8 In addition to the ability of HCV to trigger the TLR3 pathway,9, 10 the increased number of Th17 cells appears to be associated with the severity of liver inflammation in chronic HCV patients and treatment of infected patients with pegylated BGB324 nmr interferon (IFN)-α and ribavirin reduced the level of Th17-related cytokines.11 As one of the crucial factors for Th17 differentiation, thymic stromal lymphopoietin (TSLP), a member of the common γ-chain cytokine, is capable of activating (conditioning) DCs, thereby stimulating naïve T cells to differentiate into Th2 cells.12 In addition, DCs treated with both TSLP and poly (I:C) activate naïve T cells and

differentiate into Th2 and Th17 cells.8, 13 Thus, TSLP-activated DCs, which are known to be strong inducers of Th2 responses, can simultaneously induce Th17 cells under certain pathological conditions. In this report EPZ 6438 we demonstrate that the infection of hepatic cells in vitro by HCV triggers robust TSLP production and this HCV-induced production of TSLP is regulated in an nuclear factor kappa B (NFκB)-dependent manner. TSLP secreted by HCV-infected cells activates and conditions human

monocyte-derived DCs to enhance the production of Th17 differentiating cytokines, TGF-β, IL-6, and IL-21 by the DCs. Moreover, the addition of TSLP neutralizing antibody to the coculture of monocytes/DCs with HCV-infected hepatocytes blocks the production of these cytokines. Consistent with these data, we find that the hepatocyte-derived TSLP is readily detected in liver biopsies from chronic HCV patients. Our studies suggest a novel role for the hepatocyte-derived TSLP in the generation of CD4+ Th17 effector T cells through its ability to condition DCs to drive CD4+ Th17 differentiation. The potential implications of these findings in the development of HCV-induced chronic progressive liver disease are discussed. DC, dendritic

cell; HCV, hepatitis C virus; TSLP, thymic stromal lymphopoietin. Human see more hepatoma cell lines, Huh 7.5.1, were maintained in Dulbecco’s modified Eagle’s medium (DMEM) with 10% fetal bovine serum (FBS) and penicillin/streptomycin (100 μg/mL). THP-1 cells purchased from the American Tissue Culture Collection (ATCC) were cultured in RPMI 1640 and supplements as recommended by ATCC. Liver biopsies and peripheral blood samples from chronic HCV or control patients were obtained from Dr. Hugo Rosen (University of Colorado). Blood samples were also obtained from the Virginia Blood Services. All information of age, gender, and HCV genotype were previously described.14, 15 For infection of cells with secreted HCV, Huh 7.5.1 permissive cells were seeded at 3 × 106 cells in a T75 plate for 24 hours. Cells were infected with 4 × 104 FFU (multiplicity of infection [MOI] of 0.01) of JFH-1 producing cell supernatant and cultured for 10 days in DMEM-10% FCS media.