安定导致小鼠大脑的线粒体氧应激损伤

Brain,mitochondrial oxidative injuries induced by diazepam in mouse

目的 探讨安定对大脑神经元产生毒副作用的分子机制。方法以１０ ｍｇ／ｋｇ·ｂ．Ｗ安定给小鼠进行腹腔注射，连续 ３ｄ后，取相同质量的大脑进行线粒体蛋白的分离，进行有关ＭｎＳＯＤ和细胞色素氧化酶ＩＶ活性以及线粒体ＤＮＡ断裂 程度等指标的测定。结果（１）安定导致小鼠大脑的线粒体蛋白含量降低，线粒体蛋白含量从（３．３０７±０．２４０）mg／ml下降到 （２．７４６±０．４５１）mg/ml（ｔ=２．６９０,０．０１＜Ｐ＜０．０５）；（２）安定引起神经元线粒体的ＭｎＳＯＤ活性从（７５.４６±６．６３）Ｕ／ｍｇ下降到 （５５．３０±３．８３）Ｕ／ｍｇ（ｔ＝６．４４８， Ｐ＜０．０１）；（３）安定对细胞色素氧化酶具有显著的损伤作用，５５０nm处细胞色素吸收值从 （０．１１２±０．００８）下降到 ０．０６９±０．００８（t＝８．７８２， Ｐ＜０.０１）；（４）安定能导致小鼠大脑的线粒体ＤＮＡ大量断裂。在碱裂解过程 中，０°Ｃ条件下，ＤＮＡ双链的百分比从（９２．９５±２．８２）％下降到（７９．５３±２．３３）％（ｔ=８．４６６，Ｐ＜０.０１）；在 １５℃作用１ｈ，ＤＮＡ双 链的百分比从（７７．４５±３．４９）％下降到（４３．１５±６．１）％（ｔ＝１３．

Objective To probe into the molecular mechanisms by which a single dose of diazepam causes side-effect injuries to the brain. Mehtods A single dose of diazepam (10mg/kg.b.w.) was injected daily into the abdominal cavity of the mice for 3 d. The mice receiving injection with saline served as control. The brain tissues weighing precisely 350 mg were obtained from each of the rats to isolate the mitochondria, and MnSOD and cytochrome C oxidase activities were determined. Assessment of the broken mitochondrial DNA was performed with FADU method. Results A single dose of diazepam caused the mitochondrial protein to decrease from (3.307±0.240) mg/ml to (2.7461±0.451) mg/ml (n=12, t=2.690, 0.01<P＜0.05) in the mouse brain, and MnSOD activity from (75.46±6.63) U/mg to (55.30±3.83) U/mg (n=12, t=6.448, p<0.0l). A reduction of the absorption coefficient of cytochrome C at 550 nm from 0.112±0.008 to 0.070±0.083 (n=12, t=8.782, p<0.0l) was also resulted Diazepam could disrupt the mitochondrial DNA strands, reducing the percentage of double strand DNA from 92.95%± 2.82% to 79.53%±ll.33% (t=8.466, P＜0.01) at 0°Ｃ and from 70.45%±3.49o% to 43.15%±6.01% (t=13.817, P<0.01) at 15°C. Conclusions A single dose of diazepam may lead to free radical-mediated injuries to the mouse brain, also causing mitochondrial DNA disruption by oxidative stress.