多黏菌素B对鲍曼不动杆菌脓毒症的疗效研究

Effect of Polymyxin B on Acinetobacter Baumannii Sepsis

用鲍曼不动杆菌活菌腹腔注射建立脓毒症感染模型。将造模成功的大鼠随机分为模型对照组(M组)、多黏菌素B组(PB组)、TAK-242组(TAK组)、多黏菌素B+TAK-242组(PB+TAK组),每组10只,两种药物均3mg·kg^(-1)静脉注射,另取10只正常大鼠作为空白对照组(C组),C组和M组静脉注射同剂量0.9%NaCl。给大鼠注射菌液后12h内观察一般指标,12h后进行细菌计数、检测血常规、HE染色观察组织器官,用酶联免疫吸附试验(ELISA)测定血清炎症指标,用实时荧光定量聚合酶链反应(RT-PCR)检测大鼠外周血单个核细胞(PBMC)TLR4mRNA,用蛋白免疫印迹法(WesternBlot)测定大鼠PBMC的NF-κBp65和磷酸化蛋白。结果表明:C组和M组比较、同一组内不同时间点和0h比较,6h后体温有差异;细菌计数组间比较除PB组和PB+TAK组无差异外,其他组间比较均有显著性差异(F=267.76,P<0.01);5组CRP,PCT,IL-6,TNF-α比较差异有显著性,两两比较C组和其他4组、M组和其他4组差异有显著性(P<0.01),PB+TAK组和PB组、TAK组比较CRP、IL-6有统计学意义,PB+TAK组和PB组比较TNF-α有统计学意义;各组大鼠TLR4mRNA表达水平比较差异有显著性(F=27.58,P<0.01),组间两两比较除PB组和TAK组无差异,其余组间比较差异均有显著性(P<0.05);各组大鼠NF-κBp65、p-NF-κBp65比较差异有显著性(F=34.32、61.59,P<0.01)。用药组均不同程度降低炎症因子、TLR4、目的蛋白的表达。多黏菌素B和TAK-242可以通过TLR4/NF-κB信号通路减轻鲍曼不动杆菌血液感染大鼠的炎症。

To investigate the effect of polymyxin B combined with TLR4 inhibitor TAK-242 on TLR4/NF-κB inflammatory pathway in Acinetobacter baumannii sepsis,sepsis infection model was established by intraperitoneal injection of live Acinetobacter baumannii.Building Wistar rats were randomly divided into model group(M),polymyxin B group(PB),TAK-242 group(TAK),polymyxin B+TAK-242 group(PB+TAK).There were 10 rats in each group.The injection dose of the two drugs is 3 mg·kg^(-1).Another 10 normal rats were selected as blank control group(C).Group C and M were given the same dose of 0.9%NaCl intravenously.The general index was observed within 12 h after the injection of bacterial solution.After 12 h,bacterial count,blood routine test,HE staining were performed to observe the tissues and organs of the rats.Serum inflammatory markers were determined by enzyme linked immunosorbent assay(ELISA),and TLR4 mRNA expression levels in peripheral blood mononuclear cells(PBMC)were detected by real-time fluorescence quantitative polymerase chain reaction(RT-PCR).The relative expression levels of NF-κBp65 and phosphorylated NF-κBp65 in rat PBMC were measured by Western blot.Results showed that there was statistically significant difference in body temperature between group C and group M after 6 h.Compared with 0 h at different time points in the same group,the differences in body temperature after 4 h were statistically significant.There were significant differences among the bacterial count groups except for PB and PB+TAK groups(F=267.76,P<0.01).There were significant differences in CRP,PCT,IL-6 and TNF-αamong the five groups(F=52.78,12.55,72.2,36.54,P<0.01),and there were significant differences between group C and the other four groups and between group M and the other four groups(P<0.01).CRP and IL-6 were statistically significant in PB+TAK group,PB and TAK group,and TNF-αwas statistically significant in PB+TAK group and PB group(P<0.05).There were significant differences in TLR4 mRNA expression levels among the groups(F=27.58,P<0.01),and there were significant differences among the other groups except for PB group and TAK group(P<0.05).The relative expression levels of NF-κBp65 and p-NF-κBp65 were significantly different among the groups(F=34.32,61.59,P<0.01).The expressions of inflammatory factors,TLR4 and target protein were decreased in different degrees in the treatment group.Polymyxin B and TAK-242 can reduce inflammation in Acinetobacter baumannii infected rats through TLR4/NF-κB signaling pathway.