Effects of Ethanol on TNF Cytotoxicity

乙醇对 TNF 细胞毒性的影响

• 批准号: 6769874
• 负责人: JOHN G PASTORINO
• 金额: $ 18.32万
• 依托单位: THOMAS JEFFERSON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-07-01 至 2007-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6769874
• 关键词: apoptosis; biological signal transduction; cytotoxicity; enzyme activity; ethanol; laboratory rat; liver cells; membrane permeability; mitochondrial membrane; mitogen activated protein kinase; pathologic process; phosphatidylinositol 3 kinase; phosphorylation; tissue /cell culture; tumor necrosis factor alpha

DESCRIPTION (provided by applicant): Ethanol exposure promotes the development of alcoholic liver disease (ALD) but the mechanisms underlying ethanol-induced hepatotoxicity remain poorly understood. An important cellular site manifesting damage brought on by ethanol is the mitochondria. Proinflammatory cytokines such as tumor necrosis factor alpha (TNFalpha or TNF) have been linked to many of the associated damage and repair processes seen in ALD. TNFalpha prompts the opening of the mitochondrial permeability transition pore (MPT). The MPT is the regulatable opening of a large, nonspecific pore across the outer and inner mitochondrial membrane. Development of the MPT has been implicated in both necrotic and some forms of apoptotic cell death. Inhibition of the MPT prevents many of the typical manifestations seen during apoptosis. Liver mitochondria isolated from chronically ethanol fed rats are more susceptible to MPT pore opening induced by various triggers than mitochondria isolated from control animals. In preliminary results we demonstrate an enhancement of TNFalpha cytotoxicity in hepatocytes isolated from chronically ethanol-fed rats compared to their pair-fed controls. Similarly, a 48h exposure of HepG2 cells to ethanol enhanced susceptibility to TNFalpha cytotoxicity. The data suggest that ethanol enhances TNFalpha cytotoxicity, at least in part, by promoting the MPT. The general goals of this project are to investigate the mechanisms by which ethanol potentiates TNFalpha induced cytotoxicity. Our working hypothesis is that ethanol inhibits signaling pathways that protect against TNFalpha cytotoxicity and activates pathways that promote the mitochondrial permeability transition. This theory is supported by our preliminary observations that ethanol inhibits P13-kinase dependent BAD phosphorylation induced by TNFalpha and promotes cell death in a p38 MAPK dependent and caspase 8 independent manner. In addition, ethanol exposure sensitizes mitochondria to the MPT and causes an increase in the levels of the peripheral benzodiazepine receptor, a putative component of the MPT pore. Our SPECIFIC AIMS are to #1) further define the role of ethanol in the alteration of P13-kinase activity and its influence on TNFalpha cytotoxicity and #2) delineate the role and mechanism(s) of activation of p38 MAPK in ethanol plus TNFalpha cytotoxicity; #3) Define the mechanism(s) by which TNF plus ethanol cause the MPT.

描述（由申请人提供）：乙醇暴露促进发育 酒精性肝病（ALD）的发病机制，但乙醇诱发的机制 肝毒性仍知之甚少。一个重要的细胞部位表现 乙醇带来的损害是线粒体。促炎细胞因子 例如肿瘤坏死因子 α（TNFα 或 TNF）与许多疾病有关 ALD 中出现的相关损伤和修复过程。 TNFα 提示 线粒体通透性转换孔（MPT）的开放。 MPT 是 穿过外部和内部的大的非特异性孔的可调节开口 线粒体膜。 MPT 的发展涉及到以下两个方面： 坏死和某些形式的细胞凋亡。 MPT 的抑制可防止 许多细胞凋亡过程中出现的典型表现。肝脏线粒体 从长期喂食乙醇的大鼠中分离出的物质更容易受到 MPT 孔的影响 与从对照中分离的线粒体相比，各种触发因素诱导的开放 动物。在初步结果中，我们证明了 TNFα 的增强 比较从长期喂食乙醇的大鼠中分离出的肝细胞的细胞毒性 到他们的配对控制。同样，HepG2 细胞暴露于乙醇 48 小时 对 TNFα 细胞毒性的敏感性增强。数据表明，乙醇 至少部分通过促进 MPT 来增强 TNFα 细胞毒性。这 该项目的总体目标是研究以下机制： 乙醇增强 TNFα 诱导的细胞毒性。我们的工作假设是 乙醇会抑制抵抗 TNFα 的信号通路 细胞毒性并激活促进线粒体通透性的途径 过渡。这一理论得到了我们初步观察的支持： 乙醇抑制 TNFα 诱导的 P13 激酶依赖性 BAD 磷酸化 并促进 p38 MAPK 依赖性和 caspase 8 依赖性细胞死亡 方式。此外，乙醇暴露会使线粒体对 MPT 敏感， 导致外周苯二氮卓受体水平增加， MPT 孔隙的假定成分。我们的具体目标是 #1) 进一步定义 乙醇在改变P13激酶活性中的作用及其影响 TNFα 细胞毒性和 #2) 描述了 TNFα 细胞毒性的作用和机制 乙醇中 p38 MAPK 的激活加上 TNFα 细胞毒性； #3) 定义 TNF 加乙醇引起 MPT 的机制。