纳米二氧化硅对体外培养HaCaT细胞凋亡的影响

目的]比较不同粒径(15、30、100 nm)纳米二氧化硅(nano-SiO2)和常规二氧化硅(Micro-SiO2)对人皮肤表皮细胞(HaCaT)生长的抑制作用及凋亡的影响。方法]采用不同浓度(2.5、5、10μg/ml)不同粒径(15、30、100 nm)的nano-SiO2和10μg/ml的Micro-SiO2(1～5μm)染毒体外培养的HaCaT细胞24 h,同时设溶剂对照组(nano-SiO2和Micro-SiO2的分散液染毒组)。采用细胞计数试剂盒(CCK-8)法检测HaCaT细胞暴露于不同浓度、不同粒径的nano-SiO2后的细胞活力,用Annexinⅴ-PI双染法和Hoechest33342染色法测定细胞凋亡情况;使用2’,7’-二乙酰二氯荧光素荧光探针检测细胞内活性活性氧(ROS)水平。结果]不同粒径的nano-SiO2纳米对HaCaT细胞的半数抑制浓度分别为(19.4±1.3)、(27.7±1.5)、(35.9±1.6)μg/ml。当染毒浓度固定时,随着nano-SiO2粒径的减小,凋亡率逐渐增加。当粒径固定时,胞内ROS的水平也随着nano-SiO2浓度的增大而增高。相关分析表明,细胞存活率和凋亡率与胞内活性氧的水平的相关性分别为-0.952和0.898(P<0.01)。结论]nano-SiO2能抑制HaCaT细胞生长并可诱导其凋亡,这种现象的发生可能与胞内产生的活性氧水平有关。

Effects of Silicon Dioxide Nanoparticles on Apoptosis of HaCaT Cells

Objective] To investigate the effects of silicon dioxide nanoparticles（nano-SiO2） of different particle size（15,30,100 nm） and standard SiO2 particles（micro-SiO2） on proliferation and apoptosis of HaCaT cells.Method] HaCaT cells were treated by 15,30,100 nm nano-SiO2（2.5,5,10 μg/ml） particles and micro-SiO2（10 μg/ml） particles for 24 h,at the same time,the solvent control was set.The level of ROS and the effect of cell growth inhibition were assayed by 2′,7′-dichlorfluorescein-diacetate（DCFH-DA） and cell counting kit（CCK-8） Kit respectively.Apoptosis were assayed by Annexinⅴ-PI double staining and Hoechst 33342 staining method.Method] HaCaT cells were treated by 15,30,100 nm nano-SiO2（2.5,5,10 μg/ml） particles and micro-SiO2（10 μg/ml） particles for 24 h,at the same time,the solvent control was set.The level of ROS and the effect of cell growth inhibition were assayed by 2′,7′-dichlorfluorescein-diacetate（DCFH-DA） and cell counting kit（CCK-8） Kit respectively.Apoptosis were assayed by Annexinⅴ-PI double staining and Hoechest 33342 staining method.Result] As observed in the CCK-8 kit,exposure to nano-SiO2 nanoparticles for 24 h,the IC50 was 19.4 ± 1.3,27.7 ± 1.5 and 35.9±1.6 μg/ml for 15,30 and 100 nm SiO2 nanoparticles,respectively.Exposure to SiO2 nanoparticles results in a concentration-and size-dependent cytotoxicity and increase in intracellular ROS level and apoptosis rate were also observed in SiO2 nanoparticle-exposed HaCaT cells.Cytotoxicity and apoptosis in cultural HaCaT cells that is closely correlated to increased oxidative stress,the correlation coefficient was-0.952 and 0.898 respectively.Conclusion] nano-SiO2 may affect proliferation and apoptotic rates of HaCaT cells via stimulating the production of ROS in cells.