美罗培南负载微泡联合超声靶向破坏技术对大肠杆菌生物膜的作用研究  

Meropenem-loaded microbubbles combined with ultrasound-targeted disruption of Escherichia coli biofilms

作　　者：马有财 穆文博 姚粝芹 邢琼丹 曹力[2] 孙学斌[1] Ma Youcai;Mu Wenbo;Yao Liqin;Xing Qiongdan;Cao Li;Sun Xuebin(Department of Sports Medicine,the First Afiliated Hospital of Xinjiang Medical University,Urumqi 830011,China;Department of Joint Surgery,the First Affiliated Hospital of Xinjiang Medical University,Urumqi 830011,China;School of Pharmacy,Xinjiang Medical University,Urumqi 830054,China;Department of Cardiac Ultrasonography,the First Affiliated Hospital of Xinjiang Medical University,Urumqi 830011,China;Xinjiang Key Laboratory of Ultrasound Medicine,Department of Science and Technology,Xinjiang Uygur Autonomous Region,Urumqi 830054,China)

机构地区：[1]新疆医科大学第一附属医院运动医学科,乌鲁木齐830011 [2]新疆医科大学第一附属医院关节外科,乌鲁木齐830011 [3]新疆医科大学药学院,乌鲁木齐830054 [4]新疆医科大学第一附属医院心脏超声诊断科,乌鲁木齐830011 [5]新疆维吾尔自治区科技厅新疆超声医学重点实验室,乌鲁木齐830054

出　　处：《中华超声影像学杂志》2025年第3期247-255,共9页Chinese Journal of Ultrasonography

基　　金：国家自然科学基金地区基金金(82260435);天山第二批青年拔尖人才培养计划(2023TSYCQNTJ0003);教育部新疆高发病研究重点实验室(新疆医科大学)-重点项目(2023A01)。

摘　　要：目的:探究载美罗培南微泡(MEM-MBs)联合超声靶向微泡破坏(UTMD)技术对大肠杆菌及其生物膜的治疗效果及破坏作用。方法:采用薄膜水化法制备MEM-MBs,Zeta电位分析仪检测MEM-MBs表征,光学显微镜下进行形态学观察。构建大肠杆菌引起的假体周围关节感染(PJI)体外生物膜模型,SYTO59染色生物膜、DIL染色微泡后于激光共聚焦显微镜下观察生物膜形态及MEM-MBs在细菌生物膜内分布情况。采用随机数字表法将生物膜分为对照组、美罗培南(MEM)组、MEM-MBs组、UTMD组和MEM-MBs+UTMD组,每组样本量为12,分别给予相应干预后使用扫描电镜和激光共聚焦显微镜观察各处理组对大肠杆菌及其生物膜形态结构的影响,结晶紫染色后借助酶标仪测定和比较各处理组生物膜密度差异,激光共聚焦显微镜下观察各处理组生物膜厚度,激光共聚焦显微镜和涂板计数观察各处理组细菌活力的差异。结果:①构建了符合实验要求的MEM-MBs。②构建了肉眼和激光共聚焦显微镜下可见的致密的大肠杆菌生物膜,厚度(10.61±0.17)μm。③使用激光共聚焦显微镜观察到MEM-MBs可渗透入生物膜各层。④结晶紫染色结果显示对照组、MEM组、MEM-MBs组、UTMD组和MEM-MBs+UTMD组生物膜密度依次降低,MEM组与MEM-MBs组比较差异无统计学意义(P>0.05),其余组间两两比较差异有统计学意义(均P<0.05)。⑤UTMD技术和MEM-MBs+UTMD可显著破坏大肠杆菌生物膜。激光共聚焦结果显示,与对照组相比,其余各组生物膜厚度降低,仅UTMD组与MEM-MBs+UTMD组孔隙率增加(均P<0.05);与MEM组和MEM-MBs组相比,UTMD组孔隙率增加,MEM-MBs+UTMD组生物膜厚度降低,孔隙率增加(均P<0.05);与UTMD组相比,MEM-MBs+UTMD组生物膜厚度降低,孔隙率增加(均P<0.05)。⑥涂板计数及激光共聚焦显微镜结果显示,与对照组相比,MEM组、MEM-MBs组与MEM-MBs+UTMD组菌落数降低,死菌比例升高(均P<0.05)。与MEM组和MEM-MObjective:To investigate the therapeutic efficacy and disruptive effects of Meropenem(MEM)-loaded microbubbles(MBs)combined with ultrasound targeted microbubble destruction(UTMD)technology on Escherichia coli and its biofilm.Methods:MEM-MBs were prepared using the thin-film hydration method,and their characterization was assessed using a Zeta potential analyzer,with morphological observations conducted under an optical microscope.An in vitro biofilm model of periprosthetic joint infection(PJI)caused by Escherichia coli was constructed,and the morphology of the biofilm and the distribution of MEM-MBs in the bacterial biofilm were observed under a laser confocal microscope after staining the biofilm with SYTO59 staining and DIL staining for Microbubbles.The biofilm morphology and the distribution of MEM-MBs in bacterial biofilm were observed under laser confocal microscope.The biofilms were randomly divided into 5 groups using a random number table:control,Meropenem(MEM),MEM-MBs,UTMD,and MEM-MBs+UTMD,with 12 samples per group.After applying the respective interventions,scanning electron microscopy(SEM)and laser scanning confocal microscopy(LSCM)were employed to observe the effects on the morphology and structure of Escherichia coli and its biofilm.Crystal violet staining was utilized to determine and compare the biofilm density among groups using a microplate reader.LSCM was also used to observe the biofilm thickness,while both LSCM and spread plate counting were employed to assess bacterial viability differences across groups.Results:①MEM-MBs meeting the experimental requirements were successfully constructed.②A dense Escherichia coli biofilm visible under both the naked eye and LSCM was established,with a thickness of(10.61±0.17)μm and a proportion of dead bacteria within the biofilm of(16.8±0.8)%.③MEM-MBs were observed to penetrate into all layers of the biofilm using LSCM.④The results of crystal violet staining showed a decreasing trend in the biofilm density of the control group,the MEM group,the ME

关 键 词：大肠杆菌 生物膜 美罗培南 超声靶向微泡破坏技术 

分 类 号：R378[医药卫生—病原生物学]