CAJ | 학술논문

目的应用分子克隆技术建立CYP2E1基因高表达细胞株,观察三氯乙烯(TCE)对CYP2E1基因高表达细胞和正常肝细胞的毒性,探讨CYP2E1基因在TCE毒性中的作用.方法用不同剂量三氯乙烯(0、0.25、0.5、1.0、2.0和4.0 mmol/L)对CYP2E1高表达细胞和正常人肝细胞(L02细胞)进行染毒12h,观察凋亡基因(Bcl-2、caspase-3、caspase-8、caspase-9)和癌基因(c-fos、c-myc、k-ras、p53)表达变化.结果 TCE染毒引起L02肝细胞和CYP2E1高表达细胞Bcl-2表达水平下降,0.25～2.0 mmol/L TCE剂量时L02肝细胞Bcl-2表达下降50％～60％,但相同剂量组的CYP2E1高表达细胞Bcl-2表达水平明显高于正常肝细胞20％～50％; 0.5～4.0 mmol/L TCE剂量时CYP2E1高表达细胞中凋亡基因caspase-3、caspase-8、caspase-9表达水平比正常肝细胞高30％～600％,差异有统计学意义(P＜0.05或P＜0.01).TCE染毒后对L02肝细胞和CYP2E1高表达细胞的癌基因表达水平有明显差异,在0.5 ～4.0 mmol/L TCE染毒时,CYP2E1高表达细胞的c-fos、k-ras和c-myc基因表达水平明显高于L02肝细胞,差异有统计学意义(P＜0.05或P＜0.01),升高幅度达25％～120％; CYP2E1高表达细胞的p53表达水平比正常肝细胞表达下降10％～50％,差异有统计学意义(P＜0.05或P＜0.01).结论 TCE对正常肝细胞和CYP2E1高表达细胞凋亡基因和癌基因表达存在明显差异,提示CYP2E1是TCE在体内代谢的重要因素,与TCE毒性存在一定关系.
목적 응용분자극륭기술건립CYP2E1기인고표체세포주,관찰삼록을희(TCE)대CYP2E1기인고표체세포화정상간세포적독성,탐토CYP2E1기인재TCE독성중적작용.방법 용불동제량삼록을희(0、0.25、0.5、1.0、2.0화4.0 mmol/L)대CYP2E1고표체세포화정상인간세포(L02세포)진행염독12h,관찰조망기인(Bcl-2、caspase-3、caspase-8、caspase-9)화암기인(c-fos、c-myc、k-ras、p53)표체변화.결과 TCE염독인기L02간세포화CYP2E1고표체세포Bcl-2표체수평하강,0.25～2.0 mmol/L TCE제량시L02간세포Bcl-2표체하강50％～60％,단상동제량조적CYP2E1고표체세포Bcl-2표체수평명현고우정상간세포20％～50％; 0.5～4.0 mmol/L TCE제량시CYP2E1고표체세포중조망기인caspase-3、caspase-8、caspase-9표체수평비정상간세포고30％～600％,차이유통계학의의(P＜0.05혹P＜0.01).TCE염독후대L02간세포화CYP2E1고표체세포적암기인표체수평유명현차이,재0.5 ～4.0 mmol/L TCE염독시,CYP2E1고표체세포적c-fos、k-ras화c-myc기인표체수평명현고우L02간세포,차이유통계학의의(P＜0.05혹P＜0.01),승고폭도체25％～120％; CYP2E1고표체세포적p53표체수평비정상간세포표체하강10％～50％,차이유통계학의의(P＜0.05혹P＜0.01).결론 TCE대정상간세포화CYP2E1고표체세포조망기인화암기인표체존재명현차이,제시CYP2E1시TCE재체내대사적중요인소,여TCE독성존재일정관계.
Objective To investigate the effects of trichloroethylene (TCE) toxicity on the normal human liver cells (L02 cells) and hepatocytes with CYP2E1 gene overexpression which was constructed through molecular cloning technology in our laboratory,then to explore the roles of CYP2E1 gene in TCE toxicity.Methods L02 cells and hepatocytes with CYP2E1 overexpression were treated with various doses of TCE (0,0.25,0.5,1.0,2.0,4.0 mmol/L) for 12h,the expression of apoptosis genes (Bcl-2、Caspase-3、Caspase-8、Caspase-9) and oncogenes (c-fos、c-myc 、k-ras 、p53) were determined by real-time fluorescent PC R.Results Bcl-2 mRNA expression levels increased significantly in normal liver cells and CYP2E1-overexpressing cells after TCE treatment,Bcl-2 levels were 20％～50％ higher in CYP2E1-overexpressing cells than in L02 liver cells at doses of 0.25～2.0 mmol/L TCE.Caspase-3,Caspase-8 and caspase-9 mRNA expression increased by 30％～600％ in CYP2E1-overexpressing cells at doses of 0.5～4.0 mmol/L TCE when compared with L02 cells (P＜ 0.01).Additionally,c-fos、k-ras and c-myc mRNA expression levels were 25％～120％ higher in CYP2E1-overexpressing cells than in L02 cells (P＜0.01),p53 mRNA expression levels were lower 10％～50％ in CYP2E1-overexpressing cells than in L02 cells (P＜0.05 or P＜0.01).Conclusions There were significant differences for apoptosis gene and oncogene expression levels between normal liver cells and CYP2E1-overexpressing cells after they were treated with TCE,these findings indicated that CYP2E1 might play an important role in TCE metabolism in vivo.