Background:
Natural killer (NK) cells provide rapid and robust responses against viral infections. Multiple studies have shown that NK cells are also capable of antigen-specific recall to eliminate infected cells. In nonhuman primates (NHP), antigen-specific memory-like NK cell can be designed by MHC-E dependent response. During SIV infection, the stabilized MHC-E molecule may induce a potent pathway to restrict the virus via antigen-specific memory NK responses.
Methods:
To study antigen-specific NK cells in NHP, we developed a novel in vitro method to clonally expand single rhesus macaque (RM) NK cells (rhNKCL) from SIV naive or chronically infected RM. We identified SIV-derived peptides by in silico analysis and confirmed their MHC-E binding using a peptide stabilization assay on K562 cells stably expressing MHC-E (HLA-E and Mamu-E) and primary RM cells. We characterized the effector functions of NK and rhNKCL cells via intracellular staining and cytotoxicity assays in combination with K562 cell lines or autologous BLCL loaded with SIV-derived peptides.
Results:
Ten SIV-derived peptides showed MHC-E stabilization on K562 MHC-E+ (HLA-E and Mamu-E) cells and RM primary cells. Enriched NK cells exhibited an increase in IFNgamma, TNFalpha, and Ki67 during SIV acute infection against SIV-derived peptides, and a decrease in these responses in chronic infection. Interestingly, NK cells from RM with chronic SIV infection showed higher cytotoxic responses compared to NK cells from SIV-naïve RM against K562 Mamu-E+ cells loaded with SIV-derived peptides. To further investigate the existence of antigen-specific NK cells, we established a novel procedure for single-cell clonal expansion of NHP NK cells, which maintain a conserved NHP NK cell phenotype of CD3- NKG2A/C+ CD16+ CD56+ following expansion. Autologous peptide-loaded BLCL elicited a broad-spectrum of rhNKCL cytotoxic responses, with higher response for SIV ENV-derived peptide.
Conclusions:
In summary, these findings elucidate antigen-specific NK responses during SIVmac infection dependent on MHC-E-mediated SIV-derived peptide presentation. Also, we introduced a ground-breaking method for the clonal expansion of single NK cells in RM. This study may provide valuable insights into antigen-specific NK cells, including the establishment of epigenetic, metabolic, transcriptional, and phenotypic profiles. Such insights could be leveraged to enhance antiviral NK cell activity in future vaccine strategies and cell therapies.