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目的研究铜绿假单胞菌分泌物对中性粒细胞趋化功能的影响。方法用percoll密度梯度离心法分离小鼠骨髓中性粒细胞并用FACS检测其纯度;铜绿假单胞菌PAO1静置培养5 d后涡旋并离心收集其上清;通过TAXIScan-FL检测被膜上清PAO1 d5(PAO1菌株静置培养第5天)的趋化作用;不同浓度的f MLP所引起的中性粒细胞趋化运动及PAO1 d5上清稀释50倍后的趋化作用;PAO1 d5上清对f MLP所引起的中性粒细胞趋化运动的影响。结果分离的小鼠骨髓中性粒细胞纯度大于90%;TAXIScan-FL检测被膜上清有趋化作用,但是速度及方向上与f MLP及LTB4有明显差别(P<0.001);高浓度的f MLP对中性粒细胞的趋化作用减弱,PAO1 d5稀释50倍后趋化作用消失;PAO1 d5上清可以抑制f MLP所引起的中性粒细胞趋化运动。结论 PAO1d5被膜上清中存在抑制中性粒细胞趋化的物质。
Objective To study the effect of Pseudomonas aeruginosa secretion on neutrophil chemotaxis. Methods Murine bone marrow neutrophils were isolated by percoll density gradient centrifugation and detected by FACS. Pseudomonas aeruginosa PAO1 was cultured for 5 days and then vortexed and centrifuged to collect the supernatant. TAXIScan-FL was used to detect the membrane supernatant PAO1 d5 (PAO1 strain stationary culture day 5) chemotactic; different concentrations of f MLP induced neutrophil chemotaxis and PAO1 d5 supernatants diluted 50 times the chemotaxis; PAO1 d5 supernatant Effect of f MLP on Neutrophil Chemotaxis. Results The purity of bone marrow neutrophil isolated from mice was more than 90%. TAXIScan-FL showed chemotaxis in the supernatant of the membrane but there was a significant difference in speed and direction between fMLP and LTB4 (P <0.001) MLP chemotaxis to neutrophils weakened, PAO1 d5 50-fold diluted chemotaxis disappeared; PAO1 d5 supernatant can inhibit fMLP-induced neutrophil chemotactic motility. Conclusion PAO1d5 supernatants contain chemotactic substances that inhibit neutrophil chemotaxis.