US Military HIV Research Program, Walter Reed Army Institute of Research, Silver Spring, Maryland, USA.
The Henry M. Jackson Foundation for the Advancement of Military Medicine, Bethesda, Maryland, USA.
Microbiol Spectr. 2022 Jun 29;10(3):e0060422. doi: 10.1128/spectrum.00604-22. Epub 2022 May 5.
HIV-1 cure strategies aiming to eliminate persistent infected cell reservoirs are hampered by a poor understanding of cells harboring viral DNA . We describe a novel method to identify, enumerate, and characterize in detail individual cells infected using a combination of single-cell multiplexed assays for integrated proviral DNA, quantitative viral and host gene expression, and quantitative surface protein expression without any manipulation. Latently infected CD4 T cells, defined as harboring integrated provirus in the absence of spliced viral mRNA, were identified from macaque lymph nodes during acute, chronic, and combination antiretroviral therapy (cART)-suppressed simian immunodeficiency virus (SIV) infection. Latently infected CD4 T cells were most abundant during acute SIV (~8% of memory CD4 T cells) and persisted in chronic and cART-suppressed infection. Productively infected cells actively transcribing viral mRNA, by contrast, were much more labile and declined substantially between acute and chronic or cART-suppressed infection. Expression of most surface proteins and host genes was similar between latently infected cells and uninfected cells. Elevated mRNA and surface CD3 expression among latently infected cells suggest increased survival potential and capacity to respond to T cell receptor stimulation. These findings point to a large pool of latently infected CD4 T cells established very early in acute infection and upregulated host factors that may facilitate their persistence , both of which pose potential challenges to eliminating HIV-1 reservoirs. Effective combination antiretroviral therapy controls HIV-1 infection but fails to eliminate latent viral reservoirs that give rise to viremia upon treatment interruption. Strategies to eradicate latently infected cells require a better understanding of their biology and distinguishing features to promote their elimination. Tools for studying these cells from patients are currently limited. Here, we developed a single-cell method to identify cells latently infected and to characterize these cells for expression of surface proteins and host genes without manipulation, capturing their state from SIV-infected macaques. Host factors involved in cell survival and proliferation were upregulated in latently infected cells, which were abundant in the earliest stages of acute infection. These studies provide insight into the basic biology of latently infected cells as well as potential mechanisms underlying the persistence of HIV-1/SIV reservoirs to inform development of novel HIV-1 cure strategies.
旨在消除持续性感染细胞储库的 HIV-1 治愈策略受到对携带病毒 DNA 的细胞了解不足的阻碍。我们描述了一种新方法，可以在无需任何操作的情况下，使用整合前病毒 DNA 的单细胞多重分析、定量病毒和宿主基因表达以及定量表面蛋白表达的组合，来鉴定、计数和详细描述单个感染细胞。在急性、慢性和联合抗逆转录病毒治疗（cART）抑制的猴免疫缺陷病毒（SIV）感染期间，我们从猕猴淋巴结中鉴定出潜伏感染的 CD4 T 细胞，这些细胞定义为在没有剪接病毒 mRNA 的情况下携带整合前病毒。潜伏感染的 CD4 T 细胞在急性 SIV 感染时最为丰富（~记忆 CD4 T 细胞的 8%），并在慢性和 cART 抑制感染中持续存在。相比之下，积极转录病毒 mRNA 的感染细胞非常不稳定，在急性和慢性或 cART 抑制感染之间大量减少。潜伏感染细胞和未感染细胞的大多数表面蛋白和宿主基因表达相似。潜伏感染细胞中升高的 mRNA 和表面 CD3 表达表明其具有更高的生存潜力和对 T 细胞受体刺激的反应能力。这些发现指向在急性感染早期建立的大量潜伏感染 CD4 T 细胞库，并上调了可能促进其持续存在的宿主因子，这两者都对消除 HIV-1 储库构成了潜在挑战。有效的联合抗逆转录病毒治疗可控制 HIV-1 感染，但无法消除潜伏的病毒储库，这些储库在治疗中断时会引发病毒血症。消除潜伏感染细胞的策略需要更好地了解它们的生物学特性和区别特征，以促进其消除。目前用于研究这些患者细胞的工具非常有限。在这里，我们开发了一种单细胞方法来鉴定潜伏感染的细胞，并在无需操作的情况下对这些细胞进行表面蛋白和宿主基因的特征描述，从而从 SIV 感染的猕猴中捕获其状态。参与细胞存活和增殖的宿主因子在潜伏感染细胞中上调，在急性感染的早期阶段非常丰富。这些研究为潜伏感染细胞的基本生物学以及 HIV-1/SIV 储库持续存在的潜在机制提供了深入了解，为开发新的 HIV-1 治愈策略提供了信息。
