AMPKa agonist in attenuating CPT1A inhibition and alcoholic chronic pancreatitis

摘要 慢性酒精摄入/滥用每年给美国经济造成约2500亿美元的损失，约10万人死亡， 慢性胰腺炎（CP）的主要原因。酒精性慢性胰腺炎是一种严重的纤维炎症性疾病， 导致外分泌功能不全的疾病以及糖尿病和胰腺癌等严重并发症。 一些有长期饮酒史的患者甚至在疾病临床表现出来之前就已经死亡。 外分泌胰腺占胰腺的约85%，是酒精中毒的主要目标。虽然 饮酒量和持续时间是ACP发病的关键因素，其机制和 ACP的代谢基础是复杂的，仍然没有很好地理解。此外，没有有效的治疗ACP的方法， 提供给诊所。人类长期饮酒（乙醇，EtOH）会损害肝脏酒精 脱氢酶（ADH）并增加脂肪浸润和脂肪酸乙酯（FAEE， EtOH的非氧化代谢物）。最近我们发现，六个月大的肝脏ADH 1 缺乏（ADH-）与肝ADH正常（ADH+）鹿小鼠喂食慢性乙醇3个月， EtOH的高身体负荷，胰腺脂质表型失调，包括FAEE水平升高，脂毒性 炎症并模拟ACP的几种纤维免疫学特征。另外，我们发现 慢性乙醇喂养ADH- vs. ADH+鹿小鼠和EtOH处理的人胰腺腺泡细胞（hPAC）中。类似的结果也观察到， 尸检诊断为ACP的供体胰腺组织。此外，EtOH诱导的AMPKα AMPKα激动剂可减弱CPT 1A的失活和表达减少。因此，我们假设 AMPKα激动剂减弱慢性EtOH诱导的胰腺CPT 1A抑制， 胰腺脂质表型失调沿着FAEE形成和相关脂毒性 炎症和纤维化。在目标1中，我们将确定慢性EtOH是否 降低胰腺CPT 1A的表达，导致胰腺损伤，脂质表型失调，增加 在我们的ADH-鹿小鼠模型中FAEE的形成和线粒体应激。在目标2中，我们将确定 EtOH暴露可抑制CPT 1A并导致脂质表型失调和FAEE形成，细胞毒性 和线粒体应激，以确定外分泌胰腺腺泡细胞是 EtOH诱导毒性。在这两个目标中，我们将通过衰减EtOH来测量上述参数。 诱导AMPKα激动剂抑制CPT 1A和AMPKα失活。我们的项目是创新的，因为我们 将确定ADH中涉及胰腺脂质表型失调的关键脂质代谢途径/网络- 鹿小鼠酒精性胰腺炎模型使用先进的脂质组学方法。此外，调查结果显示， 预计该项目将建立胰腺CPT 1A和AMPKα信号传导与脂质失调之间的联系 表型的发病机制，并开发其治疗。

Abstract Chronic alcohol intake/abuse costing ~250 billion dollars to the US economy and ~100,000 deaths annually is the major cause of chronic pancreatitis (CP). Alcoholic chronic pancreatitis (ACP) is a serious fibro-inflammatory disorder resulting in exocrine insufficiency and such severe co-morbidities as diabetes and pancreatic cancer. Several patients with a history of chronic alcohol use die even before the disease becomes clinically manifested. The exocrine pancreas, which constitutes ~85% of the gland, is a prime target of alcohol toxicity. Although the amount and duration of alcohol consumed are key factors driving pathogenesis of ACP, the mechanism and metabolic basis of ACP are complex and still not well understood. Besides, no effective therapeutics for ACP is available for the clinics. Chronic alcohol (ethanol, EtOH) consumption in humans impairs hepatic alcohol dehydrogenase (ADH) and increases fatty infiltration and formation of fatty acid ethyl esters (FAEEs, nonoxidative metabolites of EtOH) in the pancreas. Recently we found that the six-months-old hepatic ADH1 deficient (ADH-) vs. hepatic ADH normal (ADH+) deer mice fed chronic EtOH for 3 months results in exceedingly high body burden of EtOH, dysregulated pancreatic lipid phenotype including elevated levels of FAEEs, lipotoxic inflammation and mimic several fibro-immunological characteristics of ACP. Besides, we found a decreased expression of carnitine palmitoyl transferase (CPT)1A in the pancreatic tissue of chronic EtOH fed ADH- vs. ADH+ deer mice and in EtOH treated human pancreatic acinar cells (hPACs). Similar findings were observed in pancreatic tissue of donors with a premortem diagnosis of ACP. Furthermore, EtOH-induced AMPKα deactivation and reduced expression of CPT1A was attenuated by AMPKα agonist. Therefore, we hypothesize that attenuation of chronic EtOH-induced inhibition of pancreatic CPT1A by AMPKα agonist prevents dysregulated pancreatic lipid phenotype along with formation of FAEEs and associated lipotoxic inflammation, and fibrosis in alcoholic chronic pancreatitis. In aim 1 we will determine if chronic EtOH reduces the expression of pancreatic CPT1A, causes pancreatic injury, dysregulates lipid phenotype, increases the formation of FAEEs and mitochondrial stress in our ADH- deer mouse model. In aim 2 we will determine if EtOH exposure inhibits CPT1A and causes dysregulated lipid phenotype and formation of FAEEs, cytotoxicity and mitochondrial stress in hPACs, in vitro, to establish that exocrine pancreatic acinar cells are the target of EtOH induced toxicity. In both aims, we will measure above mentioned parameters by attenuating the EtOH induced inhibition of CPT1A and AMPKα inactivation by AMPKα agonist. Our project is innovative because we will identify key lipid metabolic pathways/networks involved in dysregulated pancreatic lipid phenotype in ADH- deer mouse model of alcoholic pancreatitis using cutting edge lipidomic approach. Besides, findings of this project are expected to establish a link between pancreatic CPT1A and AMPKα signaling and dysregulated lipid phenotype in pathogenesis of ACP and develop its therapeutics.