Department of Neuroscience, Center for Neurovirology, Lewis Katz School of Medicine at Temple University, Philadelphia, PA, United States of America.
PLoS One. 2020 Nov 2;15(11):e0241667. doi: 10.1371/journal.pone.0241667. eCollection 2020.
HIV-associated CNS dysfunction is a significant problem among people with HIV (PWH), who now live longer due to viral suppression from combined anti-retroviral therapy (ART). Over the course of infection, HIV generates toxic viral proteins and induces inflammatory cytokines that have toxic effects on neurons in the CNS. Among these viral proteins, HIV Nef has been found in neurons of postmortem brain specimens from PWH. However, the source of Nef and its impact on neuronal cell homeostasis are still elusive.
Here, in using a simian immunodeficiency virus (SIV) infected rhesus macaque model of neuroHIV, we find SIV Nef reactivity in the frontal cortex, hippocampus and cerebellum of SIV-infected animals using immunohistochemistry (IHC). Interestingly, SIV-infected macaques treated with ART also showed frequent Nef positive cells in the cerebellum and hippocampus. Using dual quantitative RNAscope and IHC, we observed cells that were positive for Nef, but were not for SIV RNA, suggesting that Nef protein is present in cells that are not actively infected with SIV. Using cell specific markers, we observed Nef protein in microglia/macrophages and astrocytes. Importantly, we also identified a number of NeuN-positive neurons, which are not permissive to SIV infection, but contained Nef protein. Further characterization of Nef-positive neurons showed caspase 3 activation, indicating late stage apoptosis in the CNS neurons.
Our results suggest that regardless of ART status, Nef is expressed in the brain of SIV infected macaques and may contribute to neurological complications seen in PWH.
HIV 相关的中枢神经系统功能障碍是 HIV 感染者（PWH）面临的一个重大问题，由于联合抗逆转录病毒疗法（ART）的病毒抑制作用，他们现在的寿命更长。在感染过程中，HIV 会产生有毒的病毒蛋白，并诱导炎症细胞因子，对中枢神经系统中的神经元产生毒性作用。在这些病毒蛋白中，已在 PWH 的尸检脑组织标本的神经元中发现了 HIV Nef。然而，Nef 的来源及其对神经元细胞内稳态的影响仍不清楚。
在这里，我们使用感染了猴免疫缺陷病毒（SIV）的恒河猴神经 HIV 模型，通过免疫组织化学（IHC）发现 SIV Nef 在 SIV 感染动物的额叶皮层、海马体和小脑中有反应性。有趣的是，接受 ART 治疗的 SIV 感染猕猴的小脑和海马体中也经常出现 Nef 阳性细胞。使用双重定量 RNAscope 和 IHC，我们观察到对 Nef 呈阳性但对 SIV RNA 呈阴性的细胞，这表明 Nef 蛋白存在于未被 SIV 感染的细胞中。使用细胞特异性标志物，我们观察到 Nef 蛋白存在于小胶质细胞/巨噬细胞和星形胶质细胞中。重要的是，我们还鉴定了一些 NeuN 阳性神经元，这些神经元对 SIV 感染没有易感性，但含有 Nef 蛋白。对 Nef 阳性神经元的进一步特征分析显示 caspase 3 激活，表明中枢神经系统神经元处于晚期凋亡阶段。
我们的结果表明，无论 ART 状态如何，Nef 在 SIV 感染的猕猴大脑中均有表达，可能导致 PWH 中出现神经并发症。
