Ryanodine Receptor in Pancreatitis

项目总结/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关键作用。我们以前已经表明，异常升高的Ca 2+信号， 基底区与病理性腺泡内蛋白酶激活有关，这是 胰腺炎的发展。这种Ca 2+信号由内质网（ER）Ca 2+通道介导， Ryanodine receptor（RyR）在这个提议中，我们研究了调节这种病理性RyR Ca 2+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，并增强蛋白酶激活1，2。在初步工作中，我们证明，酒精， 胰腺炎的主要原因，触发cAMP介导的RyR磷酸化以及RyR Ca 2+释放， 增强的蛋白酶活化。因此，我们假设腺泡内蛋白酶激活和 胰腺炎，特别是酒精暴露引起的胰腺炎，是由腺泡细胞Ca 2+释放引发的 病理性激活的磷酸化RyR。在这项提议中，使用一种新的基因组合， 和药理学工具，我们将在有或无酒精的情况下，追求以下具体目标： (1)检查RyR磷酸化是否导致分离腺泡中RyR Ca 2+释放 (2)研究RyR磷酸化和RyR Ca 2+释放是否使分离的细胞中的蛋白酶活化倾向于蛋白酶活化。 腺泡 (3)确定RyR磷酸化和RyR Ca 2+释放是否倾向于蛋白酶活化， 体内胰腺炎。磷酸化对被认为介导PKA依赖性RyR Ca 2+的RyR位点的影响 释放将直接使用先前产生的携带磷酸模拟物或 RyR PKA磷酸化位点的磷酸化抗性突变。 预期这些关于RyR在病理性蛋白酶激活和胰腺炎中的作用的研究将 (1)导致对胰腺炎中异常Ca 2+信号传导理解的提高，（2）提供了 酒精和RyR Ca 2+释放可能对酒精的机制有更广泛的影响， 在多个其他器官系统的并发症，和（3）建议治疗策略，靶向RyR在 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.