Background
Natural killer (NK) cells provide immune defense against virus infection via surveillance and recognition of virally infected cells. In people living with HIV (PWH) NK cell function has been associated with viral control and is being targeted for immunotherapeutic strategies.
Methods
To determine whether NK function is impaired in PWH, in vitro NK functional assays were performed in longitudinal samples from HIV-Exposed Infected (HEI, n=17) and Uninfected (HEU, (n=9) children from Mozambique. NK were enriched from PBMC and co-cultured with CellTraceTM labeled K562, HIV-infected HUT78/SF2 (HUT) and NK-resistant CEM.NKR (CEM) cell lines for 6 hours at 4:1 ratio. Death of target cells (viability stain), cytolytic proteins (CD107a, Perforin, GranzymeB, FasL) and cytokine secretion (IFNg, TNF, MIP1b) were assessed at pre-ART (1-2m) and post-ART (12m) by flow cytometry. Expression of activating and inhibitory receptors were evaluated on NK before and after co-culture (NKG2A/C/D, NKp30/46, KLRG1, TIGIT). Adults with and without HIV (n=10 each) at a single timepoint were assessed for comparison. Mixed-effects modelling was used to determine predictors of NK killing.
Results
Overall, NK exhibited increased killing of K562 and HUT targets compared to CEM but the magnitudes were not different when comparing HEI and HEU. Paired analysis showed that HEU NK increased killing from 1m to 12m whereas HEI stayed the same or decreased. At 12m only, plasma virus load (VL) had a negative linear relationship with HUT killing (R2=0.42, p=0.016) and this was not observed with K562. Expression of the inhibitory receptor, KLRG1 on NK prior to co-culture was associated with reduced killing of K562 and HUT. After co-culture, the expression of CD107a on NK, a marker of degranulation, was highly associated with K562 target cell death but showed no relationship with HUT killing. Meanwhile, expression of the death receptor, FasL, which was present on CD56dimCD16+ NK at 5-30% was positively associated with HUT killing but showed no relationship with K562 killing. Staining of target cells showed that HUT expressed higher levels of surface Fas receptor than K562 or CEM. FasL on NK cells from adults also had a positive association with HUT killing confirming this was not a unique feature of pediatric NK cells.
Conclusions
These results suggest that chronic HIV leads to impaired NK cytolytic function against HIV infected CD4 T cells and point to a role for Fas/FasL induced cell death in viral control mediated by NK cells.