Epithelial gene expression in HIV-associated nephropathy

描述（由申请人提供）： HIV相关性肾病（HIVAN）是HIV-1感染患者慢性肾功能衰竭的最常见原因。HIVAN最显著的组织学特征是肾小管扩张。我们实验室的研究人员先前已经证明，在HIVAN患者的活检标本中，肾小管上皮细胞感染了HIV-1。然而，HIV-1感染后的细胞事件，导致HIVAN进行性肾功能衰竭还没有得到很好的理解。 我们以前已经证明，HIV-1感染可能涉及肾单位几个部分的上皮细胞，包括近端小管。我们已经从HIVAN活检标本中分离出近端肾小管上皮细胞，并用温度敏感的SV 40大T抗原使其有条件地永生化。这些细胞（HPT-1）在允许条件（33 ℃）下无限生长，并且当在37 ℃分化时，表达典型的近端小管表型标记物。 本研究的目的是确定HIV-1感染肾小管上皮细胞后差异表达的宿主基因。一旦确定了候选宿主基因，我们将绘制负责诱导这些基因表达改变的HIV-1。 在所提出的研究中，我们将使用VSV假型复制缺陷型HIV-1 gag/pol缺失构建体来转染HPT-1细胞。转导后，我们将使用代表性差异分析（RDA）和寡核苷酸表达微阵列，以确定细胞基因的差异表达后，由HIV-1转导。将通过标准分子技术确认细胞基因的差异表达。一旦宿主候选基因被确定，我们将用一系列HIV-1多基因突变体构建体转染HPT-1细胞，以定位负责细胞基因表达中观察到的改变的特定HIV-1基因。 这些研究应阐明HIV-1在HIVAN发病的基本机制。也有可能与HIVAN发病机制有关的基因可能是导致非洲裔美国人对几乎所有肾衰竭原因具有明显易感性的重要因素。

DESCRIPTION (provided by applicant): HIV-associated nephropathy (HIVAN) is the most common cause of chronic renal failure in HIV-1 infected patients. The most prominent histologic feature of HIVAN is dilatation of the renal tubules. Investigators in our laboratory have previously demonstrated, in biopsy specimens from patients with HIVAN, that renal tubular epithelial cells are infected with HIV-1. However, the cellular events following HIV-1 infection, contributing to progressive renal failure in HIVAN are not well understood. We have demonstrated previously, that HIV-1 infection may involve epithelial cells from several portions of the nephron, including the proximal tubule. We have isolated proximal tubular epithelial cells from HIVAN biopsy specimens and conditionally immortalized them with the temperature-sensitive SV40 large T antigen. These cells (HPT-1) grow indefinitely under permissive conditions (33C), and when differentiated at 37C, express typical proximal tubular phenotypic markers. The purpose of the proposed-studies is to determine the host genes that are differentially expressed following infection of renal tubular epithelial cells with HIV-1. Once candidate host genes are identified, we will map the HIV-1 responsible for inducing altered expression of these genes. In the proposed studies, we will use a VSV-pseudotyped replication defective HIV-1 gag/pol deletion construct to transduce HPT-1 cells. Following transduction, we will use representational difference analysis (RDA) and oligonucleotide expression micrarrays to determine the cellular genes that are differentially expressed following transduction by HIV-1. Differential expression of cellular genes will be confirmed by standard molecular techniques. Once host candidate genes are identified, we will transduce HPT-1 cells with a series of HIV-1 multigenic mutant constructs to map the specific HIV-1 genes responsible for the observed alterations in cellular gene expression. These studies should elucidate essential mechanisms of HIV-1 pathogenesis in HIVAN. It is also possible that genes implicated in the pathogenesis of HIVAN maybe important factors contributing to the marked predisposition of African-Americans to nearly all causes of renal failure.