Background:  Recent studies indicate that the incomplete differentiation of HIV-specific CTL is associated with the lack of virologic control in chronic HIV infection, and that CTL proliferation potential is positively associated with protective immunity in both mice and humans. We hypothesize that the incomplete differentiation of HIV-specific CTL is due to the reduced proliferation potential of these cells.

Methods:  To test this hypothesis, we examined conditions conducive to proliferation and differentiation of HIV-specific CTL in 6 HIV-infected individuals and CMV-specific CTL in 2 HIV-uninfected individuals. PBMC were CFSE labeled and stimulated with either PHA, HIV peptides, or CMV peptides. Antibodies to CD28 and CD49d were added to peptide containing cultures. Cytokines including IL2, IL7, IL15, and IL23 were added to some peptide-stimulation experiments. The proliferation and differentiation of CD8⁺ T cells were assessed 5 days later using a combination of antibodies including CD3, CD4, CD8, CD27, CD28, CD45RO and CCR7, and analyzed by FACS. The percentages of proliferating CD8⁺ T cells (Tp), and the numbers of maximum cell division (Dm) achieved were evaluated.

Results:  In both HIV-infected and -uninfected subjects, PHA stimulated a Tp of 90 to 95% and a Dm of 5 to 8. PHA stimulation led to a 2-fold decrease in the number of surface CD28 and CCR7 molecules, and a 2-fold increase in CD45RO molecules. Bi-phasic changes in CD27 expression were observed, increasing during cell divisions 1 to 3, but decreasing during divisions 4 to 7. CMV peptides stimulated a Tp of 3-5% and a Dm of 6 in HIV-uninfected subjects, and modulated the CCR7, CD45RO and CD27 markers similar to PHA.  In contrast, HIV peptide pools elicited a Tp of only 1 to 2% and a Dm of only 4 in just 1 of 6 infected subjects. In this responder, supplementation with hemopoietic cytokines IL2 and IL15 led to a 6- to 10-fold increase in the Tp of HIV peptide-specific CTL, and extended the Dm to 6, while IL7 and IL23 had no effect.

Conclusions:  Upon PHA stimulation, CD8⁺ T cells from HIV-infected individuals can proliferate, and progressively differentiate to a more mature phenotype. When stimulated with viral peptides, however, conditions optimal for the proliferation of CMV-specific CTL failed to induce proliferation of HIV-specific CTL. The addition of hemopoietic cytokines, which may be lacking in patients with HIV infection, partially corrected the defective responses of HIV-specific CTL.

Keywords: CTL; Differentiation; Proliferation