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.
