Cross-Talk between Programmed Death-1 and Suppressor of Cytokine Signaling-1 in Inhibition of IL-12 Production by Monocytes/Macrophages in Hepatitis C Virus Infection

Abstract

Hepatitis C virus (HCV) dysregulates innate immune responses and induces persistent viral infection. We previously demonstrated that HCV core protein impairs IL-12 expression by monocytes/macrophages (M/MΦs) through interaction with a complement receptor gC1qR. Because HCV core-mediated lymphocyte dysregulation occurs through the negative immunomodulators programmed death-1 (PD-1) and suppressor of cytokine signaling-1 (SOCS-1), the aim of this study was to examine their role in HCV core-mediated IL-12 suppression in M/MΦs. We analyzed TLR-stimulated, primary CD14+ M/MΦs from chronically HCV-infected and healthy subjects or the THP-1 cell line for PD-1, SOCS-1, and IL-12 expression following HCV core treatment. M/MΦs from HCV-infected subjects at baseline exhibited comparatively increased PD-1 expression that significantly correlated with the degree of IL-12 inhibition. M/MΦs isolated from healthy and HCV-infected individuals and treated with HCV core protein displayed increased PD-1 and SOCS-1 expression and decreased IL-12 expression, an effect that was also observed in cells treated with gC1qR’s ligand, C1q. Blocking gC1qR rescued HCV core-induced PD-1 upregulation and IL-12 suppression, whereas blocking PD-1 signaling enhanced IL-12 production and decreased the expression of SOCS-1 induced by HCV core. Conversely, silencing SOCS-1 expression using small interfering RNAs increased IL-12 expression and inhibited PD-1 upregulation. PD-1 and SOCS-1 were found to associate by coimmunoprecipitation studies, and blocking PD-1 or silencing SOCS-1 in M/MΦ led to activation of STAT-1 during TLR-stimulated IL-12 production. These data suggested that HCV core/gC1qR engagement on M/MΦs triggers the expression of PD-1 and SOCS-1, which can associate to deliver negative signaling to TLR-mediated pathways controlling expression of IL-12, a key cytokine linking innate and adaptive immunity.

KLRG1 Negatively Regulates Natural Killer Cell Functions through the Akt Pathway in Individuals with Chronic Hepatitis C Virus Infection

“…These results are in agreement with our previous observations that another negative signaling molecule, T cell immunoglobulin and mucin domain protein 3 (Tim-3), is upregulated on NK cells to dampen their functions, with inhibited activation (CD69), proliferation (Ki69), degranulation (CD107a), and killing activity (granzyme B) following HCV infection (data not shown). This in turn interplays with monocytes and T cells (including Th17 and Foxp3 ϩ Tregs) in immune dysregulation that contribute to viral persistence (42)(43)(48)(49). This also supports the notion that HCV may utilize KLRG1-an intrinsic mercenary of cell feedback regulation-for the purpose of virus persistence and thus facilitate chronic infection.…”

“…Additionally, the driving force for KLRG1 upregulation during HCV infection remains to be determined. To further elucidate the role of HCV in regulation of KLRG1 expression, we employed a newly established cell culture system by transfecting Huh-7 hepatocytes with the HCV JFH-1 strain in vitro to mimic the in vivo setting of early HCV infection (42,43). To this end, PBMCs or purified NKs from HS were cocultured with HCV-transfected or untransfected Huh-7 hepatocytes for 48 h, followed by flow cytometric analysis for KLRG1 expression and NK functions.…”

“…Transfection of Huh-7 hepatocytes (kindly provided by T. J. Liang, Liver Section, NIH NIDDK) with HCV JFH-1 strain (kindly provided by T. Wakita) was carried out as described previously (42,43). Prior to the coculture experiment, HCV-transfected or untransfected Huh-7 hepatocytes were serum starved for 18 h, then activated with rhIFN-␥ (0.1 g/ml; R&D Systems) for 48 h. Activated hepatocytes were removed from plates with 0.05% trypsin-EDTA and then plated at 5 ϫ 10 5 cells/well in a 12-well plate.…”

“…Flow cytometry. Procedures for detection of cell surface markers and intracellular cytokine staining were performed essentially as described previously (42,43). Briefly, PBMCs (0.2 ϫ 10 6 per well in a 96-well plate) were stimulated with 10 ng/ml recombinant human interleukin-12 (rhIL-12; eBioscience, San Diego, CA) for 18 h, followed by 1 g/ml Brefeldin A (BioLegend, San Diego, CA) 4 h prior to harvesting the cells, thus forbidding cytokine secretion.…”

Upregulation of IL-23 Expression in Patients with Chronic Hepatitis B Is Mediated by the HBx/ERK/NF-κB Pathway

“…In addition, multiorgan inflammation and premature death are observed in transgenic mice overexpressing the IL-23 p19 subunit (18). Previous studies have found that IL-12 family members, such as IL-12 and IL-27, are involved in chronic inflammation and liver damage from HBV and hepatitis C virus infections (19)(20)(21)(22). However, the regulatory mechanism of IL-23 in chronic HBV infection remains largely unknown.…”