Ryanodine Receptor in Pancreatitis

项目摘要/摘要 急性胰腺炎是一种常见的、危及生命的胰腺疾病。腺泡细胞钙信号异常 在引发这种疾病的过程中起着至关重要的作用。我们之前已经证明，在脑内钙信号异常升高 基底区与病理性腺泡内蛋白水解酶激活有关，这是一种早期和危急的事件 胰腺炎的发展。这种钙信号是由内质网(ER)钙通道介导的， 兰尼定受体(RyR)。在这项建议中，我们研究了调节这种病理性RyR钙离子的机制 在腺泡细胞中释放。我们已经证明，腺泡细胞中cAMP的增加会导致RyR磷酸化， RyR Ca~(2+)释放，并增强蛋白酶活性1，2。在初步工作中，我们证明了酒精，a 胰腺炎的主要原因，触发cAMP介导的RyR磷酸化以及RyR钙离子释放和 增强了蛋白酶的活性。因此，我们假设腺泡内蛋白水解酶的激活和 胰腺炎，尤其是酒精引起的胰腺炎，是由腺泡细胞钙离子释放引发的。 来自病理激活的、磷酸化的RyRs。在这项提议中，使用了新的基因组合 和药理工具，在酒精存在或不存在的情况下，我们将追求以下具体目标： (1)检测RyR磷酸化是否导致离体腺泡RyR钙离子释放 (2)研究RyR磷酸化和RyR Ca~(2+)释放是否易引起细胞内蛋白水解酶的激活 腺泡 (3)确定RyR磷酸化和RyR Ca~(2+)释放是否易于蛋白水解酶的激活和释放 活体胰腺炎。被认为介导PKA依赖的RyR钙离子的RyR位点的磷酸化作用 释放将直接测试使用先前产生的转基因小鼠，这些小鼠携带模拟磷或 RyR PKA磷酸化位点的抗磷突变。 预计这些关于RyR在病理性蛋白水解酶激活和胰腺炎中的作用的研究将 (1)促进对胰腺炎中异常的钙信号的理解，(2)提供一种新的联系 酒精和RyR钙离子释放可能对酒精的作用机制有更广泛的影响 多个其他器官系统的并发症，以及(3)建议针对RyR的治疗策略。 胰腺。

Project Summary/Abstract Acute pancreatitis is a common, life-threatening disorder of the pancreas. Abnormal acinar cell Ca2+ signals play a crucial role in initiating this disease. We have previously shown that abnormally elevated Ca2+ signals in the basal region are associated with pathologic intra-acinar protease activation, an early and critical event in the development of pancreatitis. This Ca2+ signal is mediated by an endoplasmic reticulum (ER) Ca2+ channel, the ryanodine receptor (RyR). In this proposal, we examine mechanisms that regulate this pathologic RyR Ca2+ release in the acinar cell. We have shown that increasing cAMP in acinar cells causes RyR phosphorylation, RyR Ca2+ release, and enhanced protease activation1, 2. In preliminary work, we demonstrate that alcohol, a leading cause of pancreatitis, triggers cAMP-mediated RyR phosphorylation as well as RyR Ca2+ release and enhanced protease activation. Therefore, we hypothesize that intra-acinar protease activation and pancreatitis, particularly that induced by alcohol exposure, are triggered by release of acinar cell Ca2+ from pathologically activated, phosphorylated RyRs. In this proposal, using a combination of novel genetic and pharmacologic tools, we will, in the presence or absence of alcohol, pursue the following Specific Aims: (1) Examine whether RyR phosphorylation causes RyR Ca2+ release in isolated acini (2) Study whether RyR phosphorylation and RyR Ca2+ release predispose to protease activation in isolated acini (3) Determine whether RyR phosphorylation and RyR Ca2+ release predispose to protease activation and pancreatitis in vivo. The effects of phosphorylation on RyR sites thought to mediate PKA-dependent RyR Ca2+ release will be directly tested using previously generated transgenic mice that harbor phospho-mimetic or phospho-resistant mutations in the RyR PKA phosphorylation site. It is anticipated that these studies on the role of the RyR in pathologic protease activation and pancreatitis will (1) lead to improved understanding of aberrant Ca2+ signaling in pancreatitis, (2) provide a novel link between alcohol and RyR Ca2+ release that may have broader implications for mechanisms contributing to alcohol's complications in multiple other organ systems, and (3) suggest treatment strategies that target the RyR in the pancreas.