Department of Pathology and Laboratory Medicine, University of Wisconsin-Madison, Madison, Wisconsin, United States of America.
Department of Microbial Pathogenesis, Yale University School of Medicine, New Haven, Connecticut, United States of America.
PLoS Pathog. 2024 Oct 28;20(10):e1012665. doi: 10.1371/journal.ppat.1012665. eCollection 2024 Oct.
Antibody-dependent cellular phagocytosis (ADCP) has been implicated in protection against HIV-1. However, methods for measuring ADCP currently rely on the phagocytosis of gp120- or gp41-coated beads that do not reflect physiologically relevant conformations of the viral envelope glycoprotein or the size of a virus-infected cell. We therefore developed a novel approach for measuring ADCP of HIV-infected cells expressing natural conformations of Env. A monocytic cell line (THP-1 cells) or primary human monocytes were incubated with a CD4+ T cell line that expresses eGFP upon HIV-1 infection in the presence of antibodies and ADCP was measured as the accumulation of eGFP+ material by flow cytometry. The internalization of HIV-infected cells by monocytes was confirmed visually by image-capture flow cytometry. Cytoskeletal remodeling, pseudopod formation and phagocytosis were also observed by confocal microscopy. We found that potent broadly neutralizing antibodies (bnAbs), but not non-neutralizing antibodies (nnAbs), mediate efficient phagocytosis of cells infected with either primary or lab-adapted HIV-1. A nnAb to a CD4-inducible epitope of gp120 (A32) failed to enable ADCP of HIV-infected cells but mediated efficient phagocytosis of gp120-coated beads. Conversely, a bnAb specific to intact Env trimers (PGT145) mediated potent ADCP of HIV-infected cells but did not facilitate the uptake of gp120-coated beads. These results underscore the importance of measuring ADCP of HIV-infected cells expressing physiologically relevant conformations of Env and show that most antibodies that are capable of binding to Env trimers on virions to neutralize virus infectivity are also capable of binding to Env on the surface of virus-infected cells to mediate ADCP.
抗体依赖的细胞吞噬作用（ADCP）被认为与 HIV-1 的保护有关。然而，目前用于测量 ADCP 的方法依赖于吞噬 gp120 或 gp41 包被的珠粒，这些方法不能反映病毒包膜糖蛋白的生理相关构象或感染病毒的细胞的大小。因此，我们开发了一种测量表达天然构象Env 的 HIV 感染细胞的 ADCP 的新方法。单核细胞系（THP-1 细胞）或原代人单核细胞与表达 eGFP 的 CD4+T 细胞系在存在抗体的情况下孵育，通过流式细胞术测量 ADCP 作为 eGFP+物质的积累。通过图像捕获流式细胞术可以直观地确认单核细胞对感染 HIV 的细胞的内化。通过共聚焦显微镜还观察到细胞骨架重塑、伪足形成和吞噬作用。我们发现，有效的广谱中和抗体（bnAbs），而不是非中和抗体（nnAbs），介导对感染原发性或实验室适应的 HIV-1 的细胞的有效吞噬作用。针对 gp120 的 CD4 诱导表位的 nnAb（A32）不能使感染 HIV 的细胞发生 ADCP，但介导 gp120 包被珠粒的有效吞噬作用。相反，针对完整 Env 三聚体的 bnAb（PGT145）介导对感染 HIV 的细胞的有效 ADCP，但不促进 gp120 包被珠粒的摄取。这些结果强调了测量表达生理相关 Env 构象的 HIV 感染细胞的 ADCP 的重要性，并表明能够结合病毒颗粒上的 Env 三聚体以中和病毒感染性的大多数抗体也能够结合感染病毒的细胞表面上的 Env 来介导 ADCP。
