A ctino bacillus actinomycetemcomitans, Porphyromonas gingivalis, Fusobacterium nucleatum Influence of Complement and Antibody on Superoxide Productio

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1 A ctino bacillus actinomycetemcomitans, Porphyromonas gingivalis, Fusobacterium nucleatum Influence of Complement and Antibody on Superoxide Production, Actinobacillus actinomycetemcomitans, Porphyromonas gingivalis and Fusobacterium nucleatum Phagocytosis and Killing by Leukocytes Kazutaka Ogiwara*, Chihomi Kato and Kazuko Saito Department of Oral Microbiology, School of Dentistry at Niigata, The Nippon Dental University 1-8 Hamaura-cho, Niigata 951, Japan Director : Prof. Kazuko Saito) ( *Present address : Department of Periodontology, The Nippon Dental University Fujimi, Chiyoda-ku, Tokyo 102, Japan The purpose of this experiment was to clarify the influence of antibody and complement on the superoxide production [Chemiluminescence (CL) response] and periodontopathic bacteria phagocytosis and killing by polymorphonuclear leukocytes (PMNs). PMNs were obtained from the peritoneal cavity of ICR mice after caseinate injection. Three species of periodontopathic bacteria were used : Actinobacillus actinomycetemcomitans Aa), Porphyromonas gingivalis ( (Pg) and Fusobacterium nucleatum (Fn). Mice were immunized with live Aa, Pg or Fn vaccine. Serum was obtained from the blood by cutting the carotid artery. Normal serum was used as complement and inactivated immune serum was used as the antibody. The antibody titers were measured by ELISA using sonicated Aa, Pg and Fn antigen. Serum with a high antibody titer was obtained. IgG levels were high when mice were immunized with either Aa or Pg, whereas IgM levels were high when immunized with Fn. The reaction mixture for the CL response consisted of luminol solution, serum, gelatin Hanks' solution, PMNs and the bacteria. After the CL response was complete, a portion of the reaction mixture was smeared on a glass slide, dried and stained with May-Gruenwald- Giemsa stain. Phagocytic activity was assessed using a microscope. Surviving bacteria were counted on solid medium after treatment with serum, antibody and PMNs alone or in combination. An increase in the CL response to Aa occurred and was dependent on the addition of complement, and the response to Pg was dependent on both complement and antibody whereas the response to Fn dependent on neither complement nor antibody. The CL response to Fn was inhibited by D- galactose, N- acetyl-d-galactosamin and mannan. Phagocytic rate of PMN and the number of Aa and Fn bacteria in PMNs was enhanced in the presence of complement. The co-operation of both complement and antibody was needed to increase the phagocytic ratio. The bactericidal effect of complement, antibody and PMN were examined in single or combination mixtures. There was no

2 marked effect on killing the bacteria in any combination. In conclusion, complement plays the role of enhancing the CL response and phagocytic activity in response to Aa. Co-operation of complement was necesary for phagocytosis of Fn. High titer antibody, complement and PMN had no bactericidal effect under the experimental conditions. Key words : PMN, Antibody, Complement, CL response, Killing

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12 Derangement of nonspecific immunity in Down syndrome subjects: Low leukocyte chemiluminescence activity after phagocytic activation. American Jounal of Medical Genetics Supple)., 7: , ( 12) Lowry, O. H., Rosebrough, N. J., Farr, A. L. and Randall, R. L. : Protein measurement with the 1) Attstrom, R.: Presence of leukocytes in crevices N.: Humoral immune response to oral microorganisms in periodontitis. Infect. Immun., 37: of healthy and flammed gingivae. J. Periodontal. Res., 5: , ) Holm, A. and Kalfus, S.: Polymorphonuclear leukocyte chemiluminescence induced by Actino- 3) Listgarten, M. A., Lai, C. H. and Evain, C. I.: Comparative antibody titers to Actinobacillus actinobacillus actinomycetemcomitans and Haemophilus aphrophilus in serum and saliva, Scand. J. Dent. mycetemcomitans in juvenile periodontitis, chronic Res., 101: , periodontitis, and periodontally healthy subjects. J. Clin. Periodontol., 8: 155, ) Mouton, C., Hammond, P. G., Slots, J. and Genco, R. J.: Serum antibodies to oral Bacteroides asaccharolyticus (Bacteroides gingivalis) : relationship 16) Whyte, G. J., Seymour, G. J., Cheung, K. and Robin- to age and periodontal disease. Infect. Immun., son, M. F. : Chemiluminescence of peripheral 31: , polymorphonuclear leukocytes from adults periodositis patients. J. Clin. Periodontol., 16: 67-74, 5) Tolo, K. and Brandtzaeg, P.: Relation between periodontal disease activity and serum antibody titers to oral bacteria, Genco, R. T. and Mergenhagen, S. E., Host-parasite interactions in perigan, D. F. : A non-lectin-like mechanism by which 17) Tuttle, R. S., Strubel, N. A., Mourad, J. and Manodontal disease, Amer. Soc. Microbiol., Washigton, D. C., , activates human lymphocytes. Oral. Microbiol. Fusobacterium nucleatum adheres to and 6) Schenkein, H. A.: The role of complement in Immunol., 7: 78-83, periodontal disease. Oral. Biol. Medicine., 2: 65-18) Mangan, D. F., Novak, M. J., Vora, S. A., Mourad, 81, J. and Kriger, P. S.: Lectinlike interactions of 7) Nisengard, R. and Jarrett, C.: Coating of subgin- Fusobacterium nucleatum with human neutrophils. gival bacteria with immunogloblins and complement. J. Periodontol., 47: , ) Bissada, N. F., Mancou-chehrPour, M., Haddow, M. and Spagnuole, P. J. : Neutrophil functional activity in juvenile and adult onset diabetic patients with mild and severe periodontitis. J. Periodont. Res., 17: , ) Temple, T. R., Kimball, H. R., Kakehashi, S. and Amen, C. R.: Host factors in periodontal disease: Periodontal manifeststions of Chediak- Higashi syndrome. J. Periodont. Res., 7: 26-27, ) Genco, R. J. and Slots, J.: Host response: Host responses in periodontal disease. J. Oral. Pathol., 14: , ) Licastro, F., Melotti, C., Parente, R., Davis, L. J., Chiricolo, M., Zannotti, M. and Barboni, F.: folin phenol reagent. J. Biol. Chem., 193: , ) Doty, S. L., Lopatin, D. E., Syed, S. A. and Smith, F. Infect. Immun., 57: , ) Slots, J. and Genco, R. J.: Microbial oathogenicity. Black-pigmented Bacteroides species, Capnocytophaga species, and Actinobacillus actinomycetemcomitans in human periodontal disease: Virulence factors in colonization, survival, and tissue destruction. J. Dent. Res., 63: , ) Wilson, M. E. and Genco, R. J.: The role of antibody, complement and neutrophils in host defense against Actinobacillus actinomycetemcomitans. Immunol. Invest., 18: , ) Greiner, D., Mayrand, D. and McBride, B. C.: Further studies of the degradation of immunoglobulins by black-pigmented bacteroides. Oral. Immunol., 4: 12-18, ) Cutler, C. W., Kalmar, J. R. and Arnold, R. R.:

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