穿心莲内酯增强庆大霉素对小鼠烧伤创面感染的防治研究

Prevention and treatment of andrographolide combined with gentamicin for burn wound infection in mice

摘要:

目的：研究穿心莲内酯联合庆大霉素对烧伤创面感染的作用。方法：建立烧伤创面铜绿假单胞菌及金黄色葡萄球菌混合感染小鼠模型，取BALB/c成年小鼠共48只，每只小鼠做4个烫伤创面，分为对照组、穿心莲内酯干预组、庆大霉素干预组及联合干预组（各组创面48个）。病理切片观察组织坏死情况，ELISA检测肿瘤坏死因子（tumor necrosis factor-α，TNF-α）、白细胞介素-6（interleukin-6，IL-6）及白细胞介素-1β（interleukin-1β，IL-1β）水平，电镜观察生物膜情况；建立混合生物膜体外模型，Western blot检测上清及菌体中外毒素A（exotoxin A，ExoA）的表达；qPCR检测exoS及exoY表达水平。结果：电镜证实建模成功并显示各干预条件下生物膜破坏情况；病理评分显示，联合干预组（4.86±0.38）预后好于庆大霉素干预组（3.29±0.49）和穿心莲内酯干预组（4.14±0.69）（P=0.000，P=0.020）；ELISA示庆大霉素干预组、穿心莲内酯干预组、联合干预组与对照组比较IL-1β、IL-6和TNF-α表达量均降低（P<0.05）；Western blot示庆大霉素干预组上清及菌体中ExoA灰度值分别为43.73±1.07和73.53±1.90，均高于联合干预组的40.99±0.57和53.37±2.31（P=0.017，P=0.000）；qPCR显示庆大霉素组exoS与exoY表达水平高于联合干预组（P=0.003，P=0.011）。结论：穿心莲内酯可增强庆大霉素对烧伤组织的修复作用，其原因与穿心莲内酯促进庆大霉素对生物膜的破坏、炎症因子及毒力因子的减少有关。

Abstract:

Objective：To study the effect of andrographolide combined with gentamicin on burn wounds infection. Methods：The burn model infected by Pseudomonas aeruginosa mixed with Staphylococcus aureus was established in 48 BALB/c mice，with four wounds in each. The wounds in each mouse underwent blank intervention（control group），gentamicin intervention（gentamicin group），andro－grapholide intervention（andrographolide group） and andrographolide combined with gentamicin intervention（combined intervention group） respectively. The tissue necrosis was observed under pathological section，and the morphology of mixed biofilm by scanning electron microscopy. The expressions of tumor necrosis factor-α（TNF-α），interleukin-6（IL-6） and interleukin-1β（IL-1β） were de－tected by ELISA. The mixed biofilm model in vitro was established. The expression of exotoxin A（ExoA） was detected by Western blot and the expressions of exoS and exoY by qPCR. Results：Electron microscopy confirmed the successful modeling and biofilm de－struction under different interventions. The pathological scores showed that the prognosis of combined intervention group（4.86±0.38） was better than that of gentamicin intervention group（3.29±0.49） and andrographolide intervention group（4.14±0.69），with P value as 0.000 and 0.02，respectively. ELISA showed that the expres－sions of IL-1β，IL-6 and TNF-α in gentamicin intervention group decreased significantly than those in combined interven－tion group（P<0.05）. Western blot showed that the gray values of ExoA in the supernatant and bacterial cells of the gentamicin intervention group were 43.73±1.07 and 73.53±1.90 respec－tively，which were significantly higher than those in the com－bined intervention group（40.99±0.57 and 53.37±2.31）（P=0.017，P=0.000）. The expressions of exoS and exoY in the gentamicin intervention group were higher than those in the combined intervention group，with P values as 0.003 and 0.011. Conclusion：Andro－grapholide enhances the effect of gentamicin on the repair of burn wounds，which is related to the effects that andrographolide can promote the destruction of biofilm，and the reduction of inflammatory factors and virulence factors.