THE ROLE OF NOTCH SIGNALING IN BILE DUCT DEVELOPMENT

描述（由申请人提供）： 肝脏和胆管发育障碍是儿科人群发病率和死亡率的主要来源。对正常肝脏和胆管发育基因的鉴定可能在未来具有潜在的治疗意义。最近确定Jag 1为Alagille综合征（AGS）（一种以胆管缺乏为主要特征的多系统发育障碍）的疾病基因，暗示Notch信号通路对正常胆管分化至关重要。小鼠模型是阐明胚胎肝脏中发生的细胞相互作用的宝贵工具。 编码Notch途径中配体的Jag 1和编码Notch细胞表面受体之一的Notch 2突变的复合杂合小鼠表现出在Alagille综合征（AGS）患者中观察到的许多特征。这些小鼠表现出先天性心脏缺陷、肾小管异常、黄疸和胆管缺乏。我们建议详细研究这种小鼠模型，以验证其作为AGS模型，并阐明Notch信号在正常胆管发育中的作用。我们将检测突变肝脏的基因表达变化，这可能有助于识别参与这一过程的其他基因。 我们的办法是双重的;首先，我们建议对Jag 1-/Notch 2-动物胆管缺乏的进展进行详细的组织学分析。在Jag 1突变的患者中，我们经常观察到胆管增生之前出现的缺乏，我们怀疑类似的过程可能发生在小鼠模型。我们提出了一系列的实验，包括免疫组织化学，以确定胆管前体细胞，TUNEL检测，以确定凋亡细胞，并测定上调或下调其他基因已知参与胆管发育，如HNF 6或HNF 1（。其次，我们提出了一种微阵列的方法来比较基因表达谱之间的正常和Jag 1-/Notch 2-肝脏。通过这种方式，我们将确定Notch信号通路的下游靶点和其他可能在导致正常肝脏和胆道系统形成的复杂细胞相互作用中发挥关键作用的基因。

DESCRIPTION (provided by applicant): Disorders of liver and bile duct development are a major source of morbidity and mortality in the pediatric population. The identification of genes orchestrating normal liver and bile duct development could have potential therapeutic implications in the future. The recent identification of Jag1 as the disease gene for Alagille syndrome (AGS), a multisystem developmental disorder involving bile duct paucity as a cardinal feature, implicates the Notch signaling pathway as being crucial for normal bile duct differentiation. A mouse model is an invaluable tool for elucidating the cellular interactions that occur in the embryonic liver. A compound heterozygous mouse for mutations in Jag1, encoding a ligand in the Notch pathway, and Notch2, encoding one of the Notch cell surface receptors, exhibits many features seen in patients affected with Alagille syndrome (AGS). These mice demonstrate congenital heart defects, renal tubular abnormalities, jaundice and bile duct paucity. We propose to study this mouse model in detail in order to verify it as a model for AGS and to elucidate the role of Notch signaling in normal bile duct development. We will assay mutant livers for changes in gene expression that could help to identify other genes involved in this process. Our approach is twofold; first, we propose to perform a detailed histologic analysis of the progression of bile duct paucity in the Jag1-/Notch2- animals. In patients with Jag1 mutations, we often observe bile duct proliferation prior to the appearance of paucity, and we suspect that a similar process may occur in the mouse model. We propose a series of experiments including immunohistochemistry to identify bile duct precursor cells, TUNEL assays to identify apoptotic cells, and assays for up- or down-regulation of other genes known to be involved in bile duct development, such as HNF6 or HNF1(. Second, we propose a microarray approach to compare gene expression profiles between normal and Jag1-/Notch2- livers. In this way, we will identify downstream targets of the Notch signaling pathway and other genes that may play a crucial role in the complex cellular interactions that result in the formation of a normal liver and biliary system.