| Co-Investigator(Kenkyū-buntansha) |
SIGEMI Hideo Dept.of Otolaryngology, Oita Medical University, Instructor, 医学部, 助手 (50271135)
KURONO Yuichi Dept.of Otolaryngology, Faculty of Medicine, Kagoshima University, Professor, 医学部, 教授 (80153427)
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| Research Abstract |
1)Mucosal immune responses against Haemophilus influenzaein human tonsils :
In order to clarify the immunological role of palatine tonsils in upper aerodigestive tract, the immune responses of tonsillar lymphocytes against P6 of nontypeable Haemophilus influenzae (NTHi) were investigated. Nasal secretions and sera were obtained frome the patients who underwent tonsillectomy and the antibody titers against P6 were determined by ELISA.Mononuclear cells were isolated from palatine tonsils by discontinuous gradient centrifugation and the numbers of antigen-specific antibody-secreting cells were determined by ELISPOT.CD4^+T cells isolated by use of anti-CD4 microbeads were incubated with P6 and the production of cytokines was determined by ELISA.The results showed that the number of P6-specific IgA-secreting cells was greater in the samples having no NTHi than that in the samples having the bacteria. The numbers of P6-specific IgA-secreting cells in tonsils were significantly correlated with
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IgA antibody titers in nasal secretions. CD4^+T cells produced Th2-and Th1-type cytokines in response to the stimulation with P6. Those findings suggest that palatine tonsils play an important role as an effector site and an inductive site for mucosal immune responses, and prevent bacterial infection in upper aerodigestive tract.
2)Mucosal immune responses against Haemophilus influenzaein mice
Mice were immunized intranasally, orally, intratracheally, or intraperitoneally with outer membrane proteins (OMP) of Haemophilus influenzae and cholera toxin (CT), and Th1/Th2 and B cell immune responses were compared to investigate the mechanisms of mucosal immune responses in the nose and the usefulness of intranasal immunization for inducing OMP-specific IgA responses. A combined vaccine of OMP and CT was administered on days 0,7, and 14. Antigen-specific antibody titers in saliva, nasal wash, bronchoalveolar lavage, and serum were determined by ELISA.On day 21, mononuclear cells isolated from nasal passages, lung, intestinal lamina propria, and spleen were analyzed by OMP-specific ELISPOT.CD4^+T cells isolated from spleen were cultured with OMP for 4 days, and the concentrations of cytokines in the culture supernatants were examined by ELISA.Following the immunization, a live bacterial suspension of the same strain as used for the preparation of OMP was inoculated into the nose, and the clearance of the bacteria from the nasal cavity was observed. Anti-OMP IgA antibody titers in saliva, nasal wash, and bronchoalveolar lavage and the numbers of OMP-specific IgA-producing cells in nasal passages were highest in mice immunized intranasally compared to the other groups of mice. Cytokine assay showed that IFN-g, IL-2, IL-5, and IL-6 were predominantly produced following intranasal immunization. In the clearance assay, significantly fewer bacteria were present in the nose of the intranasally immunized mice than in the control mice. These findings suggest that the nose is a powerful inductive and effector site for inducing IgA immune responses and that intranasal immunization enhances the clearance of pathogenic bacteria from the nose. Less
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