Cdc42信号通路异常与先天性胆道闭锁发生的关系研究

The disorders of cell cytoskeleton and junction will lead to changes in cell polarity, which often result in dysplasia during the development process. Cdc42, as the center regulator of cell polarity, plays pivotal role in cytoskeleton and junction regulation. Biliary atresia (BA), a rare neonatal liver disease, seriously threats children's lives and the pathogenesis is far from elucidated. Viral infections and immune abnormalities have long been considered to be the main cause of BA. However, even though the means of antiviral and anti-immunotherapy are significantly improved, the incidence of BA has not been effectively controlled, suggesting that they may not be the initiating factors for pathological changes of biliary tract of BA. Although, it has been reported that the disruption of cell junction and polarity was existed in in BA-children’s liver, the intrinsic link between the disruption and BA presence is still unclear. In our previous study, we also detected the disorder of cell junction and polarity, and we firstly discovered the decrease of Cdc42 and its Rho ATPase activity, which may be due to the pathogenic mutations of multiple Cdc42 signaling pathways related genes, suggesting Cdc42 signaling pathway abnormality. In addition, we speculate that the toxic effect of bile is the power, which selectively aggravates the performance of abnormal Cdc42 signaling pathway on biliary tract. This study will aim to explore the relationship between the Cdc42 signaling pathway abnormalities and the BA occurrence. And our work will be helpful to unfold the mystery of BA occurrence and provide theoretical basis for clinical treatment.

先天性胆道闭锁（BA）是一种罕见新生儿肝脏疾病，严重威胁着患儿生命。长期以来，病毒感染和免疫异常被认为是BA发生的主要原因。然而，随着抗病毒和抗免疫治疗手段的显著进步，BA发病率却不降反升，提示二者可能不是胆道器质性病变的始动因素，更像是继发产生的。人们对BA中胆道器质性病变的分子机制还知之甚少。Cdc42调控的细胞骨架和胞间连接异常可导致细胞极性改变，并往往引起发育异常。在BA肝脏中，我们检测到细胞连接和极性蛋白表达紊乱、Cdc42表达水平及其GTPase酶活性明显下降，其原因可能与多个Cdc42信号通路相关基因的致病突变有关。进而我们推测胆汁的毒性作用可能是Cdc42通路异常仅表现为胆道损伤而非某种“综合征”的重要原因。本研究将通过临床标本、转基因小鼠模型和体外3D细胞培养模型，探讨Cdc42信号通路异常与BA发生的关系，为阐明BA发生机制及早期（产前）诊断提供理论依据。

先天性胆道闭锁 (BA) 是一种破坏性的罕见新生儿闭塞性胆管病，可影响肝内、肝外胆管，严重威胁患儿生命，同时为其家庭带来严重的精神和经济负担。尽管近年来一些研究报道了BA发生的相关机制，但很大程度上BA的发病机理仍是未知的。通过免疫荧光染色和蛋白质印迹探索了早期活检 BA 肝脏中的细胞连接和极性复合物，我们发现 Cdc42 作为关键的细胞连接和极性调节蛋白在 BA 肝脏中显着降低。因此，为了研究 Cdc42 在 BA 发育中的作用，我们构建了肝脏特异性和他莫昔芬诱导的胆管细胞特异性 Cdc42 缺失的转基因小鼠。我们进一步评估了胆汁酸在加重 Cdc42 不足小鼠肝脏胆道损伤中的作用，结果发现 BA 肝脏中胆管细胞和肝细胞中细胞连接和极性复合物的组装存在显着缺陷。这种缺陷的特点是细胞连接蛋白的无序定位，包括 ZO1、β-catenin、E-cadherin 和 claudin-3。 Cdc42 及其活性形式 Cdc42-GTP 作为一种小的 Rho GTPase 来协调极性复合物与 Par6/Par3/αPKC 的组装，在 BA 肝脏中显着减少。胎儿小鼠胆管细胞中的选择性 Cdc42 缺乏导致与人类 BA 肝脏中发现的相似的组织学变化，包括肝内和肝外胆管阻塞、上皮萎缩以及细胞连接和极性复合物的破坏。胆汁酸的减少显着改善了 Cdc42 突变小鼠的组织学和血清学指标。我们的研究结果表明，BA 与肝细胞中细胞连接和极性复合物的组装受损密切相关，这可能是由 Cdc42 不足引起的，并因胆汁酸腐蚀而加剧。