Background:  Interleukin-21 (IL-21) plays important roles in regulation of T, B, and natural killer (NK) cells. We hypothesized that IL-21 may elicit cellular anti-HIV immune responses by influencing the cytotoxic potential of CD8 T cells via regulation of perforin (PFN) expression.

Methods:  Fresh peripheral blood mononuclear cells of healthy donors (n = 5) and HIV-infected individuals (n = 6) on potent antiretroviral therapy were cultured with IL-21 or IL-15 alone, anti-CD3 monoclonal antibody or HIV-gag peptide pool or combinations thereof for 5 hours. Intracellular PFN and cell surface CD107a expression in CD8 T cells and maturation subsets, defined by CD45RA and CD62L expression, were examined by flow cytometry. Unpaired t test was used for statistical analyses.  

Results:  Baseline PFN values were variable in HIV-infected subjects (19.2 to 40.5%) and controls (7.7 to 45.7%). In HIV-infected donors, IL-21 induced up-regulation of PFN expression in CD8 T cells after 5 hours’ stimulation as compared to cells cultured in medium (percentage of PFN+CD8 T cells 31.1±5.8 vs 16±2.7%, respectively; p <0.05), without inducing other T-cell activation markers. Real-time polymerase chain reaction (RT-PCR) analysis confirmed induction of PFN expression at the mRNA level in CD3 T cells. IL-15 also up-regulated PFN in CD8 T cells but the effect was significantly greater with IL-21 (1.5- ±0.05-fold increase vs 1.9- ±0.02-fold increase, respectively; p <0.001). In CD8 T-cell memory subsets, increased PFN expression was observed in both, central memory (unstimulated, 5.5±1.5% vs IL-21-cultured, 16.6±4%; p <0.05) and effector memory (unstimulated, 21.1±2.6% vs IL-21-cultured, 45.5±7.8%; p <0.05) cells from HIV-infected individuals. In healthy subjects PFN expression in CD8 T cells showed a similar trend after culture with IL-21, but the increase was not significant. HIV-gag stimulated patient CD8 T cells had elevated PFN expression in the presence of IL-21, which was further enhanced with IL-21 + IL-15 stimulation. Basal expression of surface CD107a in CD8 T cells was equivalent in patients and controls (1.8±0.3% and 0.55±0.2%, respectively) and was not augmented by IL-21 or HIV gag stimulation, but increased significantly by anti-CD3 monoclonal antibody activation (patients 6.7±2%; p <0.05; and controls, 8.6±2%; p <0.01), concordant with loss of PFN expression.

Conclusions:  These data suggest a potential role for IL-21 in boosting anti-viral immunity by building up intracellular perforin stores, without inducing activation or degranulation of memory CD8 T cells.