Autophagy in Alcoholic Pancreatitis

项目总结： 酗酒是引发人类和啮齿动物胰腺炎的主要危险因素。实验证据 表明饮酒导致溶酶体的不稳定，并增加了酶原的脆性 腺泡细胞。细胞可以通过去除这些受损的溶酶体和酶原来适应和保护自己 自噬和溶酶体生物发生的颗粒(ZGS)。然而，这些保护机制受到了损害。 在长期饮酒之后。大麻素受体2(CB2)激动剂具有强大的抗炎作用 对小鼠实验性胰腺炎的影响及保护作用。然而，潜在的分子机制 CB2激动剂通过哪些途径预防胰腺炎尚不清楚。此外，还缺少的是 具有最佳药代动力学且无主要毒性的CB2激动剂的可获得性 酒精性胰腺炎模型。在我们的初步研究中，我们建立了一种新的酒精性胰腺炎小鼠 使用最近建立的慢性加急性酗酒(Gao-binger)方案的模型。高狂欢诱导 小鼠胰腺的重要发病机制，包括水肿，ZGS增加，表达增加 炎性细胞因子、炎性细胞浸润增加和胰酶释放 (淀粉酶和脂肪酶)进入血液。我们还发现，高糖降低了胰腺组织的表达。 大麻素受体2(CB2)并使溶酶体生物发生的主要调节因子TFEB失活。精挑细选的 CB2激动剂激活小鼠胰腺中的TFEB，增加培养的腺泡细胞的自噬通量。这里 我们提出了一个新的假设，即乙醇代谢导致CB2表达减少并受损 TFEB介导的溶酶体生物发生，自噬不足，脆性ZGS堆积，受损 溶酶体，以及随后的胰腺炎。我们的长期目标是了解 酒精对胰腺腺泡细胞溶酶体生物发生的损害，以确定修复步骤 途径是酒精性胰腺炎的干预点。这项建议的目标是理解 CB2激动剂如何激活TFEB介导的溶酶体生物发生和选择性吞噬酵母菌和溶血素 预防酒精引起的胰腺炎。利用CB2受体方面的免费专业知识 药理学/毒理学(NIAAA内部研究计划)和溶酶体生物发生和建模 体外和体内的酒精性胰腺炎(堪萨斯大学医学中心)我们建议测试一组 新型、高选择性和口服CB2激动剂确保快速转化为临床应用 用于治疗酒精性胰腺炎。我们将确定选择性CB2激动剂如何激活TFEB- 介导的溶酶体生物发生及其改善自噬不足以清除受损的能力 慢性饮酒引起的溶酶体和ZGS。这项建议汇集了研究方面的专业知识 这将解决酒精性胰腺炎的关键研究问题和CB2的潜在干预 激动剂，否则是不可能的。

Project Summary  Alcohol abuse is a major risk factor in initiating pancreatitis in humans and in rodents. Experimental evidence indicates that alcohol consumption leads to the destabilization of lysosomes and increases zymogen fragility in acinar cells. Cells can adapt and protect themselves by removing these damaged lysosomes and zymogen granules (ZGs) by autophagy and lysosomal biogenesis. However, these protective mechanisms are impaired after chronic alcohol consumption. Cannabinoid receptor 2 (CB2) agonists have potent anti-inflammatory effects and protect against experimental pancreatitis in mice. However, the underlying molecular mechanisms by which CB2 agonists protect against pancreatitis are not known. Moreover, what is also missing is the availability of CB2 agonists with optimal pharmacokinetics and absence of major toxicity as tested in relevant alcoholic pancreatitis models. In our preliminary studies, we established a novel alcohol pancreatitis mouse model using the recently established chronic plus acute alcohol binge (Gao-binge) protocol. Gao-binge induces significant pathogenesis in mouse pancreas, including edema, increased ZGs, elevated expression of inflammatory cytokines, increased infiltration of inflammatory cells and release of pancreatic enzymes (amylase and lipase) into the blood. We also demonstrated that Gao-binge decreases expression of pancreatic cannabinoid receptor 2 (CB2) and inactivates TFEB, a master regulator of lysosomal biogenesis. A selective CB2 agonist activates TFEB in mouse pancreas and increases autophagic flux in cultured acinar cells. Here we propose a novel hypothesis that ethanol metabolism leads to decreased CB2 expression and impaired TFEB-mediated lysosomal biogenesis, insufficient autophagy, accumulation of fragile ZGs, damaged lysosomes, and subsequent pancreatitis. Our long-term goal is to understand the molecular mechanisms for alcohol impairment of lysosomal biogenesis in pancreatic acinar cells, in order to identify steps in the repair pathway that are points for intervention in alcoholic pancreatitis. The objective of this proposal is to understand how CB2 agonists activate TFEB-mediated lysosomal biogenesis and selective zymophagy and lysophagy to protect against alcohol-induced pancreatitis. Using complimentary expertise in CB2 receptor pharmacology/toxicology (NIAAA intramural research program) and modeling of lysosomal biogenesis and alcoholic pancreatitis in vitro and in vivo (University of Kansas Medical Center) we propose to test a set of novel, highly selective and orally available CB2 agonists assuring quick translation toward clinical application for the treatment of alcoholic pancreatitis. We will determine how the selective CB2 agonists activate TFEB- mediated lysosomal biogenesis and their ability to improve the insufficient autophagy to remove damaged lysosomes and ZGs induced by chronic alcohol consumption. This proposal brings together research expertise that will address key research questions on alcoholic pancreatitis and the potential intervention by CB2 agonists, which would not otherwise be possible.