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Abstract：
Congenital hepatic fibrosis (CHF), a genetic cholangiopathy characterized by fibropolycystic changes in the biliary tree, is caused by mutations in the PKHD1 gene, leading to defective fibrocystin (FPC), changes in planar cell polarity (PCP) and increased β-catenin-dependent chemokine secretion. In this study, we aimed at understanding the role of Scribble (a protein involved in PCP), Yes-associated protein (YAP), and β-catenin in the regulation of the fibroinflammatory phenotype of FPC-defective cholangiocytes. Immunohistochemistry showed that compared with wild type (WT) mice, in FPC-defective (Pkhd1del4/del4 ) mice nuclear expression of YAP/TAZ in cystic cholangiocytes, significantly increased and correlated with connective tissue growth factor (CTGF) expression and pericystic fibrosis, while Scribble expression on biliary cyst cells was markedly decreased. Cholangiocytes isolated from WT mice showed intense Scribble immunoreactivity at the membrane, but minimal nuclear expression of YAP, which conversely increased, together with CTGF, after small interfering RNA (siRNA) silencing of Scribble. In FPC-defective cholangiocytes, inhibition of YAP nuclear import reduced β-catenin nuclear expression, and CTGF, integrin β6, CXCL1, and CXCL10 mRNA levels, whereas inhibition of β-catenin signaling did not affect nuclear translocation of YAP. Notably, siRNA silencing of Scribble and YAP in WT cholangiocytes mimics the fibroinflammatory changes of FPC-defective cholangiocytes. Conditional deletion of β-catenin in Pkhd1del4/del4 mice reduced cyst growth, inflammation and fibrosis, without affecting YAP nuclear expression. In conclusion, the defective anchor of Scribble to the membrane facilitates the nuclear translocation of YAP and β-catenin with gain of a fibroinflammatory phenotype. The Scribble/YAP/β-catenin axis is a critical factor in the sequence of events linking the genetic defect to fibrocystic trait of cholangiocytes in CHF.
摘要：
先天性肝纤维化（CHF），一种遗传性胆管病变，其特征是胆道纤维多囊性改变，是由PKHD1基因突变引起的,导致有缺陷的纤维囊素（FPC），平面细胞极性（PCP）的变化和β-连环蛋白依赖性趋化因子分泌的增加。在这项研究中,我们的目的是了解Scribble（一种参与PCP的蛋白质）的作用，是相关蛋白（YAP），和β-连环蛋白在调节FPC缺陷型胆管细胞的纤维炎症表型中的作用。免疫组织化学结果显示,与野生型(WT)小鼠相比,在FPC缺陷（Pkhd1del4/del4）小鼠囊性胆管细胞中YAP/TAZ的核表达，显着增加，并与结缔组织生长因子（CTGF）表达和囊周纤维化相关，而Scribble在胆管囊肿细胞上的表达明显降低。从WT小鼠中分离的胆管细胞在膜上显示出强烈的Scribble免疫反应性，但是YAP的核表达很少，反过来增加，连同CTGF,小干扰RNA（siRNA）沉默后的Scribble。在FPC缺陷的胆管细胞中，抑制YAP核输入减少β-catenin核表达，和CTGF,整合素β6、CXCL1和CXCL10mRNA水平，而β-连环蛋白信号的抑制不影响YAP的核易位。值得注意的是,WT胆管细胞中Scribble和YAP的siRNA沉默模拟FPC缺陷型胆管细胞的纤维炎症变化。Pkhd1del4/del4小鼠中β-catenin的条件性缺失减少了囊肿生长，炎症和纤维化，不影响YAP核表达。总之,Scribble在膜上的缺陷锚定促进了YAP和β-catenin的核易位，并获得了纤维炎性表型。Scribble/YAP/β-catenin轴是将CHF中的遗传缺陷与胆管细胞的纤维囊性特征联系起来的事件序列中的关键因素。
