Recognition of hepatitis C virus RNA by toll like receptors 7 and 8 : implications for the initiation of innate immune response

Hepatitis C virus (HCV), a positive single stranded RNA (ssRNA) virus that replicates in the liver, infects 200 million people worldwide, with approximately 80% of infected individuals ultimately suffering from chronic HCV infection. Antiviral therapies, including interferon and ribavirin, have improved considerably in recent years, but are effective in only about one-half of those treated, and are associated with significant side effects and toxicity.  Innate immune defenses are essential to control viral infection; the innate response is activated through recognition of viral macromolecular motifs known as pathogen-associated molecular patterns (PAMPs) that are recognized by a multitude of Pathogen recognition receptors (PRRs). Although immune activation induced by HCV RNA or proteins has been extensively studied, the detection of HCV by the innate immune system remains poorly understood. Despite activation of the immune response early after HCV infection in vivo, the persistent increase inpatient HCV viral load suggests the failure of the immune response to control HCV infection. A better understanding of the mechanisms of immune activation induced by HCV is crucial for development of effective strategies for HCV treatment. Here we demonstrate in primary cell models that the HCV genome contained GU-rich RNA sequences that specifically trigger Toll-like receptors (TLR) 7/8, resulting in maturation of plasmacytoid dendritic cells (pDCs) and production of type I interferon (IFN), induction of inflammatory cytokines and chemokines in different antigen presenting cells (APCs). Cytokines produced by monocytes and pDCs upon stimulation of HCV-derived ssRNA inhibited HCV production in an IFN-dependent manner, whereas cytokines produced by myeloid DCs (mDCs) and macrophages had no inhibitory effect on virus production, due to a defect in IFN induction by HCV ssRNA in these cells.  TLR7/8 stimulated cytokines also down-regulated the expression of the HCV receptor CD81 on Huh7.5 in an IFN-independent manner that may restrict HCV infection. However, although HCV receptors like CD81 were widely expressed on different subsets of lymphocytes, DCs and monocytes did not respond to HCV particles, and our data indicates that only macrophages sense HCV and produce inflammatory cytokines. The recognition of HCV by macrophages is related to the expression of DC-SIGN on macrophages, and results in TLR7/8 engagement. Similar to a TLR7/8 agonist, HCV stimulation induces inflammatory cytokine production (TNF-α, IL-8, IL-6 and IL-1b) by macrophages but does not stimulate interferon-beta production in macrophages, Congruously, TLR7/8 and HCV RNA mediated cytokine production in macrophages did not inhibit HCV replication.  Our results reveal that HCV RNA has the potential to trigger TLR7/8 in DC populations, and initiate an innate immune response against HCV infection that leads to an IFN-dependent suppression of viral replication. However, HCV is able to escape detection by monocytes and DCs, which produce type I IFN and suppress HCV replication when TLR7/8 is engaged. Macrophages possess the capacity to detect HCV, in part because of surface expression of DC-SIGN. In turn, macrophages produce inflammatory cytokines that nevertheless fail to inhibit HCV replication due to lack of IFN-beta production. Evasion of antiviral defense by HCV may explain the failure of innate immunity in control of HCV infection. Moreover, inflammatory cytokine production by macrophages upon HCV stimulation in vitro suggests that activation of macrophages may contribute to inflammation in HCV infected individuals.

Keyphrases: TLR7, TLR8, CD81, DC-SIGN, Macrophages, Hepatitis C virus, Dendritic cells, Interferon, Single-strand RNA