Peripheral CD4⁺CD25⁺ T cells from HIV-infected patients suppress CD4 T-cell proliferation in response to recall antigens and HIV proteins. (A) Unfractionated CD4⁺ T cells (□) or purified CD4⁺CD25^– T cells (▪) were incubated for 5 days with 5 μg/mL tuberculin (PPD), 5 μg/mL p24 protein, or 1:50 dilution of CMV antigen before adding 0.5 μCi (0.0185 MBq) [³H] thymidine for the last 16 hours of culture. Results are expressed as median values of the stimulation indexes for 8 patients studied (*P < .05). The median (ranges) of [³H] thymidine incorporation in control cultures (medium alone) were 462 (140-824), 240 (130-377), and 136 (63-175) for unfractionated CD4⁺ cells, CD4⁺CD25^– cells, and CD4⁺CD25⁺ cells, respectively. (B) Coculture of CD4⁺CD25^– with CD4⁺CD25⁺ T lymphocytes from HIV-infected patients results in a dose-dependent suppression of antigen-specific proliferation. Purified CD4⁺CD25^– T cells were cultured for 5 days in the presence of 5 μg/mL purified tuberculin (♦) or 5 μg/mL p24 protein (⋄) and varying numbers of purified CD4⁺CD25⁺ lymphocytes. Percentage of inhibition was calculated as described in “Proliferation and suppression assays.” Results are expressed as mean (± SD) percentage of inhibition of proliferation for 5 patients studied.