HDL prevention of cholesterol crystal inflammation in HIV disease

描述(申请人提供)：艾滋病毒感染和抗逆转录病毒治疗与动脉粥样硬化性血管疾病的增加有关，在晚期表现为富含结晶胆固醇的坏死性病变。这种损伤容易发生灾难性的破裂，导致血栓血管闭塞、心肌梗死和中风，这些都是艾滋病毒感染者死亡的重要原因。长期以来的理论认为，结晶胆固醇是动脉粥样硬化的终末期产物，在显微镜下可以看出是组织切片上的裂缝。在这里，使用共聚焦荧光和激光反射显微镜的新组合提出的证据表明，与公认的教条相反，胆固醇晶体(CCS)在动脉粥样硬化的进展早期形成，这些晶体激活NLRP3炎症体，导致IL-12细胞因子的产生和促进小鼠动脉粥样硬化负荷的增加。重要的是，我们证明了健康人的血浆可以溶解巨噬细胞的胆固醇晶体，并根据我们的初步数据假设高密度脂蛋白(HDL)是防止血管壁胆固醇晶体形成的重要血清因素。此外，我们假设由HIV感染引起的个体炎症状态的变化会导致高密度脂蛋白的蛋白质组和脂肪含量的重塑，并改变颗粒防止血管壁胆固醇的能力 晶体含量和动脉粥样硬化负荷。为了探索这一假设，并检验人类心血管疾病的风险是否与高密度脂蛋白阻止胆固醇晶体形成的能力相关，我们将；1)使用一系列增加脂肪含量的饮食，在LDLR-/-小鼠动脉粥样硬化模型中建立胆固醇结晶与血浆VLDL/LDL水平成比例的关系：2)使用apoA-I-/-和基于LDLR-/-的人apoA-I转基因模型，评估apoA-I表达是否调节动脉粥样硬化病变中胆固醇结晶的倾向：3)评估重组胆固醇低密度脂蛋白急性治疗是否导致LDLR-/-小鼠动脉粥样硬化病变中已建立的胆固醇晶体退化，并测试这些高密度脂蛋白的影响是否依赖于ABCA1胆固醇外流转运体；4)测定一组正常人和HIV感染者的高密度脂蛋白颗粒蛋白质组和脂质含量，并测试与CC溶解、抗炎能力和心血管疾病风险的相关性，如冠脉计算机断层扫描所测量的那样，以及18F-脱氧葡萄糖-PET成像所评估的病变炎症状态。这项提议将检验新的假设，即高密度脂蛋白的功能与其阻止胆固醇晶体形成的能力有关。 而这一功能的变异性与个人患心血管疾病的风险相关。如果被证实是真的，通过使用质谱学来定量个人的高密度脂蛋白和脂肪含量，并将其与晶体溶解能力和心血管疾病相关联，这项工作有望识别出高密度脂蛋白功能的新生物标记物。这些生物标志物将有助于寻找治疗目标，在艾滋病毒感染的背景下减少心血管疾病，并可能在普通人群中减少心血管疾病。 公共卫生相关性：艾滋病毒感染和抗逆转录病毒治疗与动脉粥样硬化性血管疾病的增加有关，在晚期表现为富含结晶胆固醇的坏死性病变。这种损伤容易发生灾难性的破裂，导致心脏病发作和中风，这是现在艾滋病毒感染者死亡的重要原因。这里提出的证据表明，与公认的教条相反，胆固醇晶体(CCS)在动脉粥样硬化早期形成，会引起炎症，而高密度脂蛋白可以防止CC的形成和炎症。我们将研究高密度脂蛋白预防CC炎症的机制，并将在HIV+患者身上测试感染和抗逆转录病毒治疗是否破坏 高密度脂蛋白预防CC形成和炎症的能力。如果被证明是真的，这将代表着我们对高密度脂蛋白如何发挥作用以预防心血管疾病的理解的范式转变，并可能导致这种疾病的新疗法。

DESCRIPTION (provided by applicant): HIV infection and antiretroviral therapy has been associated with increased atherosclerotic vascular disease, which at an advanced stage presents as necrotic lesions rich in crystalline cholesterol. Such lesions are prone to catastrophic rupture that initiates thrombotic vessel occlusion, myocardial infarcts and stroke, now important causes of mortality in HIV infected individuals. Microscopically discernable as clefts in histologic sections, long standing theory considers crystalline cholesterol as an end stage product of atherosclerosis. Here evidence is presented using a novel combination of confocal fluorescent and laser reflection microscopy that shows, in contrast to accepted dogma, cholesterol crystals (CCs) form early in the progression of atherosclerosis and that these crystals activate the NLRP3 inflammasome, leading to IL-12 cytokine production and the promotion of atheroma burden in mice. Importantly, we show that human plasma from healthy individuals can dissolve macrophage cholesterol crystals and postulate from our preliminary data that high-density lipoproteins (HDL) are an important serum factor responsible for preventing vessel wall cholesterol crystal formation. Further, we hypothesize that variation in an individual's inflammatory status as driven by HIV infection causes remodeling of the proteomic and lipid content of HDL, and alters the ability of the particle to prevent vessel wall cholesterol crystal content and atherosclerotic burden. To explore this hypothesis and test whether risk of cardiovascular disease in man is correlated to the ability of HDL to prevent cholesterol crystal formation we will; 1) Establish that cholesterol crystallization is proportional to plasma VLDL/LDL levels in the LDLR-/- mouse model of atherosclerosis using a series of diets with increasing fat content: 2) Assess if apoA-I expression modulates the propensity of cholesterol to crystallize in atherosclerotic lesions using the apoA-I-/- and human apoA-I transgenic models on an LDLR-/- background: 3) Assess whether acute treatment with reconstituted cholesterol poor HDL causes regression of established cholesterol crystals in atherosclerotic lesions of LDLR-/- mice and test if these effects of HDL depend on ABCA1 cholesterol efflux transporter; 4) Determine the HDL particle proteomic and lipid content from a cohort of normal and HIV infected individuals and test for correlations to CC dissolution, anti-inflammatory capacity and CVD disease risk as measured by coronary computed tomography and lesion inflammatory status as assessed by 18F-fluorodeoxyglucose-PET imaging. This proposal will test the novel hypothesis that HDL functionality is linked to its ability to prevent cholesterol crystal formation and that variability in this function is correlated to an individual's risk of cardiovascular diseae. If proven true, by using mass spectrometry to quantitate an individual's HDL protein and lipid content and correlating this with crystal dissolution potency and cardiovascular disease this work expects to identify new biomarkers for HDL function. Such biomarkers will help in the search for therapeutic targets to reduce cardiovascular disease in the context of HIV infection and likely in the general population. PUBLIC HEALTH RELEVANCE: HIV infection and antiretroviral therapy has been associated with increased atherosclerotic vascular disease, which at an advanced stage presents as necrotic lesions rich in crystalline cholesterol. Such lesions are prone to catastrophic rupture, which causes heart attack and stroke, now important causes of mortality in HIV infected individuals. Here evidence is presented that shows, in contrast to accepted dogma, cholesterol crystals (CCs) form early in atherosclerosis, cause inflammation and that HDL can prevent CC formation and inflammation. We will investigate the mechanisms by which HDL prevents CC inflammation and will test in HIV+ individuals whether infection and antiretroviral therapy disrupt the ability HDL to prevent CC formation and inflammation. If shown to be true, this will represent a paradigm shift in our understanding of how HDL functions to prevent cardiovascular disease and may lead to new treatments for this disease.