Herbein G, Montaner L J, Gordon S
Sir William Dunn School of Pathology, University of Oxford, United Kingdom.
J Virol. 1996 Nov;70(11):7388-97. doi: 10.1128/JVI.70.11.7388-7397.1996.
The proinflammatory cytokine tumor necrosis factor alpha (TNFalpha) is readily detected after human immunodeficiency virus type 1 (HIV-1) infection of primary macrophages in vitro and is present in plasma and tissues of patients with AIDS. Previous studies have shown that human recombinant TNFalpha (hrTNFalpha) enhances HIV replication in both chronically infected promonocytic and T-lymphoid cell lines in vitro. We report here that in contrast to untreated tissue culture-differentiated macrophages (TCDM), in which the proviral long terminal repeat (LTR) could be detected as soon as 8 h postinfection by a PCR assay, TCDM pretreatment for 3 days by hrTNFalpha markedly delayed its appearance until 72 h after infection with the HIV-1 Ada monocytotropic strain. Moreover the inhibition of formation of the proviral LTR in HIV-1-infected TCDM was directly proportional to the concentration of hrTNFalpha used. To determine if the inhibition of LTR formation results from blockade of viral entry, we performed a reverse transcription PCR assay to detect intracellular genomic viral RNA as early as 2 h after infection. Pretreatment of primary TCDM by hrTNFalpha for 3 days and even for only 2 h inhibits 75% of the viral entry into the cells. The inhibition of viral entry by hrTNFalpha was totally abolished by the use of anti-human TNFalpha monoclonal antibody. By using TNFalpha mutants specific for each human TNFalpha receptor, we showed that the inhibition of HIV-1 entry into TCDM was mediated not through the 55-kDa TNF receptor but through the 75-kDa TNF receptor. Although prolonged (1 to 5 days) TNFalpha treatment can downregulate CD4 expression in primary human TCDM, surface CD4 levels were not reduced by 2 h of treatment and was therefore not a limiting step for HIV-1 entry. In contrast to the inhibition of viral entry into primary TCDM, pretreatment with hrTNFalpha did not modify HIV-1 entry into phytohemagglutinin A-activated peripheral blood lymphocytes. TNFalpha-pretreatment inhibited HIV-1 replication in primary TCDM but not in phytohemagglutinin A-activated peripheral blood lymphocytes as assessed by decreased reverse transcriptase activity in culture supernatants. These results demonstrate that TNFalpha is able to enhance host cellular resistance to HIV-1 infection and that selective inhibition of HIV-1 entry into primary TCDM by TNFalpha involves the 75-kDa TNF receptor but not the 55-kDa TNF receptor.
在体外，人免疫缺陷病毒1型（HIV-1）感染原代巨噬细胞后，促炎细胞因子肿瘤坏死因子α（TNFα）很容易被检测到，并且在艾滋病患者的血浆和组织中也存在。先前的研究表明，人重组TNFα（hrTNFα）在体外可增强慢性感染的前单核细胞系和T淋巴细胞系中的HIV复制。我们在此报告，与未处理的组织培养分化巨噬细胞（TCDM）不同，在TCDM中，感染后8小时通过PCR检测即可检测到前病毒长末端重复序列（LTR），而用hrTNFα对TCDM进行3天预处理可显著延迟其出现，直至感染HIV-1嗜单核细胞株Ada 72小时后才出现。此外，在HIV-1感染的TCDM中，前病毒LTR形成的抑制与所用hrTNFα的浓度直接相关。为了确定LTR形成的抑制是否源于病毒进入的阻断，我们在感染后2小时就进行了逆转录PCR检测以检测细胞内基因组病毒RNA。用hrTNFα对原代TCDM进行3天甚至仅2小时的预处理可抑制75%的病毒进入细胞。使用抗人TNFα单克隆抗体可完全消除hrTNFα对病毒进入的抑制作用。通过使用针对每个人TNFα受体的TNFα突变体，我们表明HIV-1进入TCDM的抑制不是通过55 kDa的TNF受体介导的，而是通过75 kDa的TNF受体介导的。尽管长时间（1至5天）的TNFα处理可下调原代人TCDM中的CD4表达，但2小时的处理并未降低表面CD4水平，因此不是HIV-1进入的限制步骤。与抑制病毒进入原代TCDM相反，用hrTNFα预处理并未改变HIV-1进入植物血凝素A激活的外周血淋巴细胞的情况。通过培养上清液中逆转录酶活性的降低评估，TNFα预处理可抑制原代TCDM中的HIV-1复制，但不能抑制植物血凝素A激活的外周血淋巴细胞中的HIV-1复制。这些结果表明，TNFα能够增强宿主细胞对HIV-1感染的抵抗力，并且TNFα对HIV-1进入原代TCDM的选择性抑制涉及75 kDa的TNF受体，而不涉及55 kDa的TNF受体。
