Regulation of PKC alpha in Alcoholic Myocytes

酒精性肌细胞中 PKC α 的调节

• 批准号: 6739095
• 负责人: MICHELE L SOLEM
• 金额: $ 15.7万
• 依托单位: THOMAS JEFFERSON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-05-01 至 2006-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6739095
• 关键词: RNase protection assay; alcoholic beverage consumption; alcoholism /alcohol abuse; biological signal transduction; calcium channel; cardiac myocytes; disease /disorder model; enzyme activity; enzyme mechanism; ethanol; gene expression; immunofluorescence technique; immunoprecipitation; insulinlike growth factor; laboratory rat; membrane activity; microarray technology; northern blottings; phosphatidylinositol 3 kinase; phospholipase C; protein biosynthesis; protein kinase C; protein protein interaction; protein tyrosine kinase; western blottings

DESCRIPTION (provided by applicant): This is a revised grant application to study PKC alpha in the alcoholic heart. IGF1 is an essential cytokine that is required for normal development of the heart, and it may play a critical physiological role in the normal adult heart as well as influence various pathological conditions. Our laboratory finds that IGF1 activates PKC alpha in adult rat cardiomyocytes, and this results in PKC alpha-dependent activation of MAP kinase, protein synthesis and gene expression. However, in cardiomyocytes from chronic alcohol-exposed rats, there is reduced activation of PKC alpha by IGF1, loss of IGF1-dependent protein synthesis and a significant increase in the unstimulated (basal) state of the IGF1 receptor. Since chronic alcohol exerts such a profound effect on the IGF1 signaling pathway, an examination of how PKC alpha is involved in these changes may uncover novel cellular mechanisms associated with chronic alcohol-induced damage to the heart. The first aim of this research proposal (1) will be to determine how chronic alcohol alters IGF1-induced PKC alpha activation. What are the changes brought about by chronic alcohol exposure that suppress activation of PKC alpha (e.g. alterations in the IGF1 receptor activation, including the possible involvement of PKC, and the downstream influence on the activities of phospholipase C gamma, PI3 kinase and intracellular tyrosine kinases. The second aim of this proposal (2) will be to study how alterations in IGF1 signaling affect PKC alpha-dependent regulation of protein synthesis and gene expression. Results from cDNA Expression Arrays indicate that the expression of several essential cardiac-specific genes which are involved in excitation-contraction coupling and cell metabolism (e.g. GIRK4, carnitine palmoyltyl transferase, HSP60, G2/M-specific cyclin G), are regulated by PKC alpha-dependent activation by IGF1. This part of the grant proposal will be done in collaboration with the Daniel Baugh Institute at Thomas Jefferson University, using its BioRobotics MicroGridTAS system for microarray analysis of gene expression. The last aim (3) of the proposal will examine how long term alcohol consumption alters the IGF1 signal transduction pathway using the laboratory's model, the chronic alcohol-fed rat fed a diet containing 37% alcohol for various time points of alcohol exposure (e.g. 2 weeks, 2 months, 4 months, 6 months, 10 months). A comparison of the cardiac Ca2+ channel activity, PKC activation and protein synthesis will be made. The objective is to better define the sequence of events that are associated with alcohol-impaired alterations in IGF1 signaling.

描述（由申请人提供）： 这是一项修订后的拨款申请，旨在研究酒精性心脏中的 PKC α。 IGF1是心脏正常发育所需的重要细胞因子，它可能在正常成人心脏中发挥重要的生理作用并影响各种病理状况。我们的实验室发现 IGF1 激活成年大鼠心肌细胞中的 PKC α，从而导致 MAP 激酶、蛋白质合成和基因表达的 PKC α 依赖性激活。然而，在长期暴露于酒精的大鼠的心肌细胞中，IGF1 对 PKC α 的激活减少，IGF1 依赖性蛋白质合成丧失，并且 IGF1 受体的未刺激（基础）状态显着增加。由于慢性酒精对 IGF1 信号通路产生如此深远的影响，因此检查 PKC α 如何参与这些变化可能会揭示与慢性酒精引起的心脏损伤相关的新细胞机制。本研究提案 (1) 的首要目标是确定长期饮酒如何改变 IGF1 诱导的 PKC α 激活。慢性酒精暴露会带来哪些抑制 PKC α 激活的变化（例如 IGF1 受体激活的改变，包括 PKC 的可能参与，以及下游影响） 磷脂酶 C γ、PI3 激酶和细胞内酪氨酸激酶的活性。该提案 (2) 的第二个目标是研究 IGF1 信号传导的改变如何影响 PKC α 依赖性蛋白质合成和基因表达的调节。 cDNA 表达阵列的结果表明，几个参与兴奋-收缩耦合和细胞代谢的重要心脏特异性基因（例如 GIRK4、肉碱棕榈酰基转移酶、HSP60、G2/M 特异性细胞周期蛋白 G）的表达受 PKC 调节IGF1 的 α 依赖性激活。这部分拨款提案将与托马斯杰斐逊大学 Daniel Baugh 研究所合作完成，使用其 BioRobotics MicroGridTAS 系统进行基因表达的微阵列分析。该提案的最后一个目标 (3) 将使用实验室模型来研究长期饮酒如何改变 IGF1 信号转导途径，慢性酒精喂养的老鼠在不同的酒精暴露时间点（例如 2周、2 个月、4 个月、6 个月、10 个月）。将比较心脏 Ca2+ 通道活性、PKC 激活和蛋白质合成。目的是更好地定义与酒精损伤的 IGF1 信号改变相关的事件序列。