Viral and Placental Determinants of HIV-1 Subtype C Mother-to-Child Transmission

描述(申请人提供)：本提案旨在全面描述与HIV-1 C亚型(HIV-1C)母婴传播(MTCT)相关的宿主和病毒生物学。我们假设HIV-1C母婴传播是一个由胎盘和病毒决定因素共同介导的非随机事件。为了解决这一假设，我们将：1)垂直传播包膜(Env)基因的基因型和表型，2)从存档的胎盘活检中分离出的包膜基因的特征，3)描述胎盘转移过程中的胎盘环境。所有实验都将使用之前从744名感染HIV-1C的马拉维孕妇队列中收集的样本，这些孕妇的特征是不传播(NT)、宫内(IU)母子对(MOP)和产中(IP)MOPS。在一项初步研究中，我们列举了22个MOP中HIV-1C V1A/2 env多样性，发现婴儿的变异明显少于他们的母亲，并证实HIV env多样性在母婴传播过程中受到限制。在这笔赠款的指导部分(具体目标1)，我将测试限制是由特定的env基因类型促进的假设。这将通过克隆全长包膜基因来完成：a)比较V1/V2环的长度，b)列举假定的糖基化位点，c)针对CCR5、CXCR4、CD4hi和CD4lo细胞对克隆的env基因进行假型。作为一名独立的调查者，我将专注于ID MTCT，并检验胎盘是一个独特的HIV-1C隔室的假设。使用异源双链跟踪分析，我将比较匹配的外周血、胎盘血、胎盘活检组织和脐带血中的HIV-1C env基因(特定目标2)。将使用在指导阶段获得的方法来描述特定于胎盘的变异。在特定目标3中，针对原代胎盘滋养层细胞和霍夫鲍尔细胞，对传播的和胎盘特异的变异体进行假定型。最后，在特定目标4中，将分析胎盘活检组织中HIV-1C、CCR5、CD4和CXCR4的表达。该项目的结果将全面描述艾滋病毒-1C包膜基因和有利于宫内艾滋病毒-1C母婴传播的胎盘环境。

DESCRIPTION (provided by applicant): This proposal aims to comprehensively describe host and virus biology associated with HIV-1 subtype C (HIV-1C) mother-to-child transmission (MTCT). We hypothesize that HIV-1 C MTCT is a non-stochastic event mediated by both placental and viral determinants. To address this hypothesis, we will: 1) genotype and phenotype vertically transmitted envelope (env) genes, 2) characterize envelope genes isolated from archived placental biopsies, 3) describe the placental environment during MTCT. All experiments will use previously collected samples from a well-characterized cohort of 744 HIV-1 C-infected pregnant Malawian women characterized as nontransmitting (NT), in utero (IU) mother-offspring pairs (MOPs) and intrapartum (IP) MOPs. In a preliminary study, we enumerated HIV-1 C V1 A/2 env diversity in 22 MOPs and found that infants had significantly fewer variants than their mothers and confirmed that HIV env diversity is restricted during MTCT. During the mentored portion of this grant (Specific Aim 1), I will test the hypothesis that restriction is facilitated by specific env genotypes. This will be accomplished by cloning full-length envelope genes to: a) compare V1/V2 loop lengths, b) enumerate putative glycosylation sites, c) pseudotype the cloned env genes against CCR5, CxCR4, CD4hi and CD4lo cells. As an independent investigator, I will focus on ID MTCT and test the hypothesis that the placenta is a unique HIV-1 C compartment. Using a heteroduplex tracking assay, I will compare HIV-1 C env genes from matched peripheral blood, placental blood, placental biopsies, and cord blood (Specific Aim 2). Placental specific variants will be characterized using the methods acquired during the mentored phase. In Specific Aim 3, transmitted and placental specific variants will be psuedotyped against primary placental trophoblasts and Hofbauer cells. Finally, in Specific Aim 4, placental biopsies will be analyzed for HIV-1 C, CCR5, CD4, and CXCR4 expression. The results of this project will provide a comprehensive description of the HIV-1 C envelope genes and the placental environment conducive to in utero HIV-1 C MTCT.