Analysis and Modulation of Virus-Host Interaction in Ani

Diabetes, Digestive, Kidney Diseases

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Abstract

Whereas it is generally accepted that vigorous hepatitis C virus (HCV)-specific T cell responses are associated with recovery from HCV infection and that weak responses are associated with persistent HCV infection, the mechanisms that determine the strength of the immune response are not known. Here, we examined two candidate mechanisms that are thought to influence the effectiveness of HCV-specific T cell responses and the outcome of HCV-infection.[unreadable] [unreadable] Trafficking of hepatitis C virus (HCV)-specific T cells to the liver depends on the induction of chemokine receptors on T cells and their respective ligands in the infected liver. Because HCV-specific T cells are not detectable in the liver until 4-8 weeks after infection, it has been suggested that delayed trafficking of HCV-specific T cells to the liver, the site of infection, may account for HCV persistence. [unreadable] Here, we prospectively studied the kinetics and mechanisms of T cell trafficking in 5 chimpanzees, infected with 100 CID50 monoclonal HCV 1a. A self-limited course of infection was observed in 2 chimpanzees and associated with CD8 cell-mediated IFN-gamma production at 4-6 weeks p.i. in response to overlapping HCV peptides. In contrast, this response was absent or delayed in 3 chimpanzees with chronically evolving hepatitis. Irrespective of the outcome of infection, however, staining of peripheral blood CD8 T cells with 5 HCV/MHC class I (Patr)-tetramers revealed HCV-specific CD8 cells in all chimpanzees at week 3. The majority (60-90%) of HCV/Patr-tetramer+ cells expressed CXCR3, a minority (less than 10%) expressed CCR5. The frequency of CXCR3+ cells within the HCV/Patr-tetramer+ population remained constant in all chimpanzees until week 5, after which it suddenly dropped by 30-70% suggesting trafficking of HCV-specific CXCR3+CD8+ cells into tissues. Indeed, intrahepatic CD8b-mRNA levels started to increase at the same time (week 6-9) when intrahepatic IFN-gamma mRNA and serum ALT levels started to increase. These events were preceded by an increase of intrahepatic IP-10, which began at week 2 and peaked immediately prior to the increase in CD8b and IFN-gamma, thus suggesting that T cell trafficking occurred in response to IP-10, the CXCR3 ligand. The induction of IP-10 correlated well to the induction of intrahepatic 2'5' OAS-1 and we have observed induction of IP-10 by IFN-alpha in Huh7 cells in vitro, suggesting that IP-10 was regulated by IFN-alpha. [unreadable] These data demonstrate that delayed trafficking of HCV-specific CD8 T cells to the liver is not due to lack of or late expression of the chemokine receptor CXCR3, but that it follows the apparent intrahepatic induction of IP-10 by IFN-alpha. Because expression of chemokines and chemokine receptors did not differ between self-limited and chronically evolving acute hepatitis C, functional differences in HCV-specific T cell responses rather than differences in T cell trafficking appear to determine the outcome of infection.[unreadable] [unreadable] One of these qualitative factors that influence the outcome of HCV infection is the production of IFN-gamma in the liver, the site of HCV infection. Vigorous IFN-gamma production by HCV-specific T cell has been associated with a self-limited course of infection. In addition, IFN-gamma has been shown to inhibit replication of subgenomic and genomic hepatitis C virus (HCV) RNAs in vitro and to noncytolytically suppress hepatitis B virus (HBV) replication in vivo. We therefore hypothesized that therapeutic expression of IFN-gamma in the liver may facilitate resolution of chronic hepatitis C, an infection that is rarely resolved spontaneously. [unreadable] To analyze immunomodulatory and antiviral effects of liver-specific IFN-gamma expression in vivo, we intravenously injected two persistently HCV-infected chimpanzees twice with a recombinant, replication-deficient HBV vector and subsequently with a recombinant adenoviral vector. These vectors expressed human IFN-gamma under control of HBV- and liver-specific promoters, respectively. Gene transfer resulted in a transient increase of intrahepatic IFN-gamma mRNA, without increase in serum alanine aminotransferase levels. Ex vivo analysis of peripheral blood lymphocytes demonstrated enhanced CD16 expression on T cells and upregulation of the liver-homing marker CXCR3. Moreover, an increased frequency of HCV-specific T cells was detected ex vivo in the peripheral blood and in vitro in liver biopsy-derived, antigen-nonspecifically expanded T cell lines. None of these immunologic effects were observed in the third chimpanzee injected with an HBV control vector. Despite these immunologic effects of the experimental vector, however, IFN-gamma gene transfer did not result in a significant and long-lasting decrease of HCV titers. In conclusion, liver-directed IFN-gamma gene delivery resulted in HCV-specific and nonspecific activation of cellular immune responses, but not in effective control of HCV replication.