Ca2+ waves in hepatocytes: Mechanisms and effects

项目总结 胆汁分泌是肝脏的主要功能之一。胆汁淤积，或胆汁流动障碍，是一种主要的 肝病的表现。胆汁淤积性肝病是一组重要的疾病，它们统称为 是儿科肝脏移植最常见的适应症，占所有肝脏的十分之一 移植。这个项目研究了上皮生物学中的一种直接相关的基本生物学机制。 用于调节健康和疾病中肝细胞的分泌。肝细胞内钙信号的形成 主要通过肌醇1，4，5-三磷酸(InsP3)受体(InsP3R)的钙释放，InsP3R是一种InsP3- 内质网(ER)中的门控钙通道。以II型InsP3R(InsP3R-2)为主 肝细胞中的异构体，占InsP3Rs总库的80%，最集中在一种特殊的 内质网小管膜下方的区域。肝细胞中的钙信号一般始于钙。 起源于该区域的波和肝细胞中InsP3R-2的顶端池对于调节 钙波和小管分泌物。相比之下，I型InsP3R(InsP3R-1)构成了 肝细胞中剩余20%的InsP3R池，优先定位于不同的亚细胞区域 不会影响分泌。这个项目的假设是，调节表达和表达的因素 InsP3R-2的亚细胞分布也调节肝细胞的分泌，因此胆汁淤积是通过这些调节机制实现的。 对InsP3R-2的影响。特别是，我们将测试淤胆性肝病是否在一定程度上是由于受损。 InsP3R-2的表达和/或管周靶向，进而损害心肌细胞内的极化钙波 胞浆和下游事件，包括胆汁酸转运体BSEP和有机阴离子的调节 运输机mrp2。这一假设将通过以下具体目标进行检验：(1)我们将确定 调节肝细胞InsP3R-2表达的遗传和表观遗传因素。(2)我们会决定 将InsP3R-2靶向内质网-小管膜界面的分子基础和细胞效应。(3)我们 将确定InsP3R-2的表达和/或肝细胞管周靶向是否以及如何受损 在小管胆汁淤积症期间。我们还将确定正在开发的新疗法的疗效 治疗胆汁淤积症的作用部分是通过调节InsP3R-2来实现的。总的来说，这些研究将 在我们对建立信号微域的方式的理解方面取得了新的突破 肝细胞，并有可能建立新的范例，将这些观察转化为 了解一系列临床上有意义的人类肝病的机制基础。

PROJECT SUMMARY Bile secretion is one of the principal functions of the liver. Cholestasis, or impaired bile flow, is a cardinal manifestation of liver disease. Cholestatic liver diseases are an important group of disorders, which collectively represent the most common indication for pediatric liver transplant and account for one in ten of all liver transplants. This project investigates a basic biological mechanism in epithelial biology that is directly relevant for the regulation of secretion in hepatocytes in health and disease. Calcium signals in hepatocytes are formed largely by calcium release from the inositol 1,4,5-trisphosphate (InsP3) receptor (InsP3R), which is an InsP3- gated calcium channel in the endoplasmic reticulum (ER). The type II InsP3R (InsP3R-2) is the predominant isoform in hepatocytes, constituting 80% of the total pool of InsP3Rs, and is most concentrated in a specialized region of the ER beneath the canalicular membrane. Calcium signals in hepatocytes generally begin as calcium waves that originate in this region, and the apical pool of InsP3R-2 in hepatocytes is important for mediating calcium waves and canalicular secretion. In contrast, the type I InsP3R (InsP3R-1), which constitutes the remaining 20% of the InsP3R pool in hepatocytes, preferentially localizes to a different subcellular region and does not affect secretion. The hypothesis of this project is that the factors that regulate the expression and subcellular distribution of InsP3R-2 also regulate hepatocyte secretion, so that cholestasis is mediated by these effects on InsP3R-2. In particular, we will test whether cholestatic liver diseases are due in part to impaired expression and/or peri-canalicular targeting of InsP3R-2, which in turn impairs polarized calcium waves in the cytosol and downstream events including regulation of the bile acid transporter Bsep and the organic anion transporter Mrp2. This hypothesis will be tested through the following specific aims: (1) We will determine the genetic and epigenetic factors that regulate expression of InsP3R-2 in hepatocytes. (2) We will determine the molecular basis for, and cellular effects of, targeting InsP3R-2 to the ER-canalicular membrane interface. (3) We will determine whether and how InsP3R-2 expression and/or pericanalicular targeting is impaired in hepatocytes during canalicular cholestasis. We also will determine whether the efficacy of new therapies being developed for treatment of cholestatis disorders is mediated in part by modulating InsP3R-2. Collectively, these studies will break new ground in our understanding of the ways in which signaling microdomains are established in hepatocytes, and have the potential to establish new paradigms for translating these observations into understanding the mechanistic basis for a range of clinically significant human liver diseases.