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

描述（由申请人提供）：本提案旨在全面描述与HIV-1亚型C （HIV-1C）母婴传播（MTCT）相关的宿主和病毒生物学。我们假设hiv - 1c MTCT是由胎盘和病毒决定因素介导的非随机事件。为了解决这一假设，我们将：1)基因型和表型垂直传播包膜（env）基因，2)表征从存档的胎盘活检中分离的包膜基因，3)描述MTCT期间的胎盘环境。所有实验将使用先前从744名hiv - 1c感染的马拉维孕妇中收集的样本，这些孕妇的特征为非传播性（NT）、子宫内（IU）母婴对（MOPs）和产内（IP） MOPs。在一项初步研究中，我们在22个MOPs中枚举了HIV-1 C V1 a /2的环境多样性，发现婴儿的变异明显少于母亲，并证实了母婴传播期间HIV环境多样性受到限制。在此资助的指导部分（具体目标1），我将验证特定环境基因型促进限制的假设。这将通过克隆全长包膜基因来完成：a)比较V1/V2环长度，b)枚举假定的糖基化位点，c)克隆的env基因针对CCR5、CxCR4、CD4hi和CD4lo细胞进行假型。作为一名独立研究者，我将专注于ID MTCT，并验证胎盘是一个独特的hiv - 1c隔室的假设。使用异源双工跟踪测定，我将比较来自匹配外周血、胎盘血、胎盘活检和脐带血的hiv - 1c环境基因（特异性目标2）。胎盘特异性变异将使用在指导阶段获得的方法进行表征。在Specific Aim 3中，将对原代胎盘滋养细胞和霍夫鲍尔细胞进行遗传变异和胎盘特异性变异的推测分型。最后，在特异性靶4中，胎盘活检将分析hiv - 1c、CCR5、CD4和CXCR4的表达。该项目的结果将提供一个全面的描述hiv - 1c包膜基因和胎盘环境有利于在子宫内hiv - 1c MTCT。

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.