二氧化硅对人肺腺癌A549细胞的毒性作用

Toxic Effects of Silica on Human Lung Adenocarcinoma Epithelial A549 Cells

目的探讨二氧化硅对人肺泡Ⅱ型上皮细胞来源的肺腺癌A549细胞的毒性作用。方法将A549细胞暴露于二氧化硅中24 h,观察二氧化硅对A549细胞内氧化应激指标、炎性细胞因子及非神经元型胆碱能系统关键分子的影响,采用化学比色法测定超氧化物歧化酶(SOD)及丙二醛(MDA)水平,采用酶联免疫吸附方法(ELISA)测定肿瘤坏死因子-α(TNF-α)及白细胞介素-6(IL-6)水平,使用改良Ellman法测定AChE酶活性,使用Real-time PCR法测定乙酰胆碱受体mRNA表达水平。结果CCK-8检测结果表明,1、2、4、8μg·cm^(-2)的二氧化硅刺激A549细胞24 h后,对细胞存活率均无影响(P均>0.05)。生化检测结果表明,与对照组相比,4、8μg·cm^(-2)二氧化硅暴露组SOD活性下降、IL-6水平升高,8μg·cm^(-2)二氧化硅暴露组TNF-α水平升高(P均<0.05);与对照组相比,8μg·cm^(-2)二氧化硅暴露组AChE酶活性、CHRM5及CHRNA7 mRNA表达水平均降低,CHRNA9 mRNA表达水平升高(P均<0.05)。结论二氧化硅暴露对A549细胞的毒性作用主要表现为氧化-抗氧化失衡、引发炎性反应和改变AChE酶活性及乙酰胆碱受体mRNA的表达水平。

Objective To investigate the toxicity of silica on human lung adenocarcinoma A549 cells derived from alveolar typeⅡepithelial cells.Methods A549 cells were exposed to silicon dioxide for 24 h to observe the effects of silicon dioxide on oxidative stress index,inflammatory cytokines and key molecules of non-neuronal cholinergic system in A549 cells.The levels of superoxide dismutase(SOD)and malondialdehyde(MDA)were determined by chemical colorimetry.The levels of tumor necrosis factor-α(TNF-α)and interleukin-6(IL-6)were determined by enzyma-linked immunosorbent assay(ELISA),the activity of AChE was determined by modified Ellman method,and the mRNA expression of acetylcholine receptor was determined by real-time PCR.Results CCK-8 assay showed that A549 cells were stimulated with silica of 1,2,4 and 8μg·cm^(-2) for 24 h,and the cell survival rate was not affected(P all>0.05).Biochemical test results showed that compared with the control group,SOD activity decreased and IL-6 level increased in 4 and 8μg·cm^(-2) silica exposure group,while TNF-αlevel increased in 8μg·cm^(-2) silica exposure group(P<0.05).Compared with the control group,AChE enzyme activity,CHRM5 and CHRNA7 mRNA expression levels were decreased in 8μg·cm^(-2) silica exposure group,while CHRNA9 mRNA expression level was increased(P all<0.05).Conclusion The toxic effects of silica exposure on A549 cells were mainly manifested as oxidation-antioxidant imbalance,triggering inflammatory response,changing AChE enzyme activity and acetylcholine receptor mRNA expression level.