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Murine cytomegalovirus replication in salivary glands is controlled by perforin, granzyme A and granzyme B during acute infection
Abstracts
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CMV Immune Response- Posters G3-22 Prevalence of the IEl-exon 4-specific CTL in naturally seropositive healthy individuals KATALIN BURIAN Albert Szent-Gyorgyi Medical University, Szeged, Hungary The importance of CMV pp65 protein as CTL target has been reported earlier (Gilbert et al., 1993). However, data concerning the prevalence of IE proteins, especially IEl-exon4 in CTL responses, are rather limited (Borysiewicz et al., 1988; Mark et al., 1993). Thirty-two healthy HCMV-seropositive blood donors were thus tested for CTL responses directed to HCMV IEl-exon4 and pp65. Among these subjects, 93% had pp65-specific CTL and 79% exhibited IEl-exon4-specific CTL. The above data were obtained by restimulating PBMC with a canarypox recombinant expressing the HCMV IEl-exon4 or pp65 protein, and then testing their cytolytic activity on vacciniaIE or pp65 recombinant-infected, EBV-transformed autologous and mismatched B lymphoblast target cells, respectively. To confirm the high percentage of donors in our study with IEl-exon4-specific CTL, autologous fibroblast cells infected with HCMV were also used either for restimulation of effector cells or as target cells. Since laboratory strains of HCMV might express more pp65 in infected cells than fresh HCMV isolates, which could favor the induction of pp65-specific CTL, we also used fresh HCMV isolates in the CTL assays. Fresh HCMVinfected fibroblast cells could restimulate both IEl-exon4- and pp65-specific CTL. Furthermore, fresh HCMVinfected fibroblasts were better targets for the IE-specific CTL than those infected by the Towne laboratory strain. The precursor frequencies of IEl-exon4- and pp65-spedfic CTL were analyzed by limiting dilution assay and were found comparable in the two donors tested. We concluded that not only pp65-specific, but also IEl-exon4-specific CTL may play an important role in CTL-mediated immunity after natural infection.
G3-23 Murine cytomegalovirus replication in salivary glands is controlled by perforin, granzyme A and granzyme B during acute infection LUDOVICA RIERA Dept. of Public Health and Microbiology, Medical School, Turin, Italy Previous studies have shown that both CD8+ and CD4+ T cells are involved in the control of primary Cytomegalovirus (MCMV) infection in mice and essential for clearance of virus from salivary glands. The precise role of either of these two T cell subsets in the course of MCMV infection and the establishment of latent infection is not clear. Nor is it understood by which mechanism CD8+ and CD4+ T ceils control virus replication in various organs. We show the course of MCMV virus infection by determining virus titers and histological alterations in major target tissues in wild type and immunodeficient C57BL/6 (RAG2-/-) mice and mice deficient in either perforin, granzyme A or granzyme B. At 15 days p.i., virus titers and corresponding inflammatory lesions were elevated in the salivary glands of all ko compared to C57BL/6 mice. No virus was detectable in any mice in lung and spleen tissues at this time point. At 30 days p.i., virus titers were measurable in salivary glands, lung and spleen tissues only in RAG2-/- mice, but not in other ko or wt mice. In RAG2-/- mice, tissues showed extensive inflammatory lesions and some mortality occurred. The data dernonstrate that in the acute phase of MCMV infection perforin, granzyme A and B contribute to the control of virus replication in salivary glands, but indicate that neither of the three molecules alone are indispensable for the final control of infection.
141
CMV Immune Response- Posters G3-22 Prevalence of the IEl-exon 4-specific CTL in naturally seropositive healthy individuals KATALIN BURIAN Albert Szent-Gyorgyi Medical University, Szeged, Hungary The importance of CMV pp65 protein as CTL target has been reported earlier (Gilbert et al., 1993). However, data concerning the prevalence of IE proteins, especially IEl-exon4 in CTL responses, are rather limited (Borysiewicz et al., 1988; Mark et al., 1993). Thirty-two healthy HCMV-seropositive blood donors were thus tested for CTL responses directed to HCMV IEl-exon4 and pp65. Among these subjects, 93% had pp65-specific CTL and 79% exhibited IEl-exon4-specific CTL. The above data were obtained by restimulating PBMC with a canarypox recombinant expressing the HCMV IEl-exon4 or pp65 protein, and then testing their cytolytic activity on vacciniaIE or pp65 recombinant-infected, EBV-transformed autologous and mismatched B lymphoblast target cells, respectively. To confirm the high percentage of donors in our study with IEl-exon4-specific CTL, autologous fibroblast cells infected with HCMV were also used either for restimulation of effector cells or as target cells. Since laboratory strains of HCMV might express more pp65 in infected cells than fresh HCMV isolates, which could favor the induction of pp65-specific CTL, we also used fresh HCMV isolates in the CTL assays. Fresh HCMVinfected fibroblast cells could restimulate both IEl-exon4- and pp65-specific CTL. Furthermore, fresh HCMVinfected fibroblasts were better targets for the IE-specific CTL than those infected by the Towne laboratory strain. The precursor frequencies of IEl-exon4- and pp65-spedfic CTL were analyzed by limiting dilution assay and were found comparable in the two donors tested. We concluded that not only pp65-specific, but also IEl-exon4-specific CTL may play an important role in CTL-mediated immunity after natural infection.
G3-23 Murine cytomegalovirus replication in salivary glands is controlled by perforin, granzyme A and granzyme B during acute infection LUDOVICA RIERA Dept. of Public Health and Microbiology, Medical School, Turin, Italy Previous studies have shown that both CD8+ and CD4+ T cells are involved in the control of primary Cytomegalovirus (MCMV) infection in mice and essential for clearance of virus from salivary glands. The precise role of either of these two T cell subsets in the course of MCMV infection and the establishment of latent infection is not clear. Nor is it understood by which mechanism CD8+ and CD4+ T ceils control virus replication in various organs. We show the course of MCMV virus infection by determining virus titers and histological alterations in major target tissues in wild type and immunodeficient C57BL/6 (RAG2-/-) mice and mice deficient in either perforin, granzyme A or granzyme B. At 15 days p.i., virus titers and corresponding inflammatory lesions were elevated in the salivary glands of all ko compared to C57BL/6 mice. No virus was detectable in any mice in lung and spleen tissues at this time point. At 30 days p.i., virus titers were measurable in salivary glands, lung and spleen tissues only in RAG2-/- mice, but not in other ko or wt mice. In RAG2-/- mice, tissues showed extensive inflammatory lesions and some mortality occurred. The data dernonstrate that in the acute phase of MCMV infection perforin, granzyme A and B contribute to the control of virus replication in salivary glands, but indicate that neither of the three molecules alone are indispensable for the final control of infection.