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. 2021 Dec 20:12:799124.
doi: 10.3389/fimmu.2021.799124. eCollection 2021.

Increased Platelet-CD4+ T Cell Aggregates Are Correlated With HIV-1 Permissiveness and CD4+ T Cell Loss

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Increased Platelet-CD4+ T Cell Aggregates Are Correlated With HIV-1 Permissiveness and CD4+ T Cell Loss

Xiao-Peng Dai et al. Front Immunol. .

Abstract

Chronic HIV-1 infection is associated with persistent inflammation, which contributes to disease progression. Platelet-T cell aggregates play a critical role in maintaining inflammation. However, the phenotypic characteristics and clinical significance of platelet-CD4+ T cell aggregates remain unclear in different HIV-infected populations. In this study, we quantified and characterized platelet-CD4+ T cell aggregates in the peripheral blood of treatment-naïve HIV-1-infected individuals (TNs), immunological responders to antiretroviral therapy (IRs), immunological non-responders to antiretroviral therapy (INRs), and healthy controls (HCs). Flow cytometry analysis and immunofluorescence microscopy showed increased platelet-CD4 + T cell aggregate formation in TNs compared to HCs during HIV-1 infection. However, the frequencies of platelet-CD4 + T cell aggregates decreased in IRs compared to TNs, but not in INRs, which have shown severe immunological dysfunction. Platelet-CD4 + T cell aggregate frequencies were positively correlated with HIV-1 viral load but negatively correlated with CD4 + T cell counts and CD4/CD8 ratios. Furthermore, we observed a higher expression of CD45RO, HIV co-receptors, HIV activation/exhaustion markers in platelet-CD4 + T cell aggregates, which was associated with HIV-1 permissiveness. High levels of caspase-1 and caspase-3, and low levels of Bcl-2 in platelet-CD4+ T cell aggregates imply the potential role in CD4+ T cell loss during HIV-1 infection. Furthermore, platelet-CD4 + T cell aggregates contained more HIV-1 gag viral protein and HIV-1 DNA than their platelet-free CD4 + T cell counterparts. The platelet-CD4 + T cell aggregate levels were positively correlated with plasma sCD163 and sCD14 levels. Our findings demonstrate that platelet-CD4 + T cell aggregate formation has typical characteristics of HIV-1 permissiveness and is related to immune activation during HIV-1 infection.

Keywords: HIV-1; T cell loss; infection; permissiveness; platelet-CD4+ T cell aggregates.

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Conflict of interest statement

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Figures

Figure 1
Figure 1
Detection of platelet-CD4 + T cell aggregates in HCs, TNs, IRs, and INRs. (A) Representative gating strategy for the flow cytometry analysis of platelet-CD4 + T cell aggregates and platelet-free CD4 + T cells from a TN patient. CD42a + CD4 + T cells represent platelet-CD4 + T cell aggregates while CD42a-CD4 + T cells represent platelet-free CD4 + T cells. (B) Representative image of circulating platelet-CD4 + T cell aggregates that came separately from HC and HIV-1-infected patients with or without ART as obtained from imaging flow cytometry. Platelets are recognized by anti-CD42a FITC (blue) while CD4 + T cells are recognized by anti-human CD3 BV510 (purple) and anti-human CD4 BV421 (green). (C) Representative images of platelet-CD4 + T cell aggregates in lymph nodes from an HIV-1-infected patient without ART as visualized via immunofluorescence microscopy. (D) Comparison of platelet-CD4 + T cell aggregate frequencies in CD4 + T cells from HCs (n=19), TNs (n=28), IRs (n=21) and INRs (n=12). (E) Median fluorescence intensity (MFI) values of CD42a were measured in all four groups. (F–H) Correlation between (F) HIV-1 viral load, (G) absolute CD4 + T cell counts, and (H) CD4/CD8 ratio with platelet-CD4 + T cell aggregate frequencies in TNs (n=28). *P < 0.05, **P ≤ 0.01, ***P ≤ 0.001, ****P ≤ 0.0001.
Figure 2
Figure 2
Phenotype of platelet-CD4 + T cell aggregates in HCs, TNs, IRs, and INRs. The phenotypic characteristics of platelet-CD4 + T cell aggregates were analyzed, including platelet activation, CD45RO expression, HIV-1 co-receptors, immune checkpoint receptors, activation markers, and pyroptosis/apoptosis markers, as well as anti-apoptotic proteins from HCs, TNs, IRs, and INRs. Platelet-CD4+ T cell aggregates and platelet-free CD4+ T cells were compared with regards to their expression of CD62P (A) [HCs (n=19), TNs (n=20), IRs (n=16), and INRs (n=11)], CD45RO (B) [HCs (n=11), TNs (n=18), IRs (n=14), and INRs (n=11)], CCR5 (C) [HCs (n=17), TNs (n=18), IRs (n=21), and INRs (n=12)], CXCR4 (D) [HCs (n=13), TNs (n=13), IRs (n=17), and INRs (n=12)], CD38/HLA-DR (E) [HCs (n=11), TNs (n=13), IRs (n=17), and INRs (n=11)], PD-1 (F) [HCs (n=8), TNs (n=17), IRs (n=17), and INRs (n=11)], caspase-1 (G) [HCs (n=16), TNs (n=15), IRs (n=15), and INRs (n=10)], caspase-3 (H) [HCs (n=16), TNs (n=15), IRs (n=16), and INRs (n=10)], and Bcl-2 (I) [HCs (n=10), TNs (n=16), IRs (n=16) and INRs (n=11)]. *P < 0.05, **P ≤ 0.01, ***P ≤ 0.001, ****P ≤ 0.0001.
Figure 3
Figure 3
Elevated levels of HIV-1 p24 and DNA in platelet-CD4+ T cell aggregates. (A) Representative images of p24 expression in platelet-CD4+ T cell aggregates via imaging flow cytometry from a treatment-naïve patient. Top row: p24 visualized only in the platelets of platelet-CD4+ T cell aggregates. Middle row: p24 visualized only in the CD4+ T cells. Bottom row: p24 visualized in both platelets and CD4+ T cells. (B) Representative gating strategy for p24 expression in platelet-CD4+ T cell aggregates and their counterparts from TNs and HCs. (C) P24 expression in platelet-CD4+ T cell aggregates and their counterparts in different groups of treatment-naïve HIV-1-infected subjects. CD4+ T cell counts>350 cells/μL (n=9), 200≤CD4+ T cell counts ≤ 350 cells/μL (n=11), CD4+ T cell counts<200 cells/μL (n=6). (D) Median fluorescence intensity (MFI) values of p24 were compared between platelet-CD4+ T cell aggregates and their counterparts in TNs (n=26). (E) Correlation between HIV-1 viral load and platelet-CD4+ T cell aggregates expressing p24 in TNs (n=26). (F) Platelet-CD4+ T cell aggregates and their counterparts were isolated from PBMCs of IRs (n=7); HIV-1 DNA was detected in these two cell fractions. *P < 0.05, **P ≤ 0.01, ***P ≤ 0.001.
Figure 4
Figure 4
An increased frequency of platelet-CD4 + T cell aggregates is positively correlated with sCD14 and sCD163 during HIV-1 infection. (A) Serum level of sCD14 from HCs (n=19), TNs (n=28), IRs (n=21) and INRs (n=12). (B) Serum level of sCD163 from HCs (n=19), TNs (n=28), IRs (n=21) and INRs (n=12). (E) Frequencies of CD38+HLA-DR+CD4+T cell in CD4+ T cells from HCs (n=11), TNs (n=13), IRs (n=17) and INRs (n=11). Correlation between (C) plasma sCD14 levels (n=28), (D) plasma sCD163 levels (n=28), and (F) CD38+HLA-DR+CD4+ T cells (n=13) and the frequencies of platelet-CD4+ T cell aggregates in CD4+ T cells. *P < 0.05, **P ≤ 0.01, ***P ≤ 0.001, ****P ≤ 0.0001.

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