Lipid Metabolism in the Aging Immune System

描述(申请人提供)：血脂在老年人的许多主要疾病(动脉粥样硬化、癌症、关节炎)以及衰老本身中起着基本的作用。具体地说，磷脂酸(PA)在细胞功能中发挥着核心作用，它通过合成磷脂来改变膜结构，并作为一种强大的细胞内信号分子。近年来，细胞内脂质结合蛋白--酰基辅酶A结合蛋白(ACBP)已成为PA合成和细胞内信号转导的重要调节因子。老年人T细胞中磷脂的变化与免疫功能降低有关。问题：衰老导致T细胞磷脂质量和脂肪酸组成发生显著变化，然而，导致这些变化的生化和分子机制(S)尚不清楚。目的：1)确定3-磷酸甘油酰基转移酶(GPAT)生物合成减少是否至少部分导致T细胞增殖减少；2)确定ACBP在老年T细胞功能障碍中PA生物合成中的作用；3)评估ACBP在老年T细胞亚群功能障碍中的作用。方法：青年大鼠和老年大鼠随机喂饲饲料。由于内质网(ER)和线粒体是PA从头合成的主要部位，因此将从脾T细胞中分离内质网(微粒体)和线粒体。这项工作将集中在甘油-3-磷酸的酰化为溶血磷脂酸，然后分别在(GPAT)和溶血磷脂酸酰基转移酶的作用下生成PA。将设计实验来确定衰老对GPAT的影响本质上是转录前(基因表达)还是转录后(磷酸化，蛋白质水平)。这一点意义重大，因为GPAT是PA生物合成中的限速酶。ACBP在介导T细胞功能减退中的作用将通过检测过表达的ACBP是否改变T细胞的分泌以及衰老对ACBP水平的影响来评估，分别采用蛋白质和基因表达的免疫印迹分析和RT-PCR。ACBP对CD4+、CD8+T细胞亚群和记忆/幼稚T细胞功能随年龄的影响将通过将ACBP导入那些随年龄改变ACBP表达的老年T细胞群体并通过流式细胞仪检测细胞因子的产生来确定。结果：这些实验将深入了解PA合成和ACBP表达或功能的变化是否是年龄相关性T细胞增殖丧失的重要机制解释。其次，将深入了解ACBP是否在特定T细胞亚群的年龄依赖性功能障碍中发挥作用。益处：确定衰老对ACBP和PA合成的影响可能最终导致旨在维持最佳免疫功能的饮食和/或药物策略的开发，从而提高老年人的生活质量。

DESCRIPTION (provided by applicant): Lipids play a fundamental role in many of the major diseases (atherosclerosis, cancer, arthritis) in the elderly as well as in aging itself. Specifically, phosphatidic acid (PA) plays a central role in cell function by modifying membrane structure via synthesis into phospholipids and serving as a potent intracellular signaling molecule. Recently, the intracellular lipid binding protein, acyl-CoA binding protein (ACBP), has emerged as an important regulator of PA synthesis and intracellular signaling. Phospholipid alterations in T-cells are associated with reduced immune function in the elderly. Problem: Aging results in significant changes in phospholipid mass and fatty acid composition in T-cells, however, the biochemical and molecular mechanism(s) causing these alterations is not known. Purpose: 1) Determine if reduced PA biosynthesis via glycerol-3-phosphate acyltransferase (GPAT) accounts, at least in part, for reduced T-cell proliferation; 2) Determine the role of ACBP in PA biosynthesis in aged T-cell dysfunction; 3) Assess the role of ACBP in aged T-cell subset dysfunction. Methods: Young and old rats fed ad libitum diets will bell used. Endoplasmic reticulum (ER) (microsomes) and mitochondria will be isolated from the splenic T-cells because the ER and mitochondria are the major sites of de novo PA biosynthesis. The work will focus on the acylation of glycerol-3-phosphate to lysophosphatidic acid then to PA by (GPAT) and lysophosphatidic acid acyltransfcrase respectively. Experiments will be designed to determine if the aging effects on GPAT are pre- (gene expression) or posttranscriptional (phosphorylation, protein levels) in nature. This is significant because GPAT is the rate-limiting enzyme in PA biosynthesis. The influence of ACBP in mediating reduced T-cell function with age will be assessed by examining whether over-expressed ACBP alters T-cell oroliferation and what impact aging has on ACBP levels by analyzing protein and gene expression by western immunoblotting_and RT-PCR, respectively. ACBP's influence on CD4+ and CD8+ T-cell sub sets and memory/naive-T-cell function with age will be determined by transfecting ACBP into those aged T-cell populations which exhibit altered ACBP expression with age and examining cytokine production by flow cytometry. Outcome: The experiments will give insight into whether alterations in PA synthesis and ACBP expression or function are an important mechanistic explanation for the age-dependent loss of T-cell proliferation. Secondly, insight will be gained into whether ACBP plays a role in the age-dependent dysfunction of specific T-cell subsets. Benefit: Characterizing the influence of aging on ACBP and PA synthesis may ultimately lead to the development of dietary and/or pharmacologic strategies aimed at maintaining optimal immune function hence improving the quality of life in the elderly.