Roles for Pkd1l1 in bile duct development

项目总结 胆道闭锁(BA)是1岁以下婴儿进行任何实质性器官移植的主要适应症。 几十年来一直回避病因学和病理生理学的重大发现。BA是一种新生儿肝病，最好是 以肝内外胆管纤维炎性阻塞为特征。通过外显子 具有异向(偏侧)特征的BA个体(具有BA脾的个体)的子集的测序 畸形综合征)，一些参与者被确认为睫毛基因的双等位破坏性突变 PKD1L1(多囊肾病1样1)。探讨PKD1L1受损的机制后果 在人类的信号传递中，我们培育了一种肝内胆管细胞限制性Pkd1l1Fl/Fl；AFP-Cre(LKO)小鼠。 最近公布的数据表明，在发育中的小鼠肝脏中缺乏Pkd1l1会导致早期胆汁 成年时形态异常和胆管周围纤维炎症增强，更多发生在远端环境中 胆管结扎后梗阻(BDL)。这些组织学特征强烈地模仿了人类BA肝脏中的特征， 提供了一个新的机会，以发现特定的细胞和分子洞察BA的快速和深刻 发病机制。为了更彻底地理解，必须揭示两个范例 Pkd1l1，从而阐明了BA的分子发病机制。第一，在发展中缺少Pkd1l1的后果 导致成年期胆管纤维炎症的胆道树未知。第二，Pkd1l1的作用机制 胆管细胞内的信号，当不存在时，有助于胆道病理，仍有待阐明。 这一建议中最重要的假设是Pkd1l1是胆汁正常发育所必需的。 和信号。这一假设将通过以下两个目标进行检验。目标1探索了 以及发育中的产前和出生后早期肝脏胆管畸形的特征。这一目标 将利用最近开发的Pkd1L1Null/Fl(NullF1)和Pkd1L1Null/Fl；AFP-cre(NullLKO)小鼠品系。目标2是 用分离的胆管细胞研究一套确定Pkd1l1-相互作用因子和信号的体外实验 结果，特别是对各种胆汁酸的反应，在分离的Pkd1L1F1和LKO胆管细胞中。 综上所述，我们预计这两个目标将发现胆道疾病的新的细胞和分子机制。 肌束发育和信号传递。此外，来自这些基于Pkd1l1的鼠标模型的信息将 帮助提供支持性的临床前证据，以解决目前缺乏有效的医疗疗法的问题 巴。

PROJECT SUMMARY Biliary atresia (BA) is the main indication to transplant any solid organ in infants less than 1 year of age and has eluded major discoveries of etiology and pathophysiology for decades. BA is a neonatal liver disease that is best characterized by fibroinflammatory obstruction of both intra- and extrahepatic bile ducts. Through exome sequencing of a subset of BA individuals with heterotaxic (laterality) features (those with the BA Splenic Malformation syndrome), several participants were identified with biallelic damaging mutations in the ciliary gene PKD1L1 (Polycystic kidney disease 1 like 1). To explore mechanistic consequences of impaired PKD1L1 signaling in humans, we developed an intrahepatic cholangiocyte-restricted Pkd1l1Fl/Fl;Afp-Cre (LKO) mouse. Recently published data indicates that the absence of Pkd1l1 in the developing mouse liver leads to early biliary dysmorphology and enhanced peribiliary fibroinflammation at adult ages, moreso in the setting of distal obstruction after bile duct ligation (BDL). These histologic features strongly mimic those seen in human BA livers, offering a novel opportunity to discover specific cellular and molecular insights into BA’s rapid and profound pathogenesis. There are two paradigms that must be uncovered to provide a more thorough understanding of Pkd1l1, and thus the molecular pathogenesis of BA. First, the consequences of absent Pkd1l1 in the developing biliary tree that leads to adulthood biliary fibroinflammation are unknown. Second, the mechanism of Pkd1l1 signaling within cholangiocytes, which when absent contributes to biliary pathology, remains to be elucidated. The overarching hypothesis in this proposal is that Pkd1l1 is required for proper biliary development and signaling. This hypothesis will be tested through the following two aims. Aim 1 explores the delineation and characterization of early bile duct dysmorphology in developing prenatal and early postnatal livers. This aim will utilize the recently developed Pkd1l1Null/Fl (NullFl) and Pkd1l1Null/Fl;Afp-cre (NullLKO) mouse lines. Aim 2 is an in vitro set of experiments with isolated cholangiocyte studies to define Pkd1l1-interactors and signaling consequences, specifically in response to various bile acids, in isolated Pkd1l1Fl/Fl and LKO cholangiocytes. Taken together, we anticipate that these two aims will discover new cellular and molecular mechanisms of biliary tract development and signaling. In addition, information stemming from these Pkd1l1-based mouse models will help provide supportive pre-clinical evidence to address the current paucity of effective medical therapeutics in BA.