吲哚菁绿光热治疗耐甲氧西林葡萄球菌感染的研究

目的　探讨吲哚菁绿（ICG）的光热治疗（PTT）性能及对耐甲氧西林葡萄球菌的杀灭效果。方法　利用紫外和荧光光谱对ICG的光物理性质进行表征；测定不同质量浓度ICG（0、12.5、25、50、100和200 mg/L）的光热升温曲线，评估ICG的光热转化性能；取小鼠乳腺癌4T1细胞与不同质量浓度的ICG溶液（0、50、100、200和500 mg/L）混合，通过MTT实验检测ICG的细胞毒性；取1×1010 CFU/mL的耐甲氧西林的金黄色葡萄球菌加入到96孔板中，将实验分为7组：PBS组、ICG组（500 mg/L）、激光照射组（1.5 W/cm2的808 nm激光照射10 min）、ICG+0.33 W/cm2激光照射组、ICG+0.66 W/cm2激光照射组、ICG+1 W/cm2激光照射组、ICG+1.5 W/cm2激光照射组，观察并记录存活的菌落数，分析基于ICG的PTT杀菌效果。结果　紫外吸收光谱证实ICG在808 nm处具有较强的光吸收能力；ICG溶液（质量浓度为200 mg/L）经功率密度为1.5 W/cm2的808 nm近红外激光照射10 min后，温度可升高23.3 ℃，而纯水的温度仅升高了4.8 ℃。ICG不同质量浓度组即50、100、200、500 mg/L组的细胞生存率分别为81.87%±2.69%、81.47%±2.94%、82.92%±2.61%、80.44%±2.98%，组间差异无统计学意义（F=0.402，P＞0.05）。不同功率密度激光照射组即0.33、0.66、1、1.5 W/cm2组的细胞生存率分别为97.21%±1.59%、94.92%±1.98%、93.89%±1.38%、93.03%±2.31%，组间差异无统计学意义（F=2.864，P＞0.05）。杀菌实验中，与PBS组比较，ICG+激光照射组随着激光功率密度的升高，细菌存活数量逐渐下降，差异有统计学意义（F=133.800，P＜0.01）。结论　ICG具有良好的生物相容性和光热治疗性能，在近红外激光照射下对耐甲氧西林葡萄球菌具有较强的杀灭效果，具有较好的临床转化应用价值。

Experimental study of indocyanine green photothermal in the treatment of methicillin-resistant Staphylococcus Aureus infection

Objective　To investigate the sterilizing effect of photothermal therapy (PTT) based on indocyanine green (ICG) on methicillin-resistant Staphylococcus Aureus. Methods　Ultraviolet and fluorescence spectra were used to characterize the photophysical property of ICG. The photothermal heating curves of ICG with different concentrations (0, 12.5, 25, 50, 100 and 200 mg/L) were determined to evaluate the photothermal conversion performance of ICG. Mouse breast cancer 4T1 cells were mixed with different concentrations of ICG solution (0, 50, 100, 200 and 500 mg/L), and the cytotoxicity of ICG was measured by MTT assay. The concentration of methicillin-resistant Staphylococcus Aureus was 1×1010 CFU/mL, and the experiment was divided into the 7 groups: the PBS group, the ICG group (500 mg/L), the laser irradiation group (808 nm laser irradiation of 1.5 W/cm2 for 10 min), the ICG+0.33 W /cm2 laser irradiation group, the ICG+0.66 W/cm2 laser irradiation group, the ICG+1 W/cm2 laser irradiation group and the ICG+ 1.5 W/cm2 laser irradiation group. The number of surviving colonies was calculated and the sterilizing effect of ICG-based PTT was analyzed. Results The ultraviolet absorption curve showed that ICG had strong absorption band at 808 nm. The temperature of ICG solution (200 mg/L) can be increased by 23.3 ℃ after irradiation by 808 nm near-infrared laser with 1.5 W/cm2 for 10 min, while the temperature of pure water only increased by only 4.8 ℃ at the same experimental condition. The relative survival rates of ICG groups with different concentrations (50 mg/L, 100 mg/L, 200 mg/L and 500 mg/L) were 81.87%±2.69%, 81.47%±2.94%,82.92%±2.61% and 80.44%±2.98%, respectively. There were no significant differences between the groups (F=0.402, P＞0.05). The survival rates of different laser irradiation groups (0.33, 0.66, 1 and 1.5 W/cm2) were 94.92%±1.98%, 93.89%±1.38% and 93.03%±2.31% respectively, and there was no significant difference between the groups (F=2.864, P＞0.05). In the sterilization experiment, compared with the PBS group, the number of surviving bacteria significantly decreased gradually with the increase of the laser power density in the ICG+ laser irradiation group (F=133.800, P＜0.01). Conclusion　ICG has a great biocompatibility and an excellent photothermal sterilizing effect under near-infrared laser irradiation to methicillin-resistant Staphylococcus Aureus, which exhibits a potential clinical application.