PM2.5联合中波紫外线处理对HaCaT细胞的影响

Effects of PM2.5 combined with UVB treatment on human keratinocytes HaCaT

目的研究PM2.5联合中波紫外线(UVB)处理对人皮肤角质形成细胞Ha Ca T的影响。方法收集环境中PM2.5并进行其金属成分分析;MTT法测定PM2.5对Ha Ca T细胞的半数抑制浓度(IC50);将细胞分为空白对照组(NC组)、单纯IC50浓度的PM2.5处理组(PM2.5组)、单纯30 m J/cm2的UVB照射组(UVB组)和PM2.5联合UVB处理组(联合处理组),MTT法检测不同处理组对细胞活力的抑制;流式细胞术检测不同处理对细胞凋亡的影响;Western blot检测不同处理后PARP、LC3的蛋白表达变化。结果 PM2.5样品中金属成分有Ca、Ba、Al、Cu、Pb等;PM2.5对Ha Ca T细胞的IC50约为300μg/m L;不同处理组之间,在处理后24 h,对细胞活力的抑制差异显著(P<0.001),其中联合处理组细胞对比NC组活力最低(P<0.001);流式分析结果显示,与NC组比,PM2.5组、UVB组以及联合处理组细胞凋亡率增加(P<0.01),联合处理组凋亡率较PM2.5组增加(P<0.05),但低于UVB照射组(P<0.01);Western blot显示PM2.5组、UVB组以及联合处理组LC3-Ⅱ、PARP蛋白较NC组均有增加,联合处理组PARP较UVB组减少,LC3-Ⅱ较PM2.5组、UVB组增加。结论 PM2.5能增加UVB对Ha Ca T细胞的损伤,其主要可能通过增加细胞自噬,而非凋亡方式。

Objective To study the effect of PM2.5 combined with UVB treatment on human keratinocytesHaCaT. Methods The PM2.5 in the air was collected in Guangzhou and the metal ingredients were analyzed. Thecells were divided into four groups：negative control group（NC group）,simple PM2.5 treatment group with theconcentration of IC50（PM2.5 group）,simple UVB irradiation group with the dose of 30 mJ/cm^2（UVB group）and PM2.5 combined with UVB treatment group（combined treatment group）. The effects of different treatments on cellviability were measured by MTT assay and those of different treatments on apoptosis were detected by flowcytometry. The expressions of PARP and LC3 protein were detected by Western blot. Results The metal componentsin PM2.5 samples included Ca,Zn,Ba,Al,Cu,Pb,etc. After the treatment of PM2.5 on HaCaT cells,weconcluded that the IC50 was about 300 μg/mL. The inhibitory effect on cell viability after 24 h in different groupsshowed significant difference（P〈0.001） and the viability of the combined treatment group was the lowest（P〈0.001）. Flow cytometry analysis showed that compared with that of NC group,the apoptosis rate of PM2.5 group（P〈0.01）,UVB group（P〈0.01）and the combined treatment group（P〈0.01）increased,but theapoptosis rate in the combined treatment group was higher than that of PM2.5 group（P〈0.05）,but lower thanthat in UVB group（P〈0.01）. Western blot showed that the level of LC3-Ⅱ and PARP in another three groupswas higher than that of NC group;PARP in the combined treatment group was lower than that in UVB groupand LC3-Ⅱ increased compared with that in PM2.5 and UVB group. Conclusion PM2.5 can increase theharm of UVB on HaCaT cells and the main mechanism may be through increasing autophagy rather than apop-tosis.