中国药理学与毒理学杂志
中國藥理學與毒理學雜誌
중국약이학여독이학잡지
CHINESE JOURNAL OF PHARMACOLOGY AND TOXICOLOGY
2012年
2期
138-144
,共7页
林勃%姜丽平%耿成燕%仲来福
林勃%薑麗平%耿成燕%仲來福
림발%강려평%경성연%중래복
8-羟基喹啉酮%氧化应激%谷胱甘肽%细胞,HepG2
8-羥基喹啉酮%氧化應激%穀胱甘肽%細胞,HepG2
8-간기규람동%양화응격%곡광감태%세포,HepG2
copper 8-quinolinolate%oxidative stress%glutathione%cells,HepG2
目的 评估8-羟基喹啉酮( CuQ)对HepG2细胞的DNA损伤作用并阐明其可能的作用机制.方法 CuQ 0 ～4 μmol·L-1处理HepG2细胞不同时间后,通过单细胞凝胶电泳实验检测细胞DNA损伤；分光光度法测定过氧化氢酶活性；苯二醛法测定细胞内谷胱甘肽(GSH)水平；硫代巴比妥酸反应物(TBARS)法检测细胞内脂质过氧化水平；Western印迹法检测NF-κB p65的变化；免疫组化方法检测细胞内8-羟基脱氧鸟苷(8-OHdG)的表达水平.结果 HepG2细胞与CuQ 0.5～4μmol· L-1作用1h后,DNA的迁移距离明显增加(P＜0.05),提示CuQ可引起DNA链断裂.CuQ能够造成细胞内GSH水平以及过氧化氢酶活性的降低.随着CuQ剂量的增加及染毒时间的延长,NF-κB由细胞浆逐渐转移至细胞核.CuQ还可以引起细胞内TBARS水平增高及8-OHdG表达水平的增强.采用GSH合成特异抑制剂DL-甲硫氨酸磺酰亚胺(BSO)预处理细胞,可明显增强CuQ对HepG2细胞DNA的损伤(P＜0.05).结论 CuQ可造成HepG2细胞氧化性DNA损伤,其作用机制与氧化应激及NF-κB p65在细胞核蓄积增高有关.
目的 評估8-羥基喹啉酮( CuQ)對HepG2細胞的DNA損傷作用併闡明其可能的作用機製.方法 CuQ 0 ～4 μmol·L-1處理HepG2細胞不同時間後,通過單細胞凝膠電泳實驗檢測細胞DNA損傷；分光光度法測定過氧化氫酶活性；苯二醛法測定細胞內穀胱甘肽(GSH)水平；硫代巴比妥痠反應物(TBARS)法檢測細胞內脂質過氧化水平；Western印跡法檢測NF-κB p65的變化；免疫組化方法檢測細胞內8-羥基脫氧鳥苷(8-OHdG)的錶達水平.結果 HepG2細胞與CuQ 0.5～4μmol· L-1作用1h後,DNA的遷移距離明顯增加(P＜0.05),提示CuQ可引起DNA鏈斷裂.CuQ能夠造成細胞內GSH水平以及過氧化氫酶活性的降低.隨著CuQ劑量的增加及染毒時間的延長,NF-κB由細胞漿逐漸轉移至細胞覈.CuQ還可以引起細胞內TBARS水平增高及8-OHdG錶達水平的增彊.採用GSH閤成特異抑製劑DL-甲硫氨痠磺酰亞胺(BSO)預處理細胞,可明顯增彊CuQ對HepG2細胞DNA的損傷(P＜0.05).結論 CuQ可造成HepG2細胞氧化性DNA損傷,其作用機製與氧化應激及NF-κB p65在細胞覈蓄積增高有關.
목적 평고8-간기규람동( CuQ)대HepG2세포적DNA손상작용병천명기가능적작용궤제.방법 CuQ 0 ～4 μmol·L-1처리HepG2세포불동시간후,통과단세포응효전영실험검측세포DNA손상；분광광도법측정과양화경매활성；분이철법측정세포내곡광감태(GSH)수평；류대파비타산반응물(TBARS)법검측세포내지질과양화수평；Western인적법검측NF-κB p65적변화；면역조화방법검측세포내8-간기탈양조감(8-OHdG)적표체수평.결과 HepG2세포여CuQ 0.5～4μmol· L-1작용1h후,DNA적천이거리명현증가(P＜0.05),제시CuQ가인기DNA련단렬.CuQ능구조성세포내GSH수평이급과양화경매활성적강저.수착CuQ제량적증가급염독시간적연장,NF-κB유세포장축점전이지세포핵.CuQ환가이인기세포내TBARS수평증고급8-OHdG표체수평적증강.채용GSH합성특이억제제DL-갑류안산광선아알(BSO)예처리세포,가명현증강CuQ대HepG2세포DNA적손상(P＜0.05).결론 CuQ가조성HepG2세포양화성DNA손상,기작용궤제여양화응격급NF-κB p65재세포핵축적증고유관.
OBJECTIVE To assess the DNA damage of copper 8-quinolinolate (CuQ) and to elucidate the plausible mechanisms.METHODS HepG2 cells were treated with CuQ0-4 μmol·L-1 for different time,DNA damage was measured by Comet assay.Catalase (CAT) activity,glutathione(GSH) level and thiobarbituric acid reactive substances (TBARS) were measured.NF-κB was examined using Western blotting.8-Hydroxydeoxyguanosine (8-OHdG) was measured by immunoperoxidase staining analysis.RESULTS CuQ 0.5 -4 μmol·L-1 caused significant increase of DNA migration in HepG2 cells.CuQ significantly decreased levels of GSH and activity of CAT in HepG2 cells (P ＜0.05).Moreover,CuQ significantly increased accumulation of the p65 subunit of NF-κB into nucleus,levels of lipid peroxidation product TBARS and the formation of 8-OHdG (P ＜0.05).The intracellular GSH level was modulated by pre-treatment with buthionine-(S,R)-sulfoximine (BSO),depletion of GSH in HepG2 cells pre-treated with BSO dramatically increased susceptibility of HepG2 cells to CuQ-induced DNA damage.CONCLUSION CuQ exerts DNA damage by oxidative stress and increases accumulation of p65 subunit of NF-κB into nucleus in HepG2 cells.
