6A). It is noteworthy that in the three woodchucks that received IL-12 only, a decrease of wIL-10 and wPD-L1 was observed, and in two of these animals this was associated with an increase in IFN-γ expression (Fig. 6A). This result suggests ABT-263 mw that the combination of IL-12 treatment and block of Treg activity may cause/induce/have a detrimental effect. The expression

of Class I, β2-m, CD3, and CD8 molecules, however, remained unchanged in all animals (Supporting Fig. 3A). Untreated control woodchucks had no changes in the expression of these molecules (Supporting Fig. 3B), indicating that the increase in the immunosuppressive environment of the liver was due to the applied treatment. In chronic hepatitis B the failure to mount an efficient immune response against viral antigens allows continuing viral replication and progression of liver damage. T-cell function is particularly affected in this condition. The mechanism underlying impaired T-cell reactivity is not fully understood. Immunotherapy of chronic HBV infection aims at activating antiviral

T-cell immunity to promote seroconversion and long-lasting control of viral replication. In a previous study we treated woodchucks with chronic WHV infection with a gene transfer vector to express intrahepatically the immunostimulatory cytokine IL-12 under the control of an inducible promoter.18 Idelalisib chemical structure We observed that the induction of IL-12 expression resulted in a significant reduction of viral load and find more stimulation of specific anti-WHV immune response. More important, the activation of antiviral immunity was associated with a reduction of WHV covalently closed circular DNA (cccDNA) forms from the liver that are mainly responsible for the maintenance of persistent viral infection. However, the decrease in viral load was only observed in woodchucks with viremias lower than 1010 vg/mL. Woodchucks with higher viremia did not respond to treatment, and surprisingly, at the end of IL-12 administration hepatic expression of FoxP3 was significantly increased, whereas in the responder

animals FoxP3 expression was clearly reduced. Further analysis showed that peripheral blood lymphocytes obtained from woodchucks with high viral load failed to respond to IL-12 stimulation. In the present study we found that the frequency of Treg in liver of WHV chronically infected woodchucks was higher than that observed in uninfected animals. The increase in hepatic Treg was accompanied by significantly higher expression of antiinflammatory cytokines such as TGF-β1 and IL-10, and immunosuppressive molecules such as PD-1 and PD-L1. Thus, similar to chronic HBV infection, persistent infection in WHV infection is associated with a strong immunosuppressive environment within the liver.