This result is consistent with the observation that LCMV-induced IFN-α secretion in mice has been shown to increase STAT1 expression in NK cells, resulting in preferential STAT1 over STAT4 phosphorylation.6, 9, 11 It also extends the findings by Miyagi et al. on preferential STAT1 phosphorylation in HCV-infected patients,11 because we show that IFN-α exposure in vivo results in increased pSTAT1 levels, and that it correlates to increased TRAIL production and degranulation and decreased IFN-γ production

(Fig. 3). The clinical relevance of IFN-α signaling in NK cells is suggested by our observation that NK cell responsiveness and refractoriness correlate with the first-phase virological response to IFN-α-based therapy (Fig. 5). This analysis of NK cells is relevant for current research on biomarkers predicting IFN responsiveness and treatment outcome. Advantages of using NK cells check details as biomarkers

of IFN responsiveness are that they are readily accessible from the peripheral blood, and that both in vivo and in vitro NK cell responsiveness can easily be assessed in a short, standardized flow-cytometry–based assay by checking pSTAT1 levels. How does our system compare to other biomarkers of IFN responsiveness? A well-established biomarker for IFN responsiveness is the intrahepatic expression of interferon-stimulated genes (ISGs). Typically, ISGs are most highly expressed PD0332991 in vivo pretreatment in HCV-infected patients who respond poorly to IFN-α-based therapy.17 As a potential explanation, it has been proposed that high baseline activation of the endogenous IFN system does not allow a further increase of ISGs during IFN-based therapy, possibly because the ISG response has already reached maximal levels and/or inhibitory autocrine feedback mechanisms have been induced.18 In contrast to these ISG data, we did not find any evidence that pretreatment pSTAT1 levels or in vitro inducibility differed among HCV-infected patients (data not shown). Thus,

pSTAT1 induction is an independent measure for IFN responsiveness and may complement ISG analysis. How does the NK cell response correlate to the treatment response? Because all patients in our study achieved an EVR to PegIFN/RBV Flucloronide therapy at week 12, we were not able to assess NK cell responses in the context of the ultimate treatment outcome. On the other hand, we believe that late time points of PegIFN/RBV therapy are less relevant for our study, because NK cells exhibited their greatest response within the first days of therapy in parallel to the first-phase virological response (Figs. 1-4). Our data clearly indicate that near-maximal NK cell activation can be reached within hours of the first injection of PegIFN, because the response to additional in vitro stimulation with IFN-α was significantly reduced at later treatment time points (Fig. 4).