，探索了TAM对PF和腹膜间皮细胞EMT进程的作用和机制。.项目执行期间的研究发现：（1）通过高糖诱导建立了腹膜间皮细胞体外转分化的细胞模型，高糖刺激明显促进腹膜间皮细胞中ER和Vimentin的表达，抑制E-cadherin的表达。（2）TAM处理明显缓解高糖诱导的腹膜间皮细胞中EMT相关标志物的表达变化，提示TAM抑制高糖诱导的腹膜间皮细胞间充质转分化的作用。（3）通过高糖腹膜透析液诱导构建C57BL/6 小鼠腹膜纤维化模型，建模42天后的小鼠腹膜组织的厚度明显增加，间皮下组织疏松，有大量的胶原纤维沉积。（4）在小鼠模型中，TAM处理显著缓解小鼠腹膜纤维化的进程，腹膜厚度及胶原沉积明显改善，腹膜间质细胞EMT进程也明显缓解。（5）β-catenin/GSK-3β通路在腹膜纤维化及间皮细胞间质转化中明显被激活，而使用β-catenin抑制剂ICG-001处理，显著抑制了β-catenin/GSK-3β通路的激活，缓解腹膜间皮细胞的转分化及腹膜纤维化程度。（6）TAM对腹膜间皮细胞中β-catenin/GSK-3β通路具有重要的调节作用，TAM显著抑制了腹膜间皮细胞EMT过程中β-catenin/GSK-3β通路的激活，从而改善PF进程。.项目执行期间，共发表SCI论文5篇；申请并授权实用新型专利1项；协助培养硕士研究生1名。