SINGLE-CELL ANALYSIS OF LYMPHOKINE IMBALANCE IN ASYMPTOMATIC HIV-1 INFECTION - EVIDENCE FOR A MAJOR ALTERATION WITHIN THE CD8(-CELL SUBSET() T)

In this study we investigated at single-cell level by how cytometry th e potential of T cell cytokine production in asymptomatic HIV-1-infect ed subjects with > 200 CD4 counts and possible correlation with T help er cell depletion and viral load. Mitogen-stimulated peripheral blood mononuclear cells from 32 HIV-1(+) patients and 16 healthy subjects we re intracytoplasmically stained for IL-2, interferon-gamma (IFN-gamma) , IL-4 or IL-10, and the frequency of cytokine-producing cells was ass essed in total T cells, CD4, CD8 and CD45RO subsets as well as in CD69 (+) CD3(+) gated lymphocytes. HIV-1(+) patients, irrespective of their degree of CD4 depletion, exhibited a major increase in IFN-gamma(+) C D8 T cells, largely due to CD28(-) cells, as well as a decrease in the capacity of CD8 T cells to produce IL-2. Patients with > 500 CD4 coun ts showed a diminished frequency of IL-4 expression in CD4 T cells and a negative correlation was found between this parameter and the ex vi vo CD4 counts in the 32 patients. Analysis of patients stratified acco rding to viral load revealed a significantly higher proportion of IL-2 -producing CD4 cells in the group with < 5000 RNA copies/ml. In short, using single-cell analysis and an antigen-presenting cell-independent stimulus, we have not been able to find any significant cytokine imba lances in the CD4 subset, suggesting that the well described T helper defects are not due to intrinsic alterations in the potential of CD4 T cells to produce cytokines. On the other hand, the major disturbances in the CD8 T lymphocytes agree with the marked activation and possibl e replicative senescence of CD8 T cells and emphasize the role of this subset in HIV immunopathogenesis.