DESCRIPTION (provided by applicant): Our long-term objectives remain to apply the fundamental concepts and broad technologies of cell and molecular biology to understand hepatic epithelial cell function and dysfunction. We continue to focus on cholangiocytes, the epithelial cells lining intrahepatic bile ducts, because of their biologic and clinical importance, and because of the new concepts, hypotheses, and techniques we have developed to study, cholangiocyte pathobiology, an underserved area of liver research. Recent evidence from our lab indicates that: (i) aquaporins (AQPs), a family of water channels, are important in ductal bile formation; and (ii) cholangiocytes contain primary cilia that act as sensory organelles and participate in normal bile formation and in biliary cystogenesis. Thus, we will test the central hypothesis that ductal bile formation: (i) is the net result of solute-driven, passive movement of water molecules through AQPs constitutively expressed in or recycled among distinct cellular compartments; (ii) is influenced by luminal mechanical, chemical, and osmotic signals sensed via primary cilia on the apical cholangiocyte membrane; and (iii) is abnormal in genetic spontaneous or experimental animal models of autosomal recessive polycystic kidney disease (ARPKD) when cholangiocyte ciliary structure and/or function is disturbed. Our three distinct but integrated specific aims test the hypotheses that: (i) ductal bile formation involves the normal function of primary cilia expressed on the apical membrane of each cholangiocyte to detect mechanical (e.g., bile flow rate), chemical (e.g., nucleotides, bile acids, glucose), and/or osmotic (e.g., bile hypo/hyperosmolarity) signals from bile; (ii) cellular expression, compartmentalization, and recycling of key 'flux' proteins (e.g., AQPs, AE2, CFTR) regulating ductal bile formation are influenced by ciliary stimulation; and (iii) abnormalities in structure, expression, and/or cellular localization of cilia-associated proteins (e.g., fibrocystin, the protein product of PKHD1, the gene mutated in ARPKD) contribute to disturbances in cholangiocyte water, solute, and ion transport promoting biliary cystogenesis. We will employ established and new methods, models, and probes, including: perfused bile duct units, isolated biliary cysts, isolated cholangiocyte cilia, spontaneous (i.e., the PCK rat) and transgenic (i.e., fibrocystin knockout mouse) animal models of ARPKD, gene silencing using small-interfering RNAs (siRNAs), novel cholangiocyte culture systems, and innovative morphologic techniques. Our results will further clarify the role of AQPs in cholangiocyte bile formation, address directly the potential importance of cholangiocyte cilia in ductal bile production, and explore the relationship of cholangiocyte cilia to possible disturbances of water, ion, and solute transport in biliary cystogenesis. Innovative aspects of our program include novel methodologies and animal models, and new concepts regarding the importance of cholangiocyte AQPs and cilia in ductal bile formation and biliary cystogenesis. We will generate information to yield new insights into normal cholangiocyte function, explore highly promising, selected aspects of cholangiocyte dysfunction, and continue to provide a broad theoretical framework for understanding and managing the cholangiopathies, a group of genetic and acquired hepatobiliary diseases in which the cholangiocyte is the principal target of diverse pathologic processes.
|Effective start/end date||12/1/78 → 3/31/18|