新型亚苄基环烷烃酮类光敏剂介导的光动力疗法对耐甲氧西林金黄色葡萄球菌生物被膜的破坏作用

Novel Benzylidene Cyclanone Photosensitizer Mediated Photodynamic Damage to Methicillin-Resistant Staphylococcus Aureus Biofilm

目的观察新型亚苄基环烷烃酮类光敏剂(cationic benzylidene cyclopentanone photosensitizers,P2)介导的光动力疗法(photodynamic therapy,PDT)对耐甲氧西林金黄色葡萄球菌(methicillin-resistant staphylococcus aureus,MRSA)生物被膜内细菌的灭活作用,并且观察PDT对生物被膜结构和多糖成分的破坏作用。方法采用不同浓度(0、10和25μM)的光敏剂P2对临床菌株MRSA0的生物被膜进行PDT处理,通过XTT法检测生物被膜内细菌的存活率。采用SYTO9、PI荧光探针及FITC-Cona荧光探针结合激光共聚焦显微镜,观察P2光敏剂浓度为10μM时,经PDT处理后生物被膜内细菌杀伤情况、以及对生物被膜的结构及多糖成分的影响。结果 PDT处理后,光敏剂浓度为10μM组和25μM组,经XTT法检测的细菌存活率分别为15.77%和6.25%。激光共聚焦显微镜观察结果显示,PDT组SYTO9探针的绿色荧光较空白对照组明显减弱;PI探针的红色荧光较空白对照组明显增强,说明细菌存活数量明显低于空白对照组。FITC-Cona荧光探针的绿色荧光较空白对照组明显减弱,说明生物被膜内多糖成分遭到明显破坏。结论 P2-PDT对MRSA生物被膜内细菌有杀伤作用,其对生物被膜内细菌杀伤率随光敏剂浓度增大而提高,其机理可能为通过破坏生物被膜内的多糖成分,从而破坏生物被膜的结构。

Objective To study the destruction effect of the cationic benzylidene cyclopentanone photosensitizers（P2） mediated photodynamic therapy（PDT） on the antibiotic resistance of methicillin resistant staphylococcus aureus biofilms.Methods The P2 photosensitizers of different concentrations（0,10 and 25 μM） of photosensitizer P2 were used to treat MRSAO biofilm in the PDT process,and the bacteria survival rate was detected by with XTT.Confocal laser scanning microscopy（CLSM） was used to observe the fluorescence emitted in the biofilm which was incubationed with SYTO9 and PI.10 μM P2 was used to treated MRSAO biofilm in the process,and the fluorescence emitted in the biofilm which was incubationed with FITC-Cona was observed by with CLSM.Results The survival rate of bacteria in MRSA biofilm decreased with the increase of the P2 concentration of P2 increased.When the concentration of P2 is was 10 μM,the bacteria survival rate dropped to 15.77%,While the P2 concentration of P2 is being 25 μM,the bacteria survival rate dropped fell to 6.25%.CLSM showed less green fluorescence（live） in PDT than in the control,and more red fluorescence（dead） in the PDT than in the control group was observed with CLSM.And CLSM showed that less green fluorescence was observed in PDT than in the control.Conclusions In the range of 0-25μM,P2-PDT can will destroy the living bacteria in MRSA biofilm,and the bacteria killing rate increases rises with the increase of P2 concentration increasing of P2.