Role of Viral Reservoirs in the Pathogenesis of HIV Disease

在过去的几年里，我们一直在研究潜伏感染的静息CD4+ T细胞和持续的病毒复制在HIV疾病发病机制中的作用，以及这种储存库对HIV感染者治疗的影响。我们以前证明，潜伏的病毒水库在休息的CD4+ T细胞区室坚持在几乎所有感染的个人接受有效的抗病毒治疗。因此，这种病毒库是体内根除HIV的主要障碍。此外，我们认识到，在接受有效抗病毒治疗的慢性感染者中，HIV持续以低水平复制，使他们在很长一段时间内持续出现病毒血症。在过去的一年里，我们的研究重点是：1）描述艾滋病毒在接受有效抗病毒治疗的感染者中持续存在的机制，2）调查在疾病的不同阶段开始抗逆转录病毒治疗的感染者中病毒储存库的衰变动力学。 首先，我们研究了长期接受有效抗逆转录病毒治疗的个体中残留HIV的存在和状态，并研究了HIV在这些个体的CD4+ T细胞中持续存在的潜在机制。我们证明，在肠道相关淋巴组织（GALT）中的CD4+ T细胞携带最高水平的HIV前病毒DNA相比，CD4 + T细胞在感染的个人接受有效的抗逆转录病毒治疗的时间较长的血液。HIV env序列的系统发育分析显示，血液和GALT相关的CD4+ T细胞之间存在活跃的交叉感染。此外，我们的数据还表明，加强抗逆转录病毒治疗，特别是通过增加新的类抗艾滋病毒药物，将是必要的，以遏制低水平的正在进行的病毒复制在慢性感染个体的组织室，以实现完全清除病毒在体内。 其次，我们研究了在疾病的不同阶段开始抗逆转录病毒治疗的感染个体中病毒储存库的衰减动力学。在一项横断面研究中，我们证明了在感染后6个月内开始抗逆转录病毒治疗的受试者中HIV前病毒DNA的中位拷贝数显著低于在感染慢性期开始抗逆转录病毒治疗的受试者（p=0.003）。为了检查携带感染性病毒的CD4+ T细胞的频率，使用来自8名感染个体的高度富集的CD4+ T细胞进行了高输入共培养试验，该试验允许检查大量细胞，在这些感染个体的细胞中未检测到可测量的HIV前病毒DNA。在感染后6个月内开始治疗的HIV感染者中携带感染性病毒的细胞频率显著低于在感染慢性期开始治疗的HIV感染者（p=0.03）。值得注意的是，从一个感染个体的外周血或GALT CD4+ T细胞中不能回收感染性病毒（每0.8 × 109个CD4+ T细胞<1个感染细胞），其中在感染早期开始抗逆转录病毒治疗。我们的数据表明，在HIV感染过程中早期开始抗逆转录病毒治疗和长期抑制病毒复制相结合，可导致HIV储库的深刻减少，从而可能导致感染个体中的病毒清除。

For the past several years, we have been investigating the role of latently infected, resting CD4+ T cells and persistent viral replication in the pathogenesis of HIV disease and the impact of this reservoir on the treatment of HIV-infected individuals. We previously demonstrated that the latent viral reservoir in the resting CD4+ T cell compartment persists in virtually all infected individuals receiving effective antiviral therapy. Consequently, this viral reservoir is a major impediment to the eradication of HIV in vivo. In addition, we realized that HIV continually replicates at low levels in chronically infected individuals who receive effective antiviral therapy that renders them consistently aviremic for prolonged periods of time. Over the past year, we have focused our research on: 1) delineation of the mechanism by which HIV persists in infected individuals receiving effective antiviral therapy for extended periods of time and 2) investigation of the dynamics of decay of viral reservoirs in infected individuals who initiate antiretroviral therapy at different stages of disease. First, we investigated the presence and status of residual HIV in individuals who had received effective antiretroviral therapy for prolonged periods of time and examined the underlying mechanisms by which HIV persists in CD4+ T cells of such individuals. We demonstrated that CD4+ T cells in the gut-associate lymphoid tissue (GALT) carried the highest level of HIV proviral DNA compared to CD4+ T cells in the blood of infected individuals receiving effective antiretroviral therapy for prolonged periods of time. Phylogenetic analyses of HIV env sequences revealed active cross-infection between blood- and GALT-associated CD4+ T cells. Furthermore, our data also suggest that intensification of antiretroviral therapy, especially through the addition of newer classes of anti-HIV drugs, will be necessary to contain low levels of on-going viral replication in the tissue compartment of chronically infected individuals in order to achieve complete clearance of virus in vivo. Second, we investigated the dynamics of decay of viral reservoirs in infected individuals who initiated antiretroviral therapy at different stages of disease. In a cross-sectional study, we demonstrated that the median copy number of HIV proviral DNA in subjects who initiated antiretroviral therapy within 6 months of infection was significantly lower compared to subjects initiating antiretroviral therapy during the chronic phase of infection (p=0.003). In order to examine the frequency of CD4+ T cells carrying infectious virus, a High Input Co-culture assay, which allows examination of large numbers of cells, was conducted using highly enriched CD4+ T cells from 8 infected individuals in whom no measurable HIV proviral DNA had been detected in their cells. The frequency of cells carrying infectious virus in HIV-infected individuals who initiated therapy within 6 months of infection was significantly lower compared to HIV-infected individuals who initiated therapy during the chronic phase of infection (p=0.03). Remarkably, no infectious virus could be recovered from the peripheral blood or GALT CD4+ T cells of one infected individual (<1 infected cell per 0.8x109 CD4+ T cells) in whom antiretroviral therapy was initiated early in infection. Our data suggest that the combination of initiation of antiretroviral therapy early in the course of HIV infection and prolonged suppression of viral replication can result in a profound diminution of HIV reservoirs that may lead to clearance of virus in infected individuals.