摘要
目的 研究超氧阴离子自由基 (O2 .- )对 NIH3T3细胞 DNA的损伤及其脂质过氧化作用和对 c- m yc基因表达的影响。方法 用黄嘌呤 -黄嘌呤氧化酶 (X- XO)系统产生 O2 .- ;测定 DNA-溴乙锭结合物荧光强度变化确定 DNA损伤程度 ;以硫代巴比妥酸法测定脂质过氧化产物丙二醛 (MDA)含量 ;以细胞原位杂交法检测 c- myc基因表达。结果 高剂量 O2 .-能直接引起 DNA断裂损伤。O2 .-作用于完整细胞 ,MDA含量及 DNA断裂损伤显著增加 ,过氧化氢酶能明显抑制此效应。中、高剂量 O2 .- 能引起明显的 c- myc基因表达。结论 O2 .- 引起细胞 DNA损伤与过氧化氢的形成有关 ,其损伤的途径可能是通过膜脂质过氧化 ;O2 .- 诱导 NIH3T3细胞 c- m yc基因表达与剂量密切相关。
To study the DNA damage,membrane lipid peroxidation,and c-myc gene expression of NIH3T3 cell induced by superoxide anion(O 2 ·-). Methods The superoxide anion(O 2 ·-)was produced by reactive system of xanthine-xanthine oxidase (X-XO)and the DNA-EB complex,malondialdehyde(MDA)content and expression of c-myc gene were measured by means of fluorescence,thiobarbitoric acid (TBA) assay and DNA-RNA situ hybridization with digoxigenin labeled probes respectively. Results The O 2 ·-of high concentration(X:20 μg,XO:2×10 -3 U) reacted with isolated DNA from NIH3T3 cell directly which caused DNA damage obviously.When the O 2 ·-of high concentration reacted with whole NIH3T3 cell,the amount of MDA increased and fluorescence intensity of DNA-EB complex decreased which mean lipid peroxidation of membrane and gene damage of the cell respectively.When the producing O 2 ·-of high concentration reactive system existing FeSO 4,the phenomena mentioned above showed obviously.Enough amount of catalase showed the inhibit effects,but it could not completely inhibit the DNA damage of whole cell.The c-myc gene expression was observed in the cell treated with O 2 ·-of middle (X:4 μg,XO:4×10 -4 U)and high concentration,but it was not observed in the cell treated with O 2 ·-of low concentration(X:0.4 μg,XO:4×10 -5 U). Conclusions DNA damage of the whole cell induced by extracellular O 2 ·- was able to act through the pathway of membrane lipid peroxidation in which hydrogen peroxide is important and the other pathways such as signal transduction of the cell might also be existed.The expression of c-myc gene of NIH3T3 cell induced by O 2 ·- was related closely to the concentration of O 2 ·-.
出处
《中国医学科学院学报》
CAS
CSCD
北大核心
2000年第3期259-262,共4页
Acta Academiae Medicinae Sinicae
基金
国家自然科学基金! (396 70 196 )
关键词
超氧阴离子自由基
DNA损伤
脂质过氧化
基因表达
superoxide anion
reactive oxygen species
DNA damage
lipid peroxidation
gene expression
