VIRAL PATHOGENESIS OF HIV ASSOCIATED NEPHROPATHY

艾滋病毒相关性肾病(HIVAN)现在是非洲第三大导致肾功能衰竭的原因 在美国，慢性肾功能衰竭是HIV-1感染者最常见的原因，肾脏疾病现在是这些患者的第四大死因。越来越多的证据支持HIV-1感染肾小球和肾小管上皮细胞在HIVAN发病机制中的直接作用。此外，对同时来自肾上皮细胞和外周血单核细胞的HIV-1外膜序列的准种分析表明，肾脏特有的亚群与HIV-1在肾间室的活跃复制一致。体外和转基因小鼠模型数据表明，在肾上皮细胞中直接表达HIV-1基因，特别是nef基因，可以引起与HIVAN相关的表型变化和与增殖和分化相关的细胞基因表达的变化。项目2将进一步明确肾上皮细胞感染在HIVAN发病机制中的直接作用。为了确定仅有肾上皮感染是否可以解释这种疾病，将检查具有替代肾脏疾病病因的艾滋病毒感染患者的肾组织。这将包括非洲人后裔以及高加索人。组织检查将包括原位DNA聚合酶链式反应和带有HIV序列的聚合酶链式反应扩增的上皮细胞的验证性激光解剖。为了进一步了解病毒进入这个独特的隔室，我们将通过激光捕获解剖直接从肾上皮获得的HIV-1包膜的表型特征。鉴于我们之前发表的工作和初步数据表明Nef和VPR对足细胞的影响，我们将使用转基因小鼠模型来研究特定细胞类型中表达的单个基因产物在发病机制中的贡献。靶向表达将通过直接表达来自特定部位启动子的转基因或使用有条件的转基因构建物来实现。此外，直接来自肾上皮的nef序列将进行基因和表型特征分析，以确定独特的多态是否与HIVAN的发生有关。拟议的研究应提供关键的 关于上皮性感染在HIVAN发病机制中的作用以及HIV-1与这一独特的宿主之间的相互作用的信息，并将影响预防这一毁灭性并发症的治疗措施。

HIV-associated nephropathy (HIVAN) is now the third leading cause of renal failure in African Americans, the most common cause of chronic renal failure in HIV-1 infected individuals and renal disease is now the fourth leading cause of death in these patients. Accumulating evidence supports a direct role of HIV-1 infection of renal glomerular and tubule epithelial cells in HIVAN pathogenesis. Furthermore, quasispecies analysis of HIV-1 envelope sequences simultaneously derived from renal epithelial cells and peripheral blood mononuclear cells show renal-specific subclusters consistent with active replication of HIV-1 in the renal compartment. In vitro and transgenic mouse model data suggest that direct expression of HIV-1 genes, particularly nef, in renal epithelium can produce phenotypic changes and alterations in expression of cell genes involved in proliferation and differentiation that are consistent with changes associated with HIVAN. Project 2 will further define the direct role of infection of renal epithelial cells in HIVAN pathogenesis. To determine if renal epithelial infection alone can account for the disease, renal tissue will be examined from HIV-infected patients who have an alternative etiology of renal disease. This will include those of African descent as well as Caucasians. Examination of tissue will include in situ DNA PCR and confirmatory laser dissection of epithelial cells with PCR amplification of HIV sequences. To further our understanding of viral lentry into this unique compartment, we will phenotypically characterize the HIV-1 envelopes directly obtained from renal epithelium by laser capture dissection. In light of our previous published work and preliminary data demonstrating the effects of Nef and, to a lesser degree, Vpr on podocytes, we will use transgeneic mouse modeling to study the contribution of individual gene products expressed in specific cell types to pathogenesis. Targeted expression will be achieved either through direct expression of the transgene from site-specific promoters or the use of conditional transgenic constructs. Furthermore, nef sequences derived directly from renal epithelium will be genotypically and phenotypically characterized to determine if unique polymorphisms are associated with the development of HIVAN. The proposed studies should provide critical information regarding the role of epithelial infection in HIVAN pathogenesis and the interaction of HIV-1 with this unique reservoir and will impact on therapeutic interventions to prevent this devastating complication.