Regulation of cholangiocytes by InsP3 receptor isoforms

描述（由申请人提供）：胆汁分泌是肝脏的主要功能之一。为了维持胆汁流动，不仅肝细胞必须分泌胆汁，而且这必须随后被胆管上皮细胞或胆管细胞进一步修饰和调节。胆管细胞功能异常导致胆汁淤积，这是肝脏疾病的主要表现。胆汁淤积性肝病占美国肝移植的20%，是儿科移植患者中最常见的肝病原因。此外，胆管细胞功能异常是导致囊性纤维化的肝脏表现的原因，囊性纤维化是最常见的遗传性疾病之一。 胆管细胞的胆汁分泌部分受胞质Ca ~（2+）调节。一般来说，细胞受随时间推移的Ca 2+信号模式和Ca 2+信号发生的细胞区域的调节。然而，很少有人知道的时间或空间方面的Ca 2+信号在胆管细胞，也不知道这些Ca 2+信号是如何调节。肌醇1，4，5-三磷酸受体（InsP 3R）介导上皮细胞中的Ca 2+信号传导，并且胆管细胞表达该受体的所有三种亚型。这一建议的假设是，钙信号在胆管细胞的亚细胞分布的InsP 3R亚型的调节。将通过以下具体目标研究这一假设：将在单通道水平上比较InsP 3R的功能和调节。 将在修饰以表达这些受体之一或组合的胆管细胞系中检查每种受体对胆管细胞中Ca 2+信号传导的贡献。这些研究结果将与本机胆管细胞的Ca 2+信号和分泌功能的组织，在孤立的微灌注胆管段确定。这项工作不仅要确定负责Ca 2+信号在胆管细胞的分子机制，但作为一个模型的信号通路的分子组织是如何负责调节小管分泌。

DESCRIPTION (provided by applicant): Bile secretion is one of the principal functions of the liver. In order to maintain bile flow, not only must hepatocytes secrete bile, but this must then be modified and conditioned further by bile duct epithelial cells, or cholangiocytes. Abnormal cholangiocytes function results in cholestasis, which is a cardinal manifestation of liver disease. Cholestatic liver diseases are responsible for 20% of liver transplants in the US, and are the most common cause of liver disease among pediatric transplant patients. In addition, abnormal cholangiocyte function is responsible for the hepatic manifestations of cystic fibrosis, one of the most common inherited diseases. Bile secretion in cholangiocytes is regulated in part by cytosolic Ca2+. In general, cells are regulated both by the pattern of Ca2+ signals over time and by the regions of the cells in which Ca2+ signals occur. However, little is known about temporal or spatial aspects of Ca2+ signaling in cholangiocytes, and nothing is known about how these Ca2+ signals are regulated. Inositol 1,4,5-trisphosphate receptors (InsP3R) mediate Ca2+ signaling in epithelia, and cholangiocytes express all three isoforms of this receptor. The hypothesis of this proposal is that Ca2+ signals in the cholangiocyte are regulated by the subcellular distribution of the InsP3R isoforms. This hypothesis will be investigated through the following specific aims: The function and regulation of the InsP3Rs will be compared at the single channel level. The contribution that each of the receptors plays to Ca2+ signaling in cholangiocytes will be examined in a bile duct cell line modified to express either one or a combination of these receptors. These findings will be related to the organization of Ca2+ signals and secretory function in native cholangiocytes, as determined in isolated microperfused bile duct segments. This work should not only identify the molecular mechanisms responsible for Ca2+ signaling in cholangiocytes, but serve as a model for how the molecular organization of signaling pathways is responsible for regulation of ductular secretion.