Role Of Viral Reservoirs In The Pathogenesis Of Hiv Dise

在过去的几年里，我们一直在研究病毒储库在感染个体中HIV疾病发病机制中的作用。尽管开发了高活性抗病毒治疗（HAART）及其在治疗HIV感染个体中的巨大成功，但已经清楚地证明，在接受HAART的大多数感染个体中，病毒复制持续存在，其中血浆病毒血症已降至检测限以下。为了更深入了解病毒宿主在艾滋病病毒感染中的作用，我们在过去一年进行了以下两项研究：1）确定低水平的正在进行的病毒复制与免疫学参数如CD 4 +/CD 8 + T细胞比率之间的关系，在血浆病毒血症已被HAART成功抑制的感染个体中，以及2）检查病毒血症与病毒血症个体中潜伏感染的静息CD 4 + T细胞中病毒表达程度的范围和HIV-1持续存在的潜在机制。在第一项研究中，我们已经证明了在接受HAART的病毒血症感染个体中，携带HIV-1前病毒DNA的CD 4 + T细胞频率与CD 4 +/CD 8 + T细胞比率之间存在统计学显著的负相关性，其中血浆病毒血症已被抑制至检测限以下较长时间（>2.5年）。HIV-1特异性细胞毒性CD 8 + T淋巴细胞（CTL）的频率与这些个体中的CD 4 +/CD 8 + T细胞比率之间没有发现相关性。我们的数据表明，持续的，低水平的，正在进行的病毒复制，虽然不足以维持HIV-1特异性CTL反应，可能部分解释为什么正常化的CD 4 +/CD 8 + T细胞比例没有实现在一些感染的个人成功地与HAART治疗。HIV-1存在于潜伏感染的静息CD 4 + T细胞中已在感染个体中得到明确证实;然而，病毒表达程度的范围和HIV-1在该潜伏病毒库中持续存在的潜在机制尚未完全阐明。在第二项研究中，我们已经表明，我们检查的大多数病毒血症患者的高度纯化的静息CD 4 + T细胞能够通过RT-PCR测量自发地体外产生无细胞HIV-1。在存在细胞增殖和病毒复制抑制剂的情况下，静息CD 4 + T细胞释放的HIV-1水平没有显著降低。相反，从大多数病毒血症患者中获得的静息CD 4 + T细胞（与从病毒血症患者中获得的细胞表型相同）未能产生无细胞HIV-1，尽管HIV-1前病毒DNA和细胞相关HIV-1 RNA的水平与病毒血症患者相当。静息CD 4 + T细胞的DNA微阵列分析表明，涉及转录调控，RNA加工和修饰，蛋白质运输和囊泡运输的一些基因在病毒血症患者的静息CD 4 + T细胞与病毒血症患者相比显著上调。这些结果表明，活跃的病毒复制，表现为可检测的血浆病毒血症，有一个显着的影响，在感染患者的静息CD 4 + T细胞的生理状态，并反过来，允许释放无细胞的HIV-1没有外源性激活刺激。此外，鉴于尽管存在细胞相关HIV-1 RNA，但大多数病毒血症患者的潜伏病毒库未产生可定量的病毒体，因此，在有效治疗期间，病毒血症患者静息CD 4 + T细胞中HIV-1 RNA转录的证据不一定被视为病毒复制持续的直接证据。

Over the past several years, we have been investigating the role of viral reservoirs in the pathogenesis of HIV disease in infected individuals. Despite development of highly active antiviral therapy (HAART) and its enormous success in the treatment of HIV-infected individuals, it has been clearly demonstrated that viral replication persists in the majority of infected individuals receiving HAART in whom plasma viremia has fallen below the limit of detection. In order to better understand the role of viral reservoirs in HIV infection, we have conducted the following two studies in the past year: 1) determination of the relationship between low levels of on-going viral replication and immunologic parameters, such as CD4+/CD8+ T cell ratios, in infected individuals in whom plasma viremia had been successfully suppressed by HAART and 2) examination of the range of the extent of viral expression and the underlying mechanisms of the persistence of HIV-1 in latently infected, resting CD4+ T cells in viremic versus aviremic individuals. In the first study, we have demonstrated a statistically significant inverse correlation between the frequency of CD4+ T cells carrying HIV-1 proviral DNA and the CD4+/CD8+ T cell ratios in aviremic infected individuals receiving HAART in whom plasma viremia had been suppressed below the limit of detection for prolonged periods of time (>2.5 years). No correlation was found between the frequency of HIV-1-specific cytotoxic CD8+ T lymphocytes (CTL) and the CD4+/CD8+ T cell ratios in those individuals. Our data suggest that persistent, low-level, on-going viral replication, although not sufficient to maintain HIV-1-specific CTL responses, may explain in part why normalization of the CD4+/CD8+ T cell ratios is not achieved in some infected individuals successfully treated with HAART. The presence of HIV-1 in latently infected, resting CD4+ T cells has been clearly demonstrated in infected individuals; however, the range of the extent of viral expression and the underlying mechanisms of the persistence of HIV-1 in this latent viral reservoir have not been fully delineated. In the second study, we have shown that highly purified resting CD4+ T cells from the majority of viremic patients whom we examined were capable of producing cell-free HIV-1 spontaneously ex vivo as measured by RT-PCR. The levels of HIV-1 released by resting CD4+ T cells were not significantly reduced in the presence of inhibitors of cellular proliferation and viral replication. In contrast, resting CD4+ T cells obtained from the majority of aviremic patients, which were phenotypically identical to those obtained from viremic patients, failed to produce cell-free HIV-1, despite levels of HIV-1 proviral DNA and cell-associated HIV-1 RNA comparable to those of viremic patients. DNA microarray analysis of resting CD4+ T cells demonstrated that a number of genes involving transcription regulation, RNA processing and modification, and protein trafficking and vesicle transport were significantly upregulated in resting CD4+ T cells from viremic patients compared to those of aviremic patients. These results suggest that active viral replication, as manifested by detectable plasma viremia, has a significant impact on the physiologic state of resting CD4+ T cells in infected patients, and in turn, allows release of cell-free HIV-1 without exogenous activation stimuli. In addition, given that no quantifiable virions were produced by the latent viral reservoir in the majority of aviremic patients despite the presence of cell-associated HIV-1 RNA, evidence for transcription of HIV-1 RNA in resting CD4+ T cells of aviremic patients should not necessarily be taken as direct evidence for ongoing viral replication during effective therapy.