Preserved function of regulatory T cells in chronic HIV-1 infection despite decreased numbers in blood and tissue

Abstract

Regulatory T cells (Tregs) are potent immune modulators, but their role in human immunodeficiency virus type 1 (HIV-1) pathogenesis remains poorly understood. We performed a detailed analysis of the frequency and function of Tregs in a large cohort of HIV-1-infected individuals and HIV-1 negative controls. While HIV "elite controllers" and uninfected individuals had similar Treg numbers and frequencies, the absolute numbers of Tregs declined in blood and gut-associated lymphoid tissue in patients with chronic progressive HIV-1 infection. Despite quantitative changes in Tregs, HIV-1 infection was not associated with an impairment of ex vivo suppressive function of flow-sorted Tregs in both HIV controllers and untreated chronic progressors.

Figure 1.

Regulatory T-cell (Treg) frequencies in peripheral blood and colonic lymphoid tissue from individuals infected with human immunodeficiency virus type 1 (HIV-1) at different stages of HIV-minus-one infection. A, Flow cytometry gating strategy: Gates are set to include CD3⁺ T lymphocytes and exclude dead cells (viability marker⁺), monocytes (CD14⁺), and B cells (CD19⁺). CD4⁺ Tregs were defined by the coexpression of CD25 and forkhead box P3 (FOXP3) using appropriate fluorescence-minus-one controls. Top right panels show representative examples of CD4⁺ Tregs in an HIV-1 elite controller and an untreated chronic HIV-1 progressor. B, CD25⁺FOXP3⁺ Treg frequencies (%Treg of CD3⁺CD4⁺ T cells) are shown. C, Absolute Treg numbers were determined and compared between the study groups: 21 HIV-1–uninfected controls (HIV⁻), 26 HIV-1 elite controllers (EC), 30 antiretroviral therapy–naive individuals with chronic untreated HIV-1 infection (CU), 10 HIV-1 positive untreated individuals with primary HIV-1 infection (AU), and 20 HIV-1 positive individuals treated with highly active antiretroviral therapy (HAART) (CT). D, Representative TissueQuest scattergrams for a healthy HIV-negative donor (upper left panel) and an untreated chronic progressor (CU) (lower left panel) with examples of lamina propria sections (upper and lower middle panels) and lymphoid follicle sections (upper and lower right panels). Images show CD4⁺ Treg staining acquired by confocal microscopy (×200 magnification). Blue represents the nucleus (4′, 6-diamidino-2-phenylindole [DAPI]); green, CD4; red, FOXP3. The frequency of FOXP3⁺ Tregs is measured among DAPI⁺CD4⁺ cells using TissueQuest software as represented on the left panels. E, F, The frequencies of CD4⁺ T cells among total cells (E) and of FOXP3⁺ Tregs among CD4⁺ T cells (F) were compared in the lamina propria and lymphoid follicles (not shown) of 12 healthy donors and 10 untreated chronic progressors.

Figure 2.

Functional assessment of CD4⁺ regulatory T cells (Tregs) isolated from individuals infected with human immunodeficiency virus type 1 (HIV-1) and healthy control subjects. A, Representative dot plot showing the sorting strategy for the CD3⁺CD4⁺ T-cell subpopulations isolated for suppression assays. Regulatory T cells are defined as CD25⁺CD127^low, and responder T cells as CD25⁻CD127⁺. B, Correlation of the frequency of CD4⁺CD25⁺CD127^low/− and CD4⁺CD25⁺FOXP3⁺ T-cell populations within the same individuals (11 HIV-1–negative individuals, 2 treated with highly active antiretroviral therapy (HAART), 7 untreated chronic progressors, and 10 elite controllers). C, Proliferation of sorted T responder cells stimulated by microbeads in the presence of different ratios of autologous Tregs in HIV-negative (solid green line), HIV-1 elite controllers (dotted red line), and chronic untreated donors (solid black line). Three individuals were studied per group. D, Representative histograms of proliferation of the carboxyfluorescein succinimidyl ester (CFSE)–loaded responder T cells in the presence of different ratios of Tregs as analyzed with the FlowJo software proliferation platform.