Chronic hepatitis C virus (HCV) infection is characterized by diminished numbers and function of HCV-reactive T cells and impaired responses to immunization. Because host response to viral infection likely involves TLR signaling, we examined whether chronic HCV infection impairs APC response to TLR ligand and contributes to the origin of dysfunctional T cells. Freshly purified myeloid dendritic cells (MDC) and plasmacytoid DC (PDC) obtained from subjects with chronic HCV infection and healthy controls were exposed to TLR ligands (poly(I:C), R-848, or CpG), in the presence or absence of cytokine (TNF-α or IL-3), and examined for indices of maturation and for their ability to activate allogeneic naive CD4 T cells to proliferate and secrete IFN-γ. TLR ligand was observed to enhance both MDC and PDC activation of naive CD4 T cells. Although there was increased CD83 and CD86 expression on MDC from HCV-infected persons, the ability of MDC to activate naive CD4 T cells in the presence or absence of poly(I:C) or TNF-α did not differ between HCV-infected and healthy control subjects. In contrast, PDC from HCV-infected persons had reduced activation marker (HLA-DR) and cytokine (IFN-α) expression upon R-848 stimulation, and these were associated with impaired activation of naive CD4 T cells. These data indicate that an impaired PDC responsiveness to TLR ligation may play an important role in the fundamental and unexplained failure to induce new T cell responses to HCV Ags and to other new Ags as a consequence of HCV infection.

, and this activity was impaired in viremic HIV infection but not in HCV infection. In heterologous PDC-NK cell assays, impaired PDC-NK cell killing activity was largely attributable to an NK cell defect, while impaired PDC-NK cell IFN-␥-producing activity was attributable to both PDC and NK cell defects. Additionally, the response of NK cells to direct IFN-␣ stimulation was defective in viremic HIV infection, and this defect was not attributable to diminished IFN-␣ receptor expression, though IFN-␣ receptor and NKP30 expression was closely associated with killer activity in viremic HIV infection but not in healthy controls. These data indicate that during uncontrolled HIV infection, PDC-dependent NK cell function is impaired, which is in large part attributable to defective IFN-␣-induced NK cell activity and not to altered IFN-␣ receptor, NKP30, NKP44, NKP46, or NKG2D expression.Immature dendritic cells (DC) are key innate mediators of the adaptive immune response. Myeloid DC (MDC) and plasmacytoid DC (PDC) have been identified as two main peripheral DC subsets (43). Numerical and functional defects in these populations have been described for both hepatitis C virus (HCV) and human immunodeficiency virus (HIV) infections, with the impairments distinctly different in each infection (3,12,19,26,33,48,53,56,57). Natural killer (NK) cells are capable of cytotoxic, cytokine-expressing, and chemokine-expressing functions (31, 38). These lymphocytes are vital during the early stages of mouse hepatic viral infection (22, 42) and during human herpes virus infection (5, 7, 10). NK cell cytotoxic function has long been known to be reduced during chronic HIV infection and in subjects with AIDS (32, 44), and unfractionated cell assays of NK cell function indicate an impaired NK cell response to alpha interferon (IFN-␣) (52). Additionally, genetic markers of NK cell phenotype are associated with disease progression rate (36). In contrast, HIVexposed but uninfected subjects appear to have enhanced NK cell function (45), and after highly active antiretroviral therapy (HAART), NK cell numbers and function appear to normalize (1, 4). In the setting of HCV infection, genetic markers of NK cell phenotype appear to predict the outcome after acute exposure (28). During chronic HCV infection, some studies indicate reduced NK cell cytotoxicity (14, 40, 55), while morerecent studies indicate normal NK cell function and reduced peripheral NK cell numbers (18,27,39).DC-NK cell bidirectional cross talk has recently been shown to play a key role in host defense (20,30,34,35,37). This cross talk can be facilitated by Toll-like receptor (TLR) signaling and results in NK cell activation, enhanced NK cell effector function, and DC maturation (15,20,21,30,49). In the setting of viremic HIV infection, recent unfractionated cell system data indicate impairment in PDC-dependent NK cell activity (11). These data may be explained by the previously described numerical defects in PDC or NK cells, though whether there are additional functional defects ...

Chronic allograft rejection is in part mediated by host T cells that recognize allogeneic antigens on transplanted tissue. One factor that determines the outcome of a T cell response is clonal size, while another is the effector quality. Studies of alloimmune predictors of transplant graft survival have most commonly focused on only one measure of the alloimmune response. Because differing qualities and frequencies of the allospecific T cell response may provide distinctly different information we analyzed the relationship between frequency of soluble antigen and allo-antigen specific memory IFN-γ secreting CD4 and CD8 T cells, their ability to secrete IL-2, and their proliferative capacity, while accounting for cognate and bystander proliferation. The results show proliferative responses primarily reflect on IL-2 production by antigen-specific T cells, and that proliferating cells in such assays entail a considerable fraction of bystander cells. On the other hand, proliferation (and IL-2 production) did not reflect on the frequency of IFN-γ producing memory cells, a finding particularly accentuated in the CD8 T cell compartment. These data provide rationale for considering both frequency and effector function of pre-transplant T cell reactivity when analyzing immune predictors of graft rejection.

The activation state, differentiation state, and functions of liver lymphocytes and perihepatic lymph nodes during chronic hepatitis C virus (HCV) infection are not well understood. Here, we performed phenotypic and functional analyses of freshly prepared lymphocytes isolated from the livers, perihepatic lymph nodes, and peripheral blood compartments of chronic HCV-infected and disease control subjects with end-stage liver disease undergoing liver transplantation. We measured lymphocyte subset frequency and memory T-cell gamma interferon (IFN-␥) and proliferative responses to HCV peptide and control viral antigens in direct ex vivo assays. We found higher frequencies of CD4 cells in the lymph node compartment than in the other compartments for both HCV-infected and disease control subjects. Lymph node CD4 and CD8 cells less commonly expressed the terminal differentiation marker CD57, a finding consistent with an earlier differentiation state. In HCV-infected subjects, HCV-specific IFN-␥-producing and proliferative responses were commonly observed in the lymph node fraction, while they were uncommonly observed in the peripheral blood or liver fractions. In contrast, control viral CD4 protein antigen and CD8 peptide antigen-specific IFN-␥ responses were commonly observed in the periphery and uncommonly observed in the lymph nodes of these same subjects. These findings are consistent with a selective defect in HCV-specific T-cell effector function or distribution in patients with advanced chronic HCV infection. The high frequency of HCV-reactive T cells in perihepatic lymph nodes indicates that a failure to generate or sustain T-lymphocyte HCV reactivity is not responsible for the paucity of functional cells even in end-stage liver disease.During hepatitis C virus (HCV) infection, both CD4 and CD8 virus-specific lymphocytes are thought to play important roles both in control/clearance of viral infection and in hepatocellular damage (7,9,13,23,26,32,33,37,38,40). Priming of these cells during acute infection and sustained activation of memory cells during chronic infection are thought to take place in regional lymph nodes that drain the tissue sites of infection. Chronic HCV infection is characterized by low frequencies of HCV-reactive T cells in circulation (16,34,(42)(43)(44). While there are a number of potential explanations for these meager responses, the relative distribution levels of HCVreactive T cells in lymph nodes, liver, and peripheral blood are not well recognized. We therefore examined the phenotype and function of HCV-specific T cells within the regional lymph nodes, livers, and peripheral blood of HCV-infected individuals, using freshly isolated tissue samples in order to obtain an improved understanding of the dominant T-cell dynamics during HCV infection and to establish a foundation for improved models of compartmentalized T-cell activation during chronic viral hepatitis. Results indicate similar paucities of functional HCV-specific T cells in the liver and periphery, while HCVreactive T cells ar...