芪风固表颗粒对肺炎克雷伯菌感染的预防和抑制作用研究

Inhibitory Effect of Qifeng Gubiao Granules on Klebsiella Pneumoniae Infection

目的:研究芪风固表颗粒对肺炎克雷伯菌(Kp)感染小鼠的预防和抑制作用。方法:BALB/C小鼠,预防给予芪风固表颗粒高(2.6 g/kg)、中(1.3 g/kg)、低(0.65 g/kg)剂量40 d后,乙醚麻醉小鼠,滴鼻感染5×10^(8)CFU/mL的Kp50μL1次,感染24 h后处死小鼠。HE染色观察给药后小鼠肺组织结构病理改变;肺泡灌洗观察不同组小鼠肺泡灌洗液内白细胞数;小鼠肺组织细菌浓度测定观察各组肺组织中细菌浓度;免疫组织化学法观察Pannexin1/P2RX7在肺组织中的表达位置;蛋白免疫印迹法检测肺组织Pannexin1/P2RX7/Caspase-11蛋白表达变化;Real-Time PCR法观察肺组织Pannexin1/P2RX7/Caspase-11 mRNA表达变化。结果:与模型组比较,HE染色结果显示,芪风固表组的肺组织病理结构有所改善,仅出现少量充血以及少量炎细胞浸润,肺组织损伤程度降低;给药组小鼠肺泡灌洗液中白细胞数降低(P<0.05,P<0.01);小鼠肺组织细菌浓度测定结果显示,芪风固表颗粒高、中剂量组较模型组细菌浓度数明显减少(P<0.05);免疫组化结果发现,芪风固表各剂量组Pannexin1、P2RX7蛋白表达降低(P<0.05,P<0.01);蛋白印迹结果显示,芪风固表高剂量组Pannexin1、P2RX7、Caspase-11和中剂量组P2RX7的表达降低(P<0.05);Real-Time PCR结果显示,芪风固表高剂量组及中剂量组Pannexin1、P2RX7、Caspase-11 mRNA表达降低(P<0.05)。结论:芪风固表颗粒可以对肺炎克雷伯菌感染引起的肺部炎性改变及损伤有明显的改善作用,机制可能与抑制Caspase-11细胞焦亡通路信号有关。

Objective:To study the preventive and inhibitory effects of Qifeng Gubiao granules on Klebsiella pneumoniae(Kp)infection in mice.Methods:BALB/C mice were given high(2.6g/kg),medium(1.3g/kg)and low(0.65g/kg)doses of Qifeng Gubiao granules for 40 days.The mice were anesthetized with ether and infected with 5×Kp50 of 10^(8)CFU/mLμL1 times,the mice were killed 24 hours after infection.HE staining was used to observe the pathological changes of lung tissue structure in mice after administration;The number of leukocytes in alveolar lavage fluid of mice in different groups was observed;The concentration of bacteria in lung tissue of mice was measured;The expression of Pannexin1/P2RX7 in lung tissue was observed by immunohistochemistry;The expression of Pannexin1/P2RX7/Caspase-11 protein in lung tissue was detected by Western blot;Real Time PCR was used to observe the expression of Pannexin1/P2RX7/Caspase-11 mRNA in lung tissue.Results:Compared with the model group,HE staining showed that the pathological structure of the lung tissue in the Qifeng Gubiao group was improved,only a small amount of congestion and inflammatory cell infiltration appeared,and the degree of lung tissue damage was reduced;The number of leukocytes in alveolar lavage fluid of mice in the treatment group decreased(P0.05,P0.01);The concentration of bacteria in the lung tissue of mice showed that the concentration of bacteria in the high and middle dosage groups of Qifeng Gubiao granules was significantly lower than that in the model group(P0.05);Immunohistochemical results showed that the expression of Pannexin1 and P2RX7 protein in each dose group of Qifenggubiao decreased(P0.05,P0.01);Western blot showed that the expression of Pannexin 1,P2RX7,Caspase-11 in high dose group and P2RX7 in middle dose group decreased(P0.05);Real Time PCR results showed that the mRNA expressions of Pannexin1,P2RX7 and Caspase-11 in the high and middle dose groups of Qifeng Gubiao decreased(P0.05).Conclusion:Qifeng Gubiao Granule can significantly improve the inflammatory changes and damage of lungs caused by Klebsiella pneumoniae infection,and the mechanism may be related to the inhibition of Caspase-11 cell scorch pathway signal.