Key words : Mycoplasma pneumoniae, Protective effect, 85KDa protein, non-hemolysis mutant Mycoplasma pneumoniae M. pneumoniae pneumonzae M. pneumoniae
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1 Key words : Mycoplasma pneumoniae, Protective effect, 85KDa protein, non-hemolysis mutant Mycoplasma pneumoniae M. pneumoniae pneumonzae M. pneumoniae (M. pneumoniae P24-S1, P24-S11)
2 M. pneumoniae FH-P2411) moniae FH-P24-S18) M. pneumoniae FH-P24-S11
3 Table 1 Protective effect in BALB/c mice of immunization mutants of M. pneumoniae with parent strain and hemolysis 8 weeks after last vaccination Table 2 Protective effect in BALB/c mice of immunization with parent strain and hemolysis mutants of M. pneumoniae 12 weeks after vaccination
4 Fig. 1 Humoral immune responses. The mean ELISA OD is represented by each point. Fig. 2 Cell mediated immune responses (MITagarose droplet method). Each column represented % of macrophage migration inhibition. Table 3 Immunological 8 weeks after last vaccination status of BALB/c mice at the time of challenge 1) Mean optical density at 405nm }SD 2) % of migration inhibition: Macrophage migration inhibition test by agarose droplet method.
5 Table 4 Immunological 12 weeks after vaccination status of BALB/c mice at the time of challenge 1) Mean optical density at 405nm }SD 2) % of migration inhibition: Macrophage migration inhibition test by agarose droplet method. Table 5 Protective effect in BALB/c mice after transfer of antiserum M. pneumoniae FH-P24 and mutant strains 1) Antiserum was prepared by injection of strain FH-P24 or mutant strains into guinea-pigs. 2) Antiserum was adjusted to equal ELISA IgG values and ten-fold dilution. Table 6 ELISA IgG value and recovery from trachea and lung of passively BALB/c mice after challenged with M. pneumoniae FH-P24
6 Mycoplasma Fig. 3 IgG-immunoblots pneumoniae免 developed with 疫 マ ウス の感 染 防御 効 果 sera from patients 689 of M.pneumoniae infection. M: marker proteins, n: M.pneumoniae pneumoniae FH-P24, b: M.pneumoniae x: P1 (165-KDa), y: 85-KDa (Coomassie P24-S1, blue し,ま た 細 胞 性 免 疫 が これ よ り低 下 した12週 後 で るP1に も防 御 率 は 低 下 し な か った と い う結 果 が 得 ら れ れ て い る が,そ た.さ らに免 疫23週 後 のP24-S1生 92KDa,62KDa21)等 IgG抗 菌 免 疫 群 で は, 体 価 は上 昇,細 胞 性 免 疫 は対 照 群 と同 じ値 に まで 低 下 した に もか か わ らず,防 御効果 に変化 は な か っ た(デ ー タ不 示).IgA,IgMに つ いて は 生 菌 免 疫8週 後,12週 られ な か った.こ 後 とも に 対 照 群 との差 は み の こ とか ら菌 の 分 離 を 指 標 に し も,ヒ stained) c: M. pneumoniae つ い て は,と く に 多 く の 報 告16) 18)がな さ れ 以 外 に も43KDa19),32KDa20), の 報 告 も み ら れ る.本 immunoblottingに よ り,親 う ち,お は み ら れ な か っ た と い う違 い が み ら れ た.こ れ が,生 可 能 性 を 示 唆 し て い る. 得 た.一 抗 血 清 で は20%の 防 御効 果 を 方 生 菌 免 疫 で は親 株 と変 らな い 防 御 効 果 を 示 したP24-S1の 抗 血 清 で 受 身 免 疫 した 群 と,効 果 の な か ったP24-S11受 果 を 示 した.す 身 免疫 群 とは ほ ぼ 同 じ結 な わ ちFH-P24と 同 じ防 御 効 果 を 得 るた め に は,両 者 と も10倍 濃 度 の 抗 血 清 を 接 種 しな けれ ぽ な ら な か った.病 原 株 攻 撃 時 に お い て 生 菌 免 疫 に よ る血 清 中 のIgG値 よ り高 い 値 を 示 す 免 疫 状 態 で あ っ て もわ ず か に20%の と どま った.こ 防御効果 に の こ とか ら も,た ん にIgG値 が高 よ そ85KDaに 菌 免 疫 に よ るP24-S1と 生 菌 免 疫 の 実 験 の 結 果 か ら細 胞 性 免 疫 よ り も む し ろlgGが た が,防 重 要 で あ る と 考 え られ,受 御 効 果 とIgG値 が平行 す るものでは ない に よ る結 果 の み で 判 定 した もの で あ るた め他 の 検 定 法 で は異 な る結 果 が得 られ る可 能 性 も否 定 で き な い. BALB/cマ ウ ス はM.pneumoniaeに 変 は お こ さ な い た め,感 指 標 と し て 判 定 し た が,再 度,本 関 わ る 因 子 を 追 求 し て ゆ き た い. 平 成 元年7月20日 proteinと よぼ れ 1) Sobeslaysky, Adsorption aminic acid 病 変 も合 せ 白 を は じめ とし て感 染 防 御 に 文 告 が あ る.な か で もattachment 菌 に対 す る感 受 性 動 物 で あ る ハ ム ス タ ー を 用 い て,肺 性 が 再 確 認 され た. 病 原 因 子 等 に関 与 す る も の と して 非 常 に 多 くの報 よ り肺 病 染 防 御 を菌 の 分 離 の み を IgGに 蛋 白 につ い て は,付 着 因 子, 身 免疫を し と い う結 果 と な っ た.細 胞 性 免 疫 に つ い て はMIT て 検 討 し,85KDa蛋 M.pneumoniaeの 対 す 間 の 差 を 生 じ させ る原 因 の一 つ で あ る い こ とだ け が 感 染 防 御 と一 致 す る の で は な く, 対 応 す る抗 原 の違 い を 検 討 す る こ との 重 要 は強 く るbandがP24-S11に 重 要 で あ ろ う と考 え られ る.そ こで,IgG抗 た 結 果,FH-P24の 株 とP24-S1で 反 応 し て い るbandの P24-S11の 体価 の 実験で トの マ イ コ プ ラ ズ マ 肺 炎 患 者 と のWestem た 感 染 防 御 で は,細 胞 性 免 疫 よ りは む しろIgGが 高 い抗 血 清 で 受 身 免 疫 した マ ウ ス を 同様 に実 験 し, a: M. P24-S11, 献 0., Prescott, B. & Chanock, of Mycoplasma receptors pneumoniae of various R.M.: to neur- cells and
7 possible role in virulence. J. Bacteriol., 96: 695 2) Collier, A.M. & Clyde, W.A. Jr.: Relationship between Mycoplasma pneumoniae and human respiratory epithelium. Infect. Immun., 3: 694 3) Powell, O. A., Hu, P. C., Wilson, R.M., Collier, A. M. & Baseman, J.B. Attachment of Mycoplasma pneumoniae to respiratory epithelium. Infect. Immun., 13: , ) Gabridge, M. G. & Taylor-Robinson, D.: Interaction of Mycoplasma pneumoniae with human lung fibroblasts: Characterization of the in vitro model. Infect. Immun., 25: , ) Cohen, G. & Sommerson, N. L.: Mycoplasma polyacrylamide gels to nitrocellulose sheets. Procedure and some applications. Proc. Natl. Acad. Sci. U. S. A., 76: , ) Hu, P. C., Huang, C., Collier, A. M. & Clyde W. A. Jr.: Demonstration of antibodies to Myco- pneumoniae: Hydrogen peroxide secretion and plasma pneumoniae attachment protein in its possible role in virulence. Ann. NY Acad. Sci., 143: 85-87, human sera and respiratory Immun., 41: , secretions. Infect. 17) Brunner H., Feldner J. & Bredt W.: Effect of monoclonal antibodies to the attachment-tip on experimental Mycoplasma pneumoniae infection of hamsters: A prelinimary report. Isr. J. 7) Yayoshi, M.: Association between Mycoplasma Med. Sci., 20: , pneumoniae hemolysis, attachment, and pulmonary 18) Jacobs, E., Drews, M., Stuhlert, A., Buttner, C., pathogenicity. Yale J. Biol. Med., 56: 685 Klein, P. J., Kist, M. & Bredt, W.: Immuno- logical reaction of guinea-pigs following 8) Yayoshi, M., Araake, M., Hayatsu, E., Kawakubo, intranasal Mycoplasma pneumoniae infection Y. & Yoshioka, M.: Characterization and Immunization with the 168 KDa adherence and pathogenicity of hemolysis mutants of Mycoplasma pneumoniae. Microbiol. Immonol., 28: , ) Yayoshi, M., Araake, M., Hayatsu, E., Takezawa, T. & Yoshioka, M.: Immunogenicity and protective effect of hemolysis mutants of Mycoplasma pneumoniae. Microbiol. Immunol., 29: , Holt, S. C.: Identification of a 32-Kilodalton protein of Mycoplasma pneumoniae associated with hemadsorption. Isr. J. Med. Sci., 23: ) Hayatsu, E.: Acquired immunity to Myco- 21) Vu, A. C., Foy, H. M., Cartwright, F. D. & Kenny, G. E.: The principal protein antigens of isolates of Mycoplasma pneumoniae as measured plasma pneumoniae pneumonia in hamsters. Microbiol. Immunol., 22: , ) Voller, A., Bidwell, D. & Bartlett, A.: In Mannual of clinical immunology (Rose, N.R. & Fiedman, M. ed.) p.506. Am. Soc. Microbiol, Washington D. C ) Harrington, J. T. Jr. & Stastny, P.: Macrophage migration from an agarose droplet: Development of a micromethod for assay of delayed hypersensitivity. J. Immunol., 110: ) Laemmli, U. K.: Cleavage of stractural proteins during the assenbly of the head of bacteriophage T4. Nature, 227: , ) Towbin, H., Staehlin, T. & Gordon, J.: Electrophoretic transfer of proteins from protein. J. Gen. Microbiol., 134: , ) Madsen, D. R., Weiner, L. B., Mcmillan, J. A., Saeed, F. A., North, J. A. & Coates, S. R.: Direct detection of Mycoplasma pneumoniae antigen in clinical specimens by a monoclonal antibody immunoblot assay. Am. J. Clin. Pathol., 89: 95-99, ) Baseman, J. B., Morrison-plummer, J., Drouillard, D., Puleo-Scheppke, B., Tryon, V. V. & by levels of immunoglobulin G in human serum are stable in strains collected over a 10-year period. Infect. Immun., 55: , 1987.
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2 94
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