Effect Of Curcumin-mediated Photodynamic Therapy On Methicillin-resistant Staphylococcus Aureus And Biofilm In Vitro

Objective:To investigate the effect of curcumin-PDT on MRSA and its biofilm structure in vitro.Methods:1.The curcumin was dissolved in 0.5% dimethyl Sulphoxide(DMSO)and formulated with different concentrations of solution(1,6.25,12.5,25 ?M).SOSG was used to test the generation of ROS induced by curcumin-PDT.2.Both bacterial suspension and MRSA biofilm were divided into 4 groups respectively: the Curcumin group,Irradiation group(PDT group),Curcumin-PDT group and the Blank control group.The concentration of curcumin in both curcumin group and irradiation group was 1,6.25,12.5,25 ?M.(1)The Curcumin group was treated with different concentration of curcumin at 37? for 30 min.(2)The Irradiation group was irradiated by 450 nm wavelength light source for 5 min(irradiance 1 W/cm squared).(3)The Curcumin-PDT group was pre-incubated with different concentration of curcumin at37? for 30 min,and then irradiated by 450 nm wavelength light source for 5 min(irradiance 1 W/cm squared).(4)the Blank control group was only treated with DMSO.The survival rates of bacteria within biofilm structure were measured by MTT assay.The optical microscope was used to observe and identify the changes of the structure of MRSA biofilm after crystal violet staining.3.Mouse osteoblasts were also divided into the Curcumin group,Irradiation group(PDT group),Curcumin-PDT group and the Blank control group,but the concentration of curcumin was 25 ?M.The Cytotoxicity induced by curcumin-PDT was tested by the optical microscope and MTT assay.Results:1.SOSG results showed that the ROS was produced in a concentration-dependent manner after treated with Curcumin-PDT.The generated ROS was increased with time,and tended to be stable in 5 min when the concentration of curcumin was constant.2.There was a remarkable killing effect of curcumin-PDT on MRSA in bothplanktonic bacteria and bacteria inside the biofilm culture,and the effect was increased in a curcumin concentration-dependent manner.The survival rate of MRSA in both planktonic bacteria and bacteria inside the biofilm culture was decreased from(95.62±1.24)%(P<0.05)and(96.60±0.89)%(P<0.05)respectively(the concentration of curcumin was 1?M)to(0.38±0.32)%(P<0.05)and(34.50±1.44)%(P<0.05)(the concentration of curcumin was 25?M).Meanwhile,the optical microscope showed that the biofilm culture was disrupted obviously in 25?M curcumin-PDT.There were no significant differences when compared to the Curcumin group,Irradiation group and Control group.3.The optical microscope showed that the growth of mouse osteoblasts in all groups was good and showed long spindle or irregular shape.There were no significant differences in morphology and density when compared to the Control group.MTT results showed that all the four groups were RGR "0" or "1" grade,and no cytotoxicity of curcumin-mediated PDT was detected.Conclusion:Our study demonstrated that curcumin-PDT has a killing effect on both MRSA planktonic bacteria and the bacteria inside the biofilm culture.The MRSA planktonic bacteria were more sensitive to curcumin-PDT than the bacteria inside the biofilm culture.Moreover,No cytotoxicity of curcumin on osteoblasts was detected in cytotoxicity test.Curcumin-mediated PDT provides a new drug choice and perspective for the treatment of MRSA-type PJI.