隐丹参酮对铜绿假单胞菌的生长及生物膜形成的抑制作用  

Inhibitory Effect of Cryptotanshinone on Biofilm Formation of Pseudomonas aeruginosa

作　　者：马书恒 缪林坪 陈荻秋 韩丽[2,3] 张富凯 贺气志 MA Shuheng;MIAO Linping;CHEN Diqiu;HAN Li;ZHANG Fukai;HE Qizhi(Changsha Medical University a.The First Clinical College,Changsha 410219,China;School of Basic Medical Science,Changsha 410219,China;Hunan Provincial Key Laboratory of the Research and Development of Novel Pharmaceutical Preparations,Changsha 410219,China)

机构地区：[1]长沙医学院第一临床学院,长沙410219 [2]长沙医学院基础医学院,长沙410219 [3]新型药物制剂研发湖南省重点实验室,长沙410219

出　　处：《激光生物学报》2024年第1期90-96,共7页Acta Laser Biology Sinica

基　　金：国家自然科学基金青年项目(82100040);湖南省教育厅科学研究项目(湘教通〔2022〕323号-22B0899);湖南省大学生创新创业训练项目(湘教通〔2021〕197号-3905);湖南省大学生创新创业训练计划重点支持领域项目(湘教通〔2023〕237号-38);长沙医学院大学生创新创业训练项目(长医教〔2023〕51号-108)。

摘　　要：为研究隐丹参酮(cryptotanshinone)对铜绿假单胞菌(Pseudomonas aeruginosa)的生长和生物膜形成的影响及其机制,采用微量稀释法测定隐丹参酮对铜绿假单胞菌的最小抑菌质量浓度(MIC),利用结晶紫染色法、倒置显微镜和激光扫描共聚焦显微镜观察隐丹参酮对铜绿假单胞菌生物膜形成的影响,再采用实时荧光定量聚合酶链反应(RT-qPCR)检测隐丹参酮对铜绿假单胞菌生物膜形成相关基因LasI、LasR、RhlI和RhlR的表达的影响。隐丹参酮对铜绿假单胞菌的MIC为800.0μg/mL;结晶紫染色后,隐丹参酮的质量浓度为800.0μg/mL时抑制菌膜效果最佳,且呈剂量依赖性(P<0.01);利用激光共聚焦显微镜观察隐丹参酮对生物膜的形成和分布,200.0μg/mL时抑菌效果显著提升。在200.0~800.0μg/mL的质量浓度范围内,800.0μg/mL的隐丹参酮对LasI、LasR、RhlI和RhlR基因的表达具有抑制作用(P<0.01)。隐丹参酮抑制铜绿假单胞菌生物膜形成机制可能是通过下调LasI等基因的表达。本研究表明,隐丹参酮可以作为群体感应抑制剂干预铜绿假单胞菌的耐药,为临床多重耐药铜绿假单胞菌(MDR-PA)的治疗提供参考。To investigate the effects of cryptotanshinone on the growth and biofilm formation of Pseudomonas aeruginosa and its mechanism,the minimum inhibitory concentration(MIC)of cryptotanshinone on P.aeruginosa was determined by microdilution,and the effect of cryptotanshinone on P.aeruginosa biofilm formation was observed by crystal violet staining,inverted microscope and laser scanning confocal microscope,and the effect of cryptotanshinone on P.aeruginosa biofilm formation was measured by real-time fluorescence quantitative polymerase chain reaction(RT-qPCR).The effect of cryptotanshinone on the expression of genes related to P.aeruginosa biofilm formation was detected by RT-qPCR.The MIC of cryptotanshinone on P.aeruginosa was 800.0μg/mL;after crystal violet staining,the best inhibition of biofilm was observed at the mass concentration of cryptotanshinone of 800.0μg/mL in a dose-dependent manner(P<0.01).The formation and distribution of biofilm by cryptotanshinone were observed by confocal laser microscopy,and the inhibition effect was significantly enhanced at 200.0μg/mL.In the range of 200.0~800.0μg/mL mass concentration,cryptotanshinone at 800.0μg/mL mass concentration inhibited the expression of LasI,LasR,RhlI and RhlR genes(P<0.01).The mechanism of cryptotanshinone inhibition of P.aeruginosa biofilm formation may be through the down-regulation of LasI and other genes expression.The present study suggests that cryptotanshinone can be used as a group-sensing inhibitor to intervene in the drug resistance of P.aeruginosa,which can provide a reference for the treatment of multi-drug-resistant P.aeruginosa(MDR-PA)in the clinic.

关 键 词：铜绿假单胞菌 生物膜 隐丹参酮 群感系统 耐药性 

分 类 号：R378.991[医药卫生—病原生物学]