Background: CMV is associated with persistent inflammation in HIV-infected persons. Here, we studied the effect of HIV expanded myeloid derived suppressor cells (MDSCs) in controlling CMV specific inflammation.
Methods: PBMCs from HIV–/CMV-seropositive (CMV+) donors were cultured in presence of heat inactivated HIV. After 5 days, CD11b+CD33+CD14+HLA DRhi (DRhi monocytes) and CD11b+CD33+CD14+HLA DR-/lo (MDSCs) cell subsets were sorted by flow cytometry. B7H4 (a negative regulator of T cell function) was silenced using siRNA and cultured with/without autologous PBMCs in presence/absence of CMV pp65 peptides (pp65; 1µg/ml). In some experiments, PBMCs were cultured with HIV/pp65 in presence/absence of neutralizing anti-IL-27 antibody. Enumeration of B7H4 on MDSCs, regulatory T-cells, intracellular IFNγ, activated forms phospho(p)-Zap70 and p-Akt were determined by flow cytometry; IFNγ and IL-27 were quantified by ELISA. Data were analyzed using two-tailed, paired Student’s t test.
Results: MDSCs cultured with autologous PBMCs exposed to pp65 caused a decrease in IFNγ production vs. controls or DRhi monocytes (p=0.002). IFNγ release was restored when MDSCs were transfected with B7H4 siRNA and cultured with PBMCs in presence of pp65 (p=0.02). MDSCs cultured with PBMCs did not alter pp65 induced activation of proximal T-cell signaling molecule Zap70 but decreased activation of Akt; this was restored when B7H4 was knocked down in MDSCs cultured with PBMCs. Culture of MDSCs with pp65 produced more IL-27 vs. control MDSCs and DRhi monocytes (p=0.05). Culture of CMV+ PBMCs with pp65 increased the frequency of CD4+IFNγ+ cells and release of IFNγ in supernatants vs. controls (p=0.04). IFNγ was further increased when PBMCs were cultured in presence of anti-IL-27 and stimulated with pp65; CMV– PBMCs did not produce IFNγ when treated with pp65. Furthermore, culture of CMV+ PBMCs with pp65 led to expansion of FoxP3+ Tregs vs. controls (p=0.02) and CMV– (p=0.003); addition of anti-IL-27 had no effect on Treg expansion. Finally, HIV expanded MDSCs had increased expression of B7H4 when compared to DRhi monocytes (p=0.03) which was inhibited in the presence of anti-IL-27 neutralizing antibody (p=0.05).
Conclusions: These findings suggest that IL-27 down regulates IFNγ during CMV infection. IL-27 induces B7H4 expression on HIV MDSCs which controls CMV induced T-cell IFNγ production by inhibiting p-Akt. IL-27 and B7H4 provide new therapeutic targets to control inflammation during HIV infection.