386 Differential Regulation of IL-7 Receptor (CD127) by TNFalpha and IL-7: Implications for HIV Immunopathogenesis A. Komsic*, C. D. Young, J. B. Angel Ottawa Hlth Res Inst, Ottawa, Canada
Background: IL-7 and its signaling via the IL-7 receptor (IL-7R/CD127) are required for normal CTL activity. Our previous observation that down-regulation of this receptor is observed on CD8+ T-cells of HIV infected individuals and is restored with effective therapy suggests that this may contribute to impaired CTL activity during HIV infection. Initial studies indicated that TNFalpha and IL-7 decrease CD127 expression whereas IL-10 and HIV gp120 do not. Further elucidating the mechanism(s) of CD127 down-regulation will provide further insight into the immunopathogenesis of HIV disease.
Methods: Magnetic bead isolated CD8+ T-cells from HIV seronegative donors were incubated with TNFalpha (0 ng/ml to 100 ng/ml) or IL-7 (0 pg/ml to 10,000 pg/ml) for 0-96 hrs for flow cytometric evaluation of CD127 surface expression, or 0-24 hrs for evaluation of CD127 RNA expression by both a semi-quantitative and a quantitative (competitive) RT-PCR. To determine if the effects of TNFalpha are direct or if they require de novo protein synthesis, experiments were performed with cycloheximide, an inhibitor of gene translation. To determine if changes in CD127 RNA were related to potential effects on RNA stability, experiments were performed with the transcriptional inhibitor actinomycin D.
Results: TNFalpha induced a time- and dose-dependent decrease in the surface expression of CD127 and the accumulation of CD127 RNA. These changes were limited to the naïve (CD45RAsup>+) subset of CD8sup>+ T-cells. IL-7 (up to 1000 pg/ml) caused a transient decrease in CD127 surface expression, whereas with higher concentrations, this response was sustained. IL-7 has no effect on CD127 RNA levels. The inhibitory activity of TNFalpha on CD127 RNA accumulation was partially reversed by cycloheximide. CD127 RNA levels at 24 hrs were decreased in the presence of both TNFalpha and actinomycin D compared to actinomycin D alone, suggesting that CD127 RNA degradation is enhanced by TNFalpha.
Conclusion: IL-7R expression is downregulated by TNFalpha and IL-7, 2 cytokines that are up-regulated during HIV infection. The mechanisms of action of these 2 cytokines appear to be distinct, indicating a complex regulation of CD127 expression and, therefore, IL-7 signaling during HIV infection.
