Isolevuglandin peptide modification and proteasomal processing is responsible for autoimmune mediated hypertension

项目摘要 据估计，世界上三分之一的人口患有高血压。在过去的十年里，它 免疫激活是高血压的原因之一。活性氧 (ROS)已经被证明是这种疾病的重要催化剂。异evuglandins（isoLG）是 由ROS形成的脂肪酸的氧化产物。这些分子共价加合到 蛋白质的赖氨酸残基。IsoLG加合蛋白由抗原呈递树突状细胞呈递 细胞（DC），并导致CD8 + T细胞活化和由此产生的高血压。这种T细胞亚群是 仅由MHC-I内呈递的肽激活，这意味着蛋白酶体在 肽加工在初步研究中，我发现异LG-加合肽明显 在系统性红斑狼疮（SLE）患者的单核细胞中富集。我提议在 假设蛋白质的IsoLG修饰是SLE高血压的原因， IsoLG修饰的肽的呈递是蛋白酶体依赖性的。在目标1中，我将检验假设 天然蛋白质的IsoLG修饰及其在树突状细胞上的呈递有助于 SLE小鼠模型中高血压和血管炎症。这将通过 利用SLE的B6.SLE123小鼠模型。将分析IsoLG修饰的肽呈递 在基线和高盐处理后通过流式细胞术检测。血压将通过以下方式测量： 颈动脉遥测将通过流式细胞术和组织学检查分析组织炎症。 分析.用异LG清除剂分子2-HOBA和高血压刺激的共同治疗将 然后执行。我预测用2-HOBA治疗SLE小鼠将减轻高血压， 组织炎症在目标2中，我将检验蛋白酶体加工肽的假设， 介导高血压和异LG加合物呈递。已知小鼠树突状细胞的治疗 细胞与氧化剂叔丁基过氧化氢（TBHP）赋予isoLG介导的敏感性， 受体小鼠高血压刺激。我发现树突状细胞与TBHP的共同治疗 而蛋白酶体抑制剂（PI）则可减弱表面异LG呈递。我将野生型小鼠与 高血压刺激和PI。IsoLG表面蛋白的呈递和组织炎症将是 测定了我还表明，蛋白酶体的活性增加，在DC的高血压小鼠。 这与炎性免疫蛋白酶体亚单位LMP7的表达增加相关。我们 已经产生了LMP7条件性敲除小鼠，并将研究LMP7缺失对 高血压和异LG-加合物呈递。 !

PROJECT SUMMARY An estimated one-third of the world’s population suffers from hypertension. Over the last decade, it has become clear that immune activation contributes to hypertension. Reactive oxygen species (ROS) have been shown to be an important activator of this disease. Isolevuglandins (isoLG) are oxidation products of fatty acids that form as a result of ROS. These molecules adduct covalently to lysine residues of proteins. IsoLG adducted proteins are presented by antigen presenting dendritic cells (DCs) and result in CD8+ T-cell activation and resultant hypertension. This subset of T-cells is exclusively activated by peptides presented within MHC-I, implying a role of the proteasome in peptide processing. In preliminary studies I have found that isoLG-adducted peptides are markedly enriched in monocytes of patients with systemic lupus erythematosus (SLE). I propose to address the hypothesis that IsoLG modification of proteins is responsible for hypertension in SLE and that the presentation of IsoLG-modified peptides is proteasome dependent. In Aim 1, I will test the hypothesis that IsoLG modification of native proteins and their presentation on dendritic cells contribute to hypertension and vascular inflammation in a mouse model of SLE. This will be accomplished by utilizing the B6.SLE123 mouse model of SLE. IsoLG modified peptide presentation will be analyzed by flow cytometry at baseline, and after treatment with high salt. Blood pressure will be measured by carotid artery telemetry. Tissue inflammation will be analyzed by flow cytometry and histologic analysis. Co-treatment with an isoLG scavenger molecule 2-HOBA and hypertensive stimulus will then be performed. I predict that treatment of SLE mice with 2-HOBA will attenuate hypertension and tissue inflammation. In Aim 2, I will test the hypothesis that proteasomal processing of peptides mediates hypertension and isoLG-adduct presentation.. It is known that treatment of mouse dendritic cells with the oxidant tert-butyl hydroperoxide (TBHP) confers isoLG-mediated sensitivity to hypertensive stimulus in recipient mice. I have found that co-treatment of dendritic cells with TBHP and proteasome inhibitor (PI) attenuates surface isoLG presentation. I will co-treat wild-type mice with hypertensive stimulus and a PI. IsoLG surface protein presentation and tissue inflammation will be measured. I have also shown that proteasomal activity is increased in DC’s from hypertensive mice. This correlates with increased expression of the inflammatory immunoproteasomal subunit LMP7. We have generated an LMP7 conditional knockout mouse and will study the effects of LMP7 deletion on hypertension and isoLG-adduct presentation in DCs. !