摘要
目的研究NOX4、NF-κB在氟化钠诱导HepG2细胞氧化应激中的作用。方法分别用浓度为0、10、20、40、80 mg/L的氟化钠处理细胞24 h,采用硫代巴比妥酸法测定细胞内丙二醛(malondialdehyde,MDA)含量,羟胺法测定细胞内总超氧化物歧化酶(superoxide dismutase,SOD)活力。免疫印迹法检测细胞内细胞核因子p65磷酸化水平和还原型烟酰胺腺嘌呤二核苷酸磷酸氧化酶4(nicotinamide adenine dinucleotide phosphate oxidase 4,NOX4)蛋白表达水平;用NOX4抑制剂夹竹桃麻素(apocynin)预处理30 min后,再用40 mg/L氟化钠处理24 h,免疫印迹法检测细胞内p65磷酸化水平和NOX4蛋白表达水平反映HepG2细胞内的氧化应激状况。结果经氟化钠处理后,20、40、80 mg/L剂量组HepG2细胞内的MDA含量较对照组相比明显升高(P<0.01),各剂量组细胞内总SOD活力均较对照组明显降低(P<0.01),40、80 mg/L剂量组NOX4蛋白表达水平和20、40、80 mg/L剂量组p65蛋白磷酸化水平均较对照组升高(P<0.01或0.05),存在着剂量效应;与单纯使用氟化钠处理组相比,用apocynin预处理细胞后,40 mg/L氟化钠诱导细胞内p65磷酸化水平和NOX4蛋白表达水平皆降低(P<0.05或0.01)。结论氟化钠诱导HepG2细胞产生氧化应激可能与激活NF-κB/NOX4途径有关。
Objective To investigate the role of NOX4 and NF-κB in oxidative stress of HepG2 cells induced by sodium fluoride Methods HepG2 cells were treated with sodium fluoride at concentrations of 0,10,20,40,and 80 mg/L for24 h,and the intramolecular malondialdehyde(MDA)content was determined by thiobarbituric acid method.The intracellular total superoxide dismutase(SOD)activity was determined by hydroxylamine method.The phosphorylation level of nuclear factor p65 and protein expression level of nicotinamide adenine dinucleotide phosphate oxidase 4(NOX4)in cells were detected by western blotting.Furthermore,pretreatment with NOX4 inhibitor apocynin for 30 min,then treated with 40 mg/L sodium fluoride for 24 h.The phosphorylation level of nuclear factor p65 and protein expression level of nicotinamide adenine dinucleotide phosphate oxidase 4(NOX4)in cells were detected by Western blotting.Results Compared with control group,MDA content in 20,40 and 80 mg/L sodium fluoride treated groups increased significantly(P<0.01).Compared with control group,the total SOD activity in HepG2 cells decreased significantly after sodium fluoride treatment(P<0.05).Compared with control group,the phosphorylation level of nuclear factor p65 of groups treated with 40 and 80 mg/L sodium fluoride and protein expression level of nicotinamide adenine dinucleotide phosphate oxidase 4(NOX4)in groups treated with 20,40 and 80 mg/L sodium fluoride increased significantly(P<0.01 or 0.05).Apocynin pretreatment significantly decreased the phosphorylation level of nuclear factor p65 and protein expression level of nicotinamide adenine dinucleotide phosphate oxidase 4(NOX4)(P<0.05 or 0.01).Conclusions Induction of oxidative stress in HepG2 cells by sodium fluoride may be associated with activation of the NF-κB/NOX4 pathway.
作者
杨成伟
章静
刘必勇
唐玉菲
王路晗
YANG Chengwei;ZHANG Jing;LIU Biyong;TANG Yufei;WANG Luhan(Anhui NO.2 Provincial People’s Hospital(Anhui Prevention&Treatment Center for Occupational Disease),Hefei,Anhui 230022,China;The Third People’s Hospital Of Hefei,Hefei,Anhui 230022,China)
出处
《职业卫生与应急救援》
2020年第6期595-598,共4页
Occupational Health and Emergency Rescue
基金
安徽省“十三五”医疗卫生重点培育实验室基金资助项目(皖卫科教2017-30)。
