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Su.40. Differential Antigen Processing Activities of PBMC Subsets Targeted by HIV Modulate HIV Epitope Production
Abstracts a rapid and broadened anamnestic response to subsequent boosting with PA83. Cynomolgus macaques were immunized intranasally on days 0 and 14 with CVD 908-htrA expressing ClyA-PA83 (n = 12) or CVD 908-htrA carrying an empty plasmid (n = 8). Three months after priming, monkeys of both groups were evenly divided to be boosted either with 85 µg of rPA83 plus alum or 0.5 mL of Biothrax (the currently licensed anthrax vaccine). All monkeys elicited serum IgG anti-S. Typhi LPS in response to the live vector priming. One week after the boost, all animals primed with S. Typhi(ClyA-PA83) exhibited remarkably high levels of anthrax toxin neutralization antibodies (TNA), regardless of the boosting agent (rPA83-alum or Biothrax). On the contrary, unprimed animals lacked detectable TNA responses (p b 0.05). TNA titers peaked on day 113 in animals primed with CVD 908-htrA (ClyA-PA83). This is the first report describing effective mucosal immune responses to B. anthracis PA83 observed in non-human primates using an S. Typhi-based prime-boost approach. doi:10.1016/j.clim.2008.03.389
Su.39. Loss of Virus-specific Memory T Cells in Coxsackievirus B3 and B4-Infected Mice Lu Li, Alfred Dufour. U.S. EPA, Cincinnati, OH There are two major types of enteroviruses: polioviruses and non-polio enteroviruses. While vaccines have effectively eliminated poliovirus infections, no vaccine is currently available for the non-polio enteroviruses. Generation of long-term surviving, pathogen specific, memory cells is critical for the development of a good vaccine. To investigate whether long-term memory Tcells are produced in response to infection by non-polio enteroviruses, coxsackieviruses B3 (CVB3) and coxsackieviruse B4 (CVB4) were intraperitoneally inoculated into adult BABL/c mice. Virus-specific memory T cells were assayed for proliferation and release interferon gamma (IFN-γ) under ex vivo stimulation with the viral pathogens. The level of IFN-γ was measured by antibody-capture chemiluminescent ELISA. Four days after stimulation, there were no detectable changes in the levels of IFN-γ from mice 17 months postexposure to CVB3 and CVB4 compared with negative control mice inoculated only with sterilized PBS buffer. In contrast, the levels of IFN-γ were markedly increased after stimulation with specific viral pathogens in mice 21/2 months post-infection with CVB3 and CVB4. These results suggest that non-polio enteroviruses might generate only short-lived virus-specific memory T cells. This would imply that vaccines for non-polio enterviruses might not provide long term protection against these viruses. This study might explain why people can repeatedly be infected by non-polio enteroviruses. doi:10.1016/j.clim.2008.03.390
Su.40. Differential Antigen Processing Activities of PBMC Subsets Targeted by HIV Modulate HIV Epitope Production Estibaliz Lazaro,1 Sasha Blue Godfrey,1 Pamela Stamegna,1 Jeremy Ho,1 Bruce Walker,2 Sylvie Le Gall. 1 1MGH and
S137 Harvard Medical School, Charlestown, MA; 2HHMI, Chevy Chase, MD Background: Immune recognition of HIV-infected cells requires peptide presentation by major histocompatibility complex I (MHC-I) at the cell surface. These peptides result from a cascade of proteolytic events involving proteasome and aminopeptidases. Although HIV infects several cellular types, the capacity of each of them to process epitopes is unknown. Here we compare epitope processing activities of primary CD4 T cells and monocytes. Methods: CD4 T cells and monocytes were enriched by magnetic immunosorting from peripheral blood monocytic cells (PBMC) of 20 healthy donors. Proteasome and aminopeptidase activities were measured for each cell subset using specific fluorogenic substrates. The capacity of the extracts to produce epitopes was assessed during the in vitro degradation of long HIV Gag or Pol peptides. Kinetics, identity and amount of digestion products were identified by HPLC and mass spectrometry. Their antigenicity was measured by 51Cr release assay with epitope-specific CTL. Results: Proteasome and aminopeptidase activities were significantly higher in monocytes than in PBMC or CD4 T cells. These differences affected the identity, amount and kinetics of peptides produced. Monocytes extracts yielded the largest and fastest production of optimal epitopes or extended antigenic peptides and consequently the antigenicity of the degradation peptides produced in monocyte extracts was 5-fold higher than that of CD4 T cells. Conclusion: Differential antigen processing activities in two PBMC subsets targeted by HIV affects the identity, amount and antigenicity of peptides produced. This may alter the capacity of CD8 T cells to recognize infected cells. doi:10.1016/j.clim.2008.03.391
Su.41. Mucosal Salmonella Typhi Prime Followed by Parenteral Subunit Vaccine Boost Can Serve as an Effective Vaccine Strategy for Early Life Immunization Karina Ramirez, Liliana Rodriguez, Katherine Davis, James Galen, Marcela Pasetti. University of Maryland School of Medicine, Baltimore, MD Neonates have a limited capacity to generate protective immunity in response to vaccination. We have shown that Salmonella-based live vector vaccines are excellent candidates to induce immune responses against a foreign antigen at very early life stages. Recognizing that biodefense vaccines will be needed for high-risk groups such as young infants and children, we investigated the immunogenicity of the licensed attenuated S. Typhi vaccine Ty21a expressing Bacillus anthracis protective antigen (Ty21a-PA). Newborn mice were immunized intranasally with Ty21a-PA on day 7 and 14 after birth, and boosted intramuscularly 14 days later with PA-alum. Control groups received Ty21a alone or PBS followed by PA-alum. Newborns primed with Ty21a-PA and boosted with PA-alum induced high levels of PA-specific IgG and toxin neutralizing antibodies that surpassed those of the unprimed group. Live vector priming elicited strong cellmediated immunity (T cell proliferation and IFN-γ secretion)
Su.39. Loss of Virus-specific Memory T Cells in Coxsackievirus B3 and B4-Infected Mice Lu Li, Alfred Dufour. U.S. EPA, Cincinnati, OH There are two major types of enteroviruses: polioviruses and non-polio enteroviruses. While vaccines have effectively eliminated poliovirus infections, no vaccine is currently available for the non-polio enteroviruses. Generation of long-term surviving, pathogen specific, memory cells is critical for the development of a good vaccine. To investigate whether long-term memory Tcells are produced in response to infection by non-polio enteroviruses, coxsackieviruses B3 (CVB3) and coxsackieviruse B4 (CVB4) were intraperitoneally inoculated into adult BABL/c mice. Virus-specific memory T cells were assayed for proliferation and release interferon gamma (IFN-γ) under ex vivo stimulation with the viral pathogens. The level of IFN-γ was measured by antibody-capture chemiluminescent ELISA. Four days after stimulation, there were no detectable changes in the levels of IFN-γ from mice 17 months postexposure to CVB3 and CVB4 compared with negative control mice inoculated only with sterilized PBS buffer. In contrast, the levels of IFN-γ were markedly increased after stimulation with specific viral pathogens in mice 21/2 months post-infection with CVB3 and CVB4. These results suggest that non-polio enteroviruses might generate only short-lived virus-specific memory T cells. This would imply that vaccines for non-polio enterviruses might not provide long term protection against these viruses. This study might explain why people can repeatedly be infected by non-polio enteroviruses. doi:10.1016/j.clim.2008.03.390
Su.40. Differential Antigen Processing Activities of PBMC Subsets Targeted by HIV Modulate HIV Epitope Production Estibaliz Lazaro,1 Sasha Blue Godfrey,1 Pamela Stamegna,1 Jeremy Ho,1 Bruce Walker,2 Sylvie Le Gall. 1 1MGH and
S137 Harvard Medical School, Charlestown, MA; 2HHMI, Chevy Chase, MD Background: Immune recognition of HIV-infected cells requires peptide presentation by major histocompatibility complex I (MHC-I) at the cell surface. These peptides result from a cascade of proteolytic events involving proteasome and aminopeptidases. Although HIV infects several cellular types, the capacity of each of them to process epitopes is unknown. Here we compare epitope processing activities of primary CD4 T cells and monocytes. Methods: CD4 T cells and monocytes were enriched by magnetic immunosorting from peripheral blood monocytic cells (PBMC) of 20 healthy donors. Proteasome and aminopeptidase activities were measured for each cell subset using specific fluorogenic substrates. The capacity of the extracts to produce epitopes was assessed during the in vitro degradation of long HIV Gag or Pol peptides. Kinetics, identity and amount of digestion products were identified by HPLC and mass spectrometry. Their antigenicity was measured by 51Cr release assay with epitope-specific CTL. Results: Proteasome and aminopeptidase activities were significantly higher in monocytes than in PBMC or CD4 T cells. These differences affected the identity, amount and kinetics of peptides produced. Monocytes extracts yielded the largest and fastest production of optimal epitopes or extended antigenic peptides and consequently the antigenicity of the degradation peptides produced in monocyte extracts was 5-fold higher than that of CD4 T cells. Conclusion: Differential antigen processing activities in two PBMC subsets targeted by HIV affects the identity, amount and antigenicity of peptides produced. This may alter the capacity of CD8 T cells to recognize infected cells. doi:10.1016/j.clim.2008.03.391
Su.41. Mucosal Salmonella Typhi Prime Followed by Parenteral Subunit Vaccine Boost Can Serve as an Effective Vaccine Strategy for Early Life Immunization Karina Ramirez, Liliana Rodriguez, Katherine Davis, James Galen, Marcela Pasetti. University of Maryland School of Medicine, Baltimore, MD Neonates have a limited capacity to generate protective immunity in response to vaccination. We have shown that Salmonella-based live vector vaccines are excellent candidates to induce immune responses against a foreign antigen at very early life stages. Recognizing that biodefense vaccines will be needed for high-risk groups such as young infants and children, we investigated the immunogenicity of the licensed attenuated S. Typhi vaccine Ty21a expressing Bacillus anthracis protective antigen (Ty21a-PA). Newborn mice were immunized intranasally with Ty21a-PA on day 7 and 14 after birth, and boosted intramuscularly 14 days later with PA-alum. Control groups received Ty21a alone or PBS followed by PA-alum. Newborns primed with Ty21a-PA and boosted with PA-alum induced high levels of PA-specific IgG and toxin neutralizing antibodies that surpassed those of the unprimed group. Live vector priming elicited strong cellmediated immunity (T cell proliferation and IFN-γ secretion)