抗坏血酸对钴纳米粒子及钴离子致成纤维细胞毒性的保护作用

摘要

目的 探讨抗坏血酸(AA)对降低钴(Co)诱导的成纤维细胞毒性作用.方法 将实验分为空白对照、Co2+、Co2++AA、钴纳米粒子(CoNPs)、CoNPs+ AA、AA 6个组.AA提前预处理1h.分别用CCK8检测CoNPs、Co2+诱导及AA处理后细胞毒性,采用荧光染色技术检测J线粒体中活性氧生成,Western blot检测相关蛋白表达,实时定量PCR检测相关分子mRNA水平.并检测细胞质中细胞色素C的水平.结果 当使用CoNPs和Co2+处理细胞后,细胞出现凋亡.CoNPs可以明显诱导活性氧(ROS)生成;促凋亡因子(Caspase 3及Bax)表达明显升高,而抗凋亡因子Bcl-2表达减少;细胞色素C及细胞凋亡诱导因子(AIF)均表达增加,并从线粒体进入细胞质中;使用AA预处理后,CoNPs所引起的这些改变可被减少.结论 AA可以通过减少ROS的生成与释放降低CoNPs导致的细胞毒性,但不能减弱Co2毒性作用.%Objective To explore the effect of L-ascorbic acid(AA) for reducing the fibroblast cells induced by cobalt(co)cytotoxicity.Methods The experiment was divided into the blank control group,Co2+ group,Co2++AA group,cobalt nanoparticles(CoNPs) group,CoNPs+AA group and AA group.The AA pretreated for 1 h was performed in advance.The CCK8 assay was used to detect the CoNPs and Co2+ induced and AA-treated cytotoxicity.The fluorescence staining was used to measure the production of ROS in mitochondria.The real-time polymerase chain reaction(PCR) and Western blot were used to measure the expression of related protein and related molecular mRNA respectively.Moreover the cytochrome-C level in cytoplasml was detected.Results After treating the cells by CoNPs and Co2+,the cells appeared apoptosis.CoNPs could significantly induce ROS generation;the proapoptotic factors(caspases-3,and Bax) were significantly increased,while the anti-apoptotic factor Bcl-2 expression was decreased;cytochrome C and AIF expressions were up-regulated and released from mitochondria into the cytoplasm,after AA pretreatment,these changes caused by CoNPs were decreased.Conclusion AA can reduce the CoNPs caused cytotoxicity by reducing ROS generation and release,but can not weaken the Co2+ toxic effect.