血液PEA检测在鉴别PA感染与定植中的作用

Study of the role of blood PEA detection in identification of PA infection and plantation

目的：探讨大鼠血液铜绿假单胞菌（pseudomonas aeruginosa，PA）外毒素（paeruginosa exotoxin A，PEA）检测在PA感染与定植鉴别中的价值。方法：8周龄雄性SD大鼠180只，经地塞米松抑制免疫预处理后，随机分为感染组（气管内注入0.4 ml铜绿假单胞菌混悬液）、定植组（经咽部滴入铜绿假单胞菌混悬液0.2 ml）、对照组（气管内注入0.4 ml生理盐水）。各组按处死时间不同，再分为3 D组、7 D组、11 D组，分别于接种后第3、7、11 d随机抽取各组20只大鼠，心脏采血法处死，无菌取出肺组织并抽取血样。左肺组织匀浆行铜绿假单胞菌培养并计数菌落；右肺组织HE染色，镜下观察肺组织结构；收集血液分别以实时荧光定量PCR法检测PEA DNA及ELISA法检测IL-6、IL-8、IFN-γ、TNF-α。结果：定植组、对照组大鼠各时间点肺组织均未见PA菌生长及明显炎症反应，定植组各时间点咽拭子培养见PA菌生长（其菌落计数均＞105 cfu/ml），感染组大鼠肺组织匀浆各时间点均可见PA菌生长（其菌落计数均＞104 cfu/ml）及明显炎症反应，其中以第3 d最为明显，至第7、11天有所下降。各时间点感染组大鼠血液PEA DNA及IL-6、IL-8、IFN-γ、TNF-α表达均有升高，与定植组及对照组表达差异均有统计学意义（P＜0.05），各时间点定植组及对照组大鼠血液PEA DNA及IL-6、IL-8、IFN-γ、TNF-α表达差异无统计学意义（P〉0.05），其中感染组大鼠血液PEA DNA及IL-6、IL-8、IFN-γ、TNF-α表达检测以第3 d最为显著。结论：血液PEA DNA检测对鉴别铜绿假单胞菌感染与定植有一定价值，其表达水平高低与大鼠肺组织PA菌菌落计数及肺组织炎症反应程度正相关，推测PEA DNA可能反映肺部感染程度及其预后。

Objective：To investigate the value of detection of blood pseudomonas aeruginosa exotoxin （PEA） in the identification of PA infection and PA plantation in rats. Method：180 male eight weeks old SD rats,after immunoinhibition by dexamethasone were randomly divided into the infection group [0.4 ml of suspension of pseudomonas aeruginosa （PA） injected into trachea], the plantation group （0.2 ml of PA instilled into pharynx） and the control group （0.4 ml of physiological saline injected into trachea） , and then divided again into 3 group, 7 d group and 11d group, 20 rats each animals were sacrificed on 3 d, 7 d and 11 d, and then the samples of pulmonary tissues and blood were collected aseptically. The PA in homogenates of the left lung were cultivated and then the bacterial colonies were counted . The right lung section was observed by optical microscope after HE staining . The blood PEA DNA and the IL-6 , IL-8 , IFN-γ, and TNF-αwere detected through the Real-time PCR and ELISA respectively. Result：The pulmonary tissue of the plantation group and control group did have not bacterial growth and inflammation response at each time point , bacterial growth （the colony counting more than 105 cfu/ml） was boserved in the Culture of Pharyngeal Swab of the plantation group of bacteria growth and the inflammatory response in pulmonary homogenate in the 3＆amp;nbsp;infection groups （the colony counting more than 104 cfu/ml）,and most notabe on 3 d, declined at 7 d and 11 d. The expression of PEA DNA , IL-6 , IL-8 , IFN-γand TNF-αin blood of the infection group were higher than those of the control group and the plantation group （P〈0.05） . And there was no significant difference between the plantation group and the control group in expression of PEA DNA, IL-6 , IL-8 , IFN-γand TNF-αin blood （P〉 0.05）. And the changes in the infection group were the most notable at 3d. Conclusion：There is some value of blood PEA in identification of PA infection and plantation , and there is positive correlation between the exprssion and the colony counting or inflammatory severity. There fore,PEA,DNA may reflects the pulmonary degree of infection and the prognosis .