MOLECULAR BASIS OF HIV LIPODYSTROPHY: ROLE OF VPR

描述(改编自申请人的摘要) 本RFA的2个R01项目的统一目标是指定一个关键字 HIV相关性脂营养不良综合征(HLS)的分子机制 以及随之而来的代谢紊乱导致其临床 表现形式。这些表现提示糖皮质激素增加。 脂肪、肌肉和肝脏的敏感性。最近的研究表明，一种HIV-1病毒 VPR蛋白增强配体介导的糖皮质激素激活 受体(GR)，并拮抗PPAR-γ介导的基因转录。 初步研究还表明，HLS患者有血脂和血糖 动力学符合VPR的这些分子效应。 基础科学R01项目的假设是a)过度表达 VPR在小鼠体内引起以GR活化为特征的代谢变化； B)VPR对脂肪生成和脂肪生成的影响在中央和中央不同。 外周脂肪细胞；c)vpr通过与 GR和PPAR-γ的转录复合体，导致激活 GR调节基因和PPAR-γ调节基因的抑制。这个 具体目标是：a)测量身体成分和脂肪、蛋白质和 利用稳定同位素/质谱仪研究转基因动物的葡萄糖代谢 小鼠在饮食控制和蛋白水解酶作用下过度表达VPR 抑制剂的使用；b)脂肪生成和脂肪生成的功能分析 从这些小鼠的中枢和外周去除的脂肪细胞；c)分子 VPR介导的GR调节机制及其作用的剖析 和PPAR-γ在人前脂肪细胞系和原代培养中的表达 来自腹部和外周脂肪库的前脂肪细胞。 临床科学R01的假设是：a)患者出现 HLS逐渐表现出持续的GR激活的代谢效应 脂肪、肌肉和肝脏，即全身脂肪分解增多，增多 肝脏脂肪生成增加，腹部脂肪中甘油三酯储存增加 蛋白质周转率和内源性葡萄糖产量在空腹和 喂食；b)这些变化是次要的，因为对 VPR作用所致的糖皮质激素。具体目标包括 对新诊断的艾滋病毒感染患者进行纵向、密集的GCRC研究 和匹配的正常受试者，以测量：a)全身脂肪动力学 (脂肪分解、脂肪生成、再酯化、极低密度脂蛋白合成、甘油三酯 利用)、局部脂肪分解、蛋白质周转和内源性葡萄糖 生产，同时喂食和禁食，使用稳定同位素/质谱分析 B)糖皮质激素对蛋白分解和脂解的敏感性；c)VPR 血浆、腹部脂肪和大腿脂肪细胞外液中的浓度； D)详细的HLS的身体成分和生化参数。 这些项目将由一个具有以下经验的协调团队执行 代谢方案和稳定同位素技术，艾滋病毒临床专家， 以及GR和VPR的分子生物学专家。他们将详细介绍一项 这种新的脂肪营养不良综合征的分子途径可能先于 使用有效的抗逆转录病毒疗法，但在临床上是显而易见的 在治疗期间由于营养摄入量增加而导致的，并转化为 临床科学：VPR介导的代谢失调机制.

DESCRIPTION (Adapted from the applicant's abstract) The unified objective of the 2 R01 projects in this RFA is to specify a key molecular mechanisms underlying HIV-associated lipodystrophy syndrome (HLS), and the consequent metabolic derangements that lead to its clinical manifestations. These manifestations suggest increased glucocorticoid sensitivity in fat, muscle and liver. Recent work shows that an HIV-1 viral protein, Vpr, potentiates ligand-mediated activation of the glucocorticoid receptor (GR), and antagonized PPAR-gamma mediated gene transcription. Preliminary studies also show that HLS patients have lipid and glucose kinetics consistent with these molecular effects of Vpr. The hypotheses for the Basic Science R01 project are that a) over expression of Vpr in mice leads to metabolic changes characteristic of GR activation; b) Vpr affects adipogenesis and lipogenesis differentially in central vs. peripheral adipocytes; c) Vpr exerts these effects by direct interactionwith the transcriptional complexes of the GR and PPAR-gamma, leading to activation of GR-regulated genes and inhibition of PPAR-gamma regulated genes. The Specific Aims are: a) measurements of body composition and lipid, protein and glucose metabolism, using stable isotopes/mass spectrometry, in transgenic mice over expressing Vpr, following dietary manipulations and protease inhibitor administration; b) functional assays of adipogenesis and lipogenesis in central vs. peripheral adipocytes removed from these mice; c) molecular dissection of the mechanism and effects of Vpr-mediated regulation of the GR and PPAR-gamma in preadipocyte cell lines and primary cultures of human preadipocytes derived from abdominal and peripheral fat depots. The hypotheses for the clinical Science R01 are that a) patients who develop HLS progressively manifest the metabolic effects of persistent GR activation fat, muscle, and liver, namely, increased whole body lipolysis, increased hepatic lipogenesis, enhanced triglyceride storage in abdominal fat, increased protein turnover, and elevated endogenous glucose production while fasting and feeding; b) these changes are secondary to increased sensitivity to glucocorticoids due to the actions of Vpr. The Specific Aims involve longitudinal, intensive GCRC studies on newly diagnosed HIV-infected patients and matched normal subjects, to measure; a) whole body lipid kinetics (lipolysis, lipogenesis, reesterification, VLDL synthesis, triglyceride utilization), regional lipolysis, protein turnover, and endogenous glucose production, while feeding and fasting, using stable isotopes/mass spectrometry b) glucocorticoid sensitivity towards proteolysis and lipolysis; c) Vpr concentrations in plasma, abdominal fat- and thigh fat-extracellular fluid; d) detailed body composition and biochemical parameters of HLS. These projects will be performed by a coordinated team experienced in metabolic protocols and stable isotope techniques, HIV clinical specialists, and experts in the molecular biology of the GR and Vpr. They will detail a molecular pathway to this novel lipodystrophic syndrome, which likely predates the use of effective anti-retroviral therapy but comes clinically obvious during therapy as a result of increased nutrient intake, and translate to clinical science the Vpr-mediated mechanism of metabolic dysregulation.