原薯蓣皂苷体外抗产NDM-1耐碳青霉烯Raoultella ornithinolytica B1645-1B的机制研究  

Mechanism of Protodioscin Against Carbapenem-resistant Raoultella Ornithinolytica B1645-1B Carrying NDM-1 in Vitro

作　　者：余春芳 严廷苇 妥娟 彭紫晶 张思涵 胡景秀 何浩 张涵肃 孟丽雪 YU Chun-fang;YAN Ting-wei;TUO Juan;PENG Zi-jing;ZHANG Si-han;HU Jing-xiu;HE hao;ZHANG Han-su;MENG Li-xue(School of Basic Medical Sciences,Hubei University of Medicine;Hubei Key Laboratory of Wudang Local Chinese Medicine Research;College of Pharmacy and Nursing,Hubei University of Medicine;First Clinical College,Hubei University of Medicine;Second Clinical College,Hubei University of Medicine,Shiyan,Hubei 442000,China)

机构地区：[1]湖北医药学院基础医学院 [2]武当特色中药研究湖北省重点实验室 [3]湖北医药学院药护学院 [4]湖北医药学院第一临床学院 [5]湖北医药学院第二临床学院,湖北十堰442000

出　　处：《湖北医药学院学报》2024年第5期463-469,共7页Journal of Hubei University of Medicine

基　　金：湖北省教育厅自然科学基金(2023AFB270);湖北省大学生创新创业训练计划项目(S202213249002);十堰市科技局项目(22Y23,22Y21)。

摘　　要：目的:研究原薯蓣皂苷(protodioscin,PD)对产碳青霉烯酶NDM-1解鸟氨酸乌拉尔菌(carbapenem-re⁃sistant Raoultella ornithinolytica)B1645-1B的体外抑菌作用及逆转耐药性机制研究。方法:采用微量肉汤稀释法测定PD对B1645-1B的最低抑菌浓度(minimum inhibitory concentration,MIC)。应用棋盘法测定中药单体PD联合亚胺培南(Imipenem,IMP)对B1645-1B的MIC,计算抑菌浓度指数(fractional inhibitory concentration,FIC)。采用实时荧光定量PCR(quantitative real-time PCR,qRT-PCR)检测PD作用菌株B1645-1B后NDM-1 mRNA合成。利用MOE(Molecular Operating Environment)软件预测PD与NDM-1蛋白分子对接。通过十二烷基硫酸钠聚丙烯酰胺凝胶电泳(sodium dodecyl sulfate-polyacrylamide gel electrophoresis,SDS-PAGE)检测PD作用纯化后菌株B1645-1B的NDM-1蛋白表达。结果:PD对B1645-1B的MIC为10000μg/mL。PD联合IMP对B1645-1B的FIC为0.75,出现协同作用。PD作用后菌株B1645-1B的NDM-1 mRNA合成抑制和酶活性降低。结论:PD可能是一种很有前途的新型有效的协同抗菌剂及NDM-1酶抑制剂。Objective To investigate the antibacterial effect of protodioscin(PD)against carbapenem-resistant Raoultella ornithinolytica B1645-1B carrying NDM-1 in vitro and its mechanism.Methods A broth microdilution method was used to determine the minimum inhibitory concentration(MIC)of PD against strain B1645-1B.The MIC of PD combined with imi⁃penem(IMP)against the strain was determined by the checkerboard method,and the fractional inhibitory concentration(FIC)index was calculated.The mRNA expression of NDM-1 in strain B1645-1B after treatment with PD was detected by quantitative real-time PCR(qRT-PCR).The Molecular Operating Environment(MOE)was used to predict the molecular docking between PD and NDM-1 protein.The protein expression of NDM-1 after treatment with PD and purification was determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis(SDS-PAGE).Results The MIC of PD against strain B1645-1B was 10000μg/mL,and PD combined with IMP had an FIC of 0.75,indicating synergy.After treatment with PD,the mRNA synthesis and enzyme activity of NDM-1 were inhibited.Conclusion PD was a promising novel and effective synergistic antimicrobial agent and NDM-1 enzyme inhibitor.

关 键 词：原薯蓣皂苷 耐碳青霉烯解鸟氨酸乌拉尔菌 NDM-1 亚胺培南 体外抑菌实验 

分 类 号：R285[医药卫生—中药学]