Antiadipogenic Influence of HIV Protease Inhibitors

拟议研究的主要目的是阐明接受HIV蛋白酶抑制剂(PI)治疗的HIV+患者发生脂营养不良综合征(LDS)的分子基础。LDS的特征是皮下脂肪组织脂肪萎缩，并伴有内脏脂肪组织肥大。腰椎间盘突出症患者经常出现胰岛素抵抗和高甘油三酯血症的迹象。我们的中心假设是，脂肪萎缩与PI对皮下脂肪细胞的选择性萎缩作用有关，从而导致内脏脂肪代偿性增加。内脏肥胖症的增加促进了胰岛素抵抗，增加了非酯化脂肪酸进入肝脏的流量，并增加了极低密度脂蛋白-甘油三酯的产生和分泌。我们使用模型前脂肪细胞系(3T3-L1和-F442a)的初步结果表明，几种PI抑制前脂肪细胞分化和/或拮抗脂肪细胞表型(促进甘油三酯丢失，脂肪细胞特异性蛋白表达丢失，并可能通过凋亡机制降低存活率)。因此，PI可能通过影响脂肪细胞的活性和/或通过抑制脂肪生成来阻止丢失的脂肪细胞的替换，从而促进LDS患者的皮下脂肪萎缩。我们推测，PI通过阻止成脂转录因子CCAAT/增强子结合蛋白(C/EBP)α和过氧化物酶体增殖物激活受体(PPAR)γ的表达来阻止脂肪形成，因为这些是PI最早抑制的可观察到的分子标志物。有待研究的潜在PI依赖效应可能阻断C/EBPalpha和/或PPARGamma的表达，包括：1)抑制C/EBPbeta/Delta依赖的C/EBPalpha和PPARGamma基因的转录激活，2)不能取消抑制C/EBP未分化蛋白(CUP)/AP-2α和Sp1依赖的C/EBPalpha基因的表达，以及3)不能失活反脂肪成脂的Wnt信号通路。我们假设皮下脂肪细胞比内脏脂肪细胞对PI的拮抗作用更敏感。人的原代脂肪细胞将从皮下和内脏库中分离出来，并检查PI诱导的不同效应。有待研究的潜在的依赖PI的机制包括：1)激活TNFpha信号转导通路的下游信号成分；2)抑制GLUT4葡萄糖转运蛋白的功能。在小鼠体内的前体脂肪细胞植入技术将被用来测试所提出的PI作用的分子机制。这项拟议研究的长期目标是促进缺乏脂肪萎缩潜力的抗艾滋病毒疗法的设计。

The primary objective of the proposed studies is to elucidate the molecular basis of the lipodystrophy syndrome (LDS) in HIV+ patients treated with HIV protease inhibitors (PIs). LDS is characterized by lipoatrophy of subcutaneous adipose tissue with accompanying hypertrophy of visceral adipose tissue. Patients with LDS often show signs of insulin resistance and hypertriglyceridemia. Our central hypothesis is that the lipoatrophy is related to a selective atrophic effect of PIs on the subcutaneous adipocyte leading to a compensatory increase in visceral adiposity. The increased visceral adiposity promotes insulin resistance, increased flux of non-esterified fatty acid to the liver and increased very low density lipoprotein- triglyceride production and secretion. Our preliminary results using model preadipocyte cell lines (3T3-L1 and -F442A) demonstrate that several PIs inhibit preadipocyte differentiation and/or antagonize the adipocyte phenotype (promote loss of triglyceride, loss of adipocyte-specific protein expression and decreased viability perhaps via an apoptotic mechanism). Thus, PIs may promote subcutaneous lipoatrophy in patients with LDS by compromising adipocyte viability and/or preventing replacement of lost adipocytes by inhibiting adipogenesis. We hypothesize that PIs block adipogenesis by preventing expression of the adipogenic transcription factors CCAAT/enhancer-binding protein (C/EBP) alpha and peroxisome proliferator-activated receptor (PPAR) gamma as these are the earliest observable molecular markers to be repressed by PIs. Potential PI-dependent effects to be investigated and that may block expression of C/EBPalpha and/or PPARgamma include: 1) Inhibition of C/EBPbeta/delta-dependent transcriptional activation of C/EBPalpha and PPARgamma gene expression, 2) Failure to derepress C/EBP undifferentiated protein (CUP)/AP-2alpha- and Sp1-dependent repression of C/EBPalpha gene expression and 3) Failure to inactivate antiadipogenic Wnt signaling pathways. We hypothesize that subcutaneous adipocytes are more susceptible than visceral adipocytes to the antagonistic effects of PIs. Human primary adipocytes will be isolated from subcutaneous and visceral depots and examined for differential PI-induced effects. Potential PI- dependent mechanisms to be investigated include: 1) Activation of downstream signaling components in the TNFalpha signal transduction pathway and 2) Inhibition of GLUT4 glucose transporter function. An in vivo preadipocyte implantation technique in mice will be used to test proposed molecular mechanisms of PI action. The long term objective of the proposed research is to facilitate the design of anti-HIV therapeutics that lack lipoatrophic potential.