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目的 了解百日咳鲍特菌Ⅰ相菌株外膜蛋白 (outermembraneproteins,OMP)生物学活性与IgE介导哮喘的关系。方法 采用去垢剂处理、DEAE 5 2和SephadexG 15 0层析法制备百日咳鲍特菌Ⅰ相菌株OMP及其组分 ,以SDS PAGE估计OMP组分的相对分子质量 (Mr)。采用ELISA检测百日咳全菌死疫苗诱导动物产生的总IgE和卵白蛋白 (ovalbumin ,OVA)特异的IgE量。采用肥大细胞脱颗粒试验观察OMP及其组分促进脱颗粒的作用、ELISA检测培养物上清中组胺水平。采用组胺敏感试验检测OMP及其组分增加小鼠对组胺致死敏感性的作用。结果 从百日咳鲍特菌OMP中分离获得Mr 为 (30、32、38和 6 9)× 10 3的 4个组分。 1～ 10 0 μg mlOMP及其组分均未显示凝集鲎变形细胞溶解物的活性。OVA和百日咳鲍特菌联合免疫豚鼠的血清总IgE和OVA特异性IgE较单用OVA免疫有明显增加 (t=2 .75～ 6 .5 9;P <0 .0 5～ 0 .0 1)。OMP及其Mr30× 10 3、Mr32× 10 3组分有明显的促豚鼠肥大细胞脱颗粒的作用 (t=4 .914～ 9.137,P <0 .0 1) ,但Mr38× 10 3、Mr6 9× 10 3组分无此活性。OMP及其Mr30× 10 3、Mr32× 10 3组分处理的上清中组胺水平明显高于阴性对照组 (t =5 .6 72～9.30 4 ,P <0 .0 1) ,但Mr38× 10 3、Mr6 9× 10 3组分无此活性。结论 百日咳?
Objective To investigate the relationship between the biological activity of outer membrane protein (OMP) of phase Ⅰ Bordetella pertussis and IgE-mediated asthma. Methods OMC and its fraction of B. pertussis phase Ⅰ strain were prepared by detergent treatment, DEAE 5 2 and SephadexG 15 0 chromatography. The relative molecular mass (Mr) of OMP fraction was estimated by SDS PAGE. Total IgE and ovalbumin (OVA) -specific IgE levels induced by whole-body pertussis vaccine were determined by ELISA. Mast cell degranulation test was used to observe the effect of OMP and its components on degranulation. ELISA was used to detect the histamine level in the culture supernatant. Histamine sensitivity test was used to detect the effect of OMP and its components on the lethality of histamine in mice. Results Four components with Mr of (30, 32, 38 and 69) × 10 3 were isolated from the B. pertussis OMP. 1 ~ 10 0 μg ml OMP and its components did not show the activity of agglutinated 鲎 alveolar cell lysate. The serum total IgE and OVA specific IgE of OVA combined with B. pertussis co-immunized guinea pigs were significantly higher than that of OVA alone (t = 2.75-6.59; P <0.05-0.01) . OMP and its Mr30 × 10 3, Mr32 × 10 3 components were significantly induced guinea pig mast cell degranulation (t = 4.1414 ~ 9.137, P <0.01), but Mr38 × 103, Mr6 9 × 10 3 components without this activity. OMP and its Mr30 × 103, Mr32 × 10 3 components of the supernatant histamine levels were significantly higher than the negative control group (t = 5.72 ~ 9.304, P <0.01), but Mr38 × 10 3, Mr6 9 × 10 3 components without this activity. Conclusion Pertussis?