Toll-like receptor-activated primary human hepatocytes produce inflammatory cytokines and different types of interferon, resulting in HCV suppression in a co-culture model

Aims: The function of the hepatic innate immune system and its role in the defence against Hepatitis C Virus infection are not well understood. Recent publications suggested that type-III interferons (IFN; Interleukin-28A, -28B and -29) play a crucial role in the host response against HCV. Aim of this study was to investigate the Toll-like receptor (TLR) signaling in primary human hepatocytes and the capacity of these cells to produce type-III IFNs and their ability to control HCV replication.

Methods: Human liver samples were obtained after tumor resection (n=16) or liver transplantation (n=14). Human hepatocytes (n=30) were isolated after perfusion and digestion of the liver. Cells were stimulated with TLR1–9 ligands for 6h to 24h, expression of tumor necrosis factor α (TNF-α), Interleukin 6 (IL-6), IL-10, IFN-α, -β, -γ,-λ and ISGs were determined by qRT-PCR or ELISA.

Results: In human hepatocytes stimulation with TLR1–9 ligands, except TLR9, led to cell-type specific induction of pro-inflammatory (TNF-α, IL-6) and anti-inflammatory cytokines (IL-10). In contrast, a slight expression of type-I, -II (IFN-α, -β, -γ) and a high expression of type-III interferons (IL-28A, IL-28B, IL-29) could only be induced after TLR3 stimulation, resulting in enhanced expression of ISGs (ISG15, IFI-T1, RSAD2, MXA). Therefore, only supernatants from TLR3-activated hepatocytes suppressed HCV replication during co-culture with hepatoma cells harbouring a subgenomic HCV replicon. The use of neutralising antibodies against type-I and -II IFNs did not alter this suppressive effect, which therefore might be mediated by type-III IFN. TLR3 ligand Poly I:C-induced IFN and ISG expression is enhanced in hepatocytes isolated from HCV-infected patients. The different types of surgery, fibrosis stage as well as serum levels of transaminase did not correlate with TLR signaling and antiviral activity in these cells.

Conclusions: Primary isolated human hepatocytes respond to TLR ligands by production of pro-inflammatory (TNF-α and IL-6) anti-inflammartory (IL-10) cytokines as well as type-I, -II and -III IFNs. TLR3-activated hepatocytes led to suppression of HCV replication in a type-III IFN-dependent manner. However, gene expression of IFNs and ISGs is higher in patients infected with HCV.