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Background:  Natural CD4⁺CD25^+hi T regulatory cells (T_R) can suppress HIV-specific T-cell immune responses in vitro. However, their in vivo dynamics during different stages of HIV infection are unknown.

Methods: Subjects included 47 newly infected (NI; <6months) and 13 chronically infected (CI) adults with HIV-1. Flow cytometry was performed using frozen peripheral blood monocuclear cells (PBMC). T cells were sorted into T_R (defined as CD3⁺CD4⁺CD25^+hiCD45RO⁺) and non-T_R CD4⁺T subsets. Foxp3 and HIV RNA levels in these subsets were quantified by real-time polymerase chain reaction (PCR) and normalized using gapdh Paired t test, analysis of variance (ANOVA), and linear regression methods.

Results: CD25^+hiCD4⁺T cells exhibited a 400-fold higher level of Foxp3 gene expression compared to CD25^-CD4⁺ cells. Among CD25^+hiCD4⁺T cells, the CD45RO⁺ subset constituted about 66%, and contained Foxp3 at a 5-fold higher level compared with the CD45RO^- subset, confirming that Foxp3-expressing regulatory T cells can be phenotypically characterized as CD25^+hiCD45RO⁺CD4⁺T cells. NI subjects had a significantly higher percentage of T_R within lymphocytes (1.8% vs 0.9%; p = 0.004) as well as higher absolute T_R cell counts (39 vs 22 cells/mm³, p = 0.005) compared to CI. T_R in HIV-uninfected subjects (n = 18) was 1.4%, which was lower compared to NI subjects (p=0.02) but higher compared to CI subjects (p = 0.08). The percentage of T_R within CD4⁺T cells remained constant (acute 7.7; chronic 7.1; HIV-uninfected 7.6). No correlation of T_R with baseline plasma HIV RNA was seen. Longitudinal analyses revealed significant increase of T_R (expressed as percentage of lymphocytes) from 1.2% before treatment to 1.5% at 24 weeks and 2.0% at 48 weeks on highly active antiretroviral therapy (HAART) (p = 0.051) However, the slope of CD4⁺T-cell count increase during therapy-related immune reconstitution was significantly steeper than that of T_R increase during the same period (p < 0.0001), suggesting that T_R-cell regeneration rate was slower compared to CD4⁺T cells. In 2 subjects, T_R had a 6.9 and 31.2-fold higher level of HIV RNA compared to non-T_R CD4⁺T cells.

Conclusions: CD4⁺CD25^+hi T regulatory cells are expanded during acute HIV infection and decrease over time in untreated chronic HIV infection, presenting a possible target for infection with HIV. T_R numbers increase during HAART; however the slower recovery of T_R may contribute to persisting defects in immune responses even when CD4⁺ T cell numbers are restored. Further studies are underway to evaluate T_R infectivity and T_R-cell function during HAART.

Keywords: T regulatory cells; HIV-1; foxp3