Role and regulation of beta-catenin in cholestatic liver disease

 描述（由申请方提供）：原发性硬化性胆管炎（PSC）是一种病因不明的罕见慢性胆汁淤积性肝病，其特征为肝内和肝外胆管的进行性炎症和纤维化。目前，没有有效的医学治疗方法，肝移植仍然是终末期PSC患者延长生命的唯一治疗选择。尽管广泛的研究，PSC的发病机制仍不完全清楚。一种高度相关的机制是毒性胆汁酸（BA）在肝脏中的积累。我们最近已经证明，肝脏特异性β-连环蛋白敲除小鼠（Alb-cre β-连环蛋白KO; KO 1）在两种胆汁淤积模型（胆管结扎（BDL）和0.1% 3，5-二乙氧羰基-1，4-二氢-可力丁（DDC）饮食）中表现出肝损伤、纤维化和非典型小管增殖（ADP）的显著减少。此外，我们观察到减少总肝BA KO 1 BDL后，伴随着增强法尼醇X受体（FXR）的激活。这导致发现β-连环蛋白通过与细胞核中的FXR结合在调节BA合成和转运中的新作用。此外，β-连环蛋白的抑制激活了通常对BA刺激无反应的FXR库。因此，该提议的首要假设是，在胆汁淤积期间抑制β-连环蛋白可以通过物理抑制FXR活化来抑制BA生物合成，从而减轻损伤和疾病的进展。此外，抑制β-连环蛋白的策略可以潜在地与利用FXR激动剂的当前治疗剂结合使用，以增强治疗的有效性。在目标1中，我们将研究通过β-连环蛋白敲低降低BA毒性对小鼠胆汁淤积性肝损伤的影响。我们推测，通过β-连环蛋白抑制减少毒性BA将延迟梗阻性、化学性和遗传性胆汁淤积模型的发病或减轻疾病的严重程度。在目标2中，我们将描述FXR/β-连环蛋白联合在胆汁淤积中的作用和调节。这将通过首先通过使用FXR-β-连环蛋白双KO小鼠验证FXR活化作为BDL后的保护机制，然后分析胆汁淤积期间FXR/β-连环蛋白缔合和活性的动力学来实现。我们还将在体内和体外确定上游Wnt信号传导对FXR/β-连环蛋白缔合的贡献。在目标3中，我们将确定FXR/β-连环蛋白联合的治疗和预后相关性。首先，我们将直接比较β-连环蛋白与FXR激动剂联合的外源性抑制与单独使用激动剂在减轻胆汁淤积性损伤中的作用。最后，我们将在移植后PSC患者活检中将β-连环蛋白定位作为FXR活性的替代物与肝功能相关联。因此，在补助金中提出的研究将进一步我们的了解-连环蛋白在胆汁淤积性疾病，如PSC和预期的结果可能有显着的预后和治疗意义的作用。

 DESCRIPTION (provided by applicant): Primary sclerosing cholangitis (PSC) is a rare, chronic, cholestatic liver disease of unknown etiology characterized by progressive inflammation and fibrosis of the intrahepatic and extrahepatic biliary ducts. Currently, there are no effective medical therapies available, and liver transplantation remains the only life-extending treatment option for patients with end-stage PSC. Despite extensive research, the pathogenetic mechanisms of PSC are still incompletely understood. One mechanism that is highly relevant is the accumulation of toxic bile acids (BA) in the liver. We have recently demonstrated that liver-specific ß-catenin knockout mice (Alb-cre ß-catenin KO; KO1) exhibit a dramatic decrease in liver injury, fibrosis, and atypical ductular proliferation (ADP) in two models of cholestasis (bil duct ligation (BDL) and 0.1% 3,5- diethoxycarbonyl-1,4-dihydro-collidine (DDC) diet). Furthermore, we observed decreased total hepatic BA in KO1 after BDL, concomitant with enhanced farnesoid X receptor (FXR) activation. This led to the discovery of a novel role for ß-catenin in regulating BA synthesis and transport through association with FXR in the nucleus. Furthermore, inhibition of ß-catenin activates a pool of FXR that is normally unresponsive to BA stimulation. Thus, the overarching hypothesis of the proposal is that suppressing ß-catenin during cholestasis may alleviate injury and progression of the disease through inhibition of BA biosynthesis by physically inhibiting FXR activation. Further, strategies that suppress ß-catenin could potentially be used in conjunction with current therapeutics that utilize FXR agonists in order to enhance the effectiveness of treatment. In aim 1, we will investigate the impact of reduced BA toxicity through ß-catenin knockdown on murine cholestatic liver injury. We hypothesize that reduction of toxic BA through ß-catenin inhibition would delay the onset or alleviate the severity of disease in obstructive, chemical, and genetic models of cholestasis. In aim 2 we will characterize the role and regulation of FXR/ß-catenin association in cholestasis. This will be accomplished by first verifying FXR activation as the protective mechanism after BDL through use of FXR-ß-catenin double KO mice, and then analyzing the dynamics of FXR/ß-catenin association and activity during cholestasis. We will also determine the contribution of upstream Wnt signaling to FXR/ß-catenin association both in vivo and in vitro. In aim 3, we will determine the therapeutic and prognostic relevance of FXR/ß -catenin association. First, we will directly compare the exogenous inhibition of ß-catenin in conjunction with an FXR agonist to the use of agonist alone in alleviating cholestatic injury. Finally, we will correlate ß-catenin localization as a surrogate for FXR activity with hepatic function in PSC patient biopsies post-transplant. Thus, the proposed studies in the grant will further our understanding of the role of -catenin in cholestatic diseases such as PSC and the expected outcomes may have significant prognostic and therapeutic implications.