BACKGROUND and AIMS: Interferon-gamma (IFN-gamma), a cytokine produced by activated natural killer cell (NK) and T lymphocytes, is an important regulator of innate and adaptive immunity during hepatitis C virus (HCV) infection. However, the cellular sources and mechanisms of IFN-gamma induction in HCV-infection are not fully understood.

METHODS: We cultured normal human peripheral blood mononuclear cells (PBMCs) with different populations of immune cells and JFH-1 HCV-infected Huh7.5 (JFH-1/Huh7.5) cells.

RESULTS: We found that PBMCs produced large amounts of IFN-gamma after co-culture with JFH-1/Huh7.5 cells. Using intracellular cytokine staining we confirmed that NK cells and NKT cells (to a lesser extent) were the major IFN-gamma producers within PBMCs. Purified NK/NKT cells did not produce IFN-gamma in response to JFH-1/Huh7.5 cells and depletion of accessory (HLA-DR+) cells prevented IFN-gamma induction in PBMCs. Through selective cell depletion of dendritic cells or monocytes from PBMCs, we determined that plasmacytoid dendritic cells (pDCs) were indispensable for NK-IFN-gamma induction and the presence of monocytes was needed for maximal NK-IFN-gamma induction. We further revealed that NK-IFN-gamma induction depended on pDC-derived IFN-alpha while other IFN-gamma inducing cytokines, IL-12 and IL-18, played minimal roles. Close contact between JFH-1/Huh7.5 cells and NK cells was required for IFN-gamma production and monocyte-derived IL-15, significantly augmented IFN-gamma induction.

CONCLUSIONS: We discovered a novel mechanism where NK cells interact with pDCs and monocytes, efficiently producing IFN-gamma in response to HCV-infected cells. This indicates that co-operation between NK cells and accessory cells is critical for IFN-gamma production and regulators of immunity during HCV infection.