大鼠局灶性脑缺血/再灌注后氧化性损伤标记8-ohdG及其修复酶rOGG1表达的研究

8-ohdG AND rOGG1mRNA EXPRESSION IN RAT WITH FOCAL CEREBRAL ISCHEMIA AND REPERFUSION

目的:检测大鼠脑缺血/再灌注后8-ohdG及rOGG1的表达,以了解神经元损伤是否与修复酶活性下降有关。方法:健康雄性Wistar大鼠,体重250±30g,随机分为2组:①造模组;②假手术组。①组大鼠用线栓法成功制作可复流的MCAO模型,2小时后再灌注。在再灌注后6小时、24小时、48小时将大鼠断头取脑,保存于液氮。均匀切成2mm厚的脑片,取第三片提取DNA或RNA。DNA样品经酶解后上高效液相-电化学检测器检测8-ohdG。RNA样品通过RT-PCR的方法探测rOGG1mRNA的表达。结果:①造模组脑缺血再灌注各时点8-ohdG的含量均较假手术组明显减少(P<0.01)。随再灌注时间的增加,脑缺血区rooG lmRNA的含量逐渐增加。②随再灌注时间的增加,造模组脑缺血区rooGlmRNA的表达量逐渐增加,但仍较假手术组明显减少(P<0.01)。结论:脑缺血区受损DNA修复不完全将导致DNA损伤积累从而引起神经元死亡。

Objectives: To invesitgate the change and effect of 8-ohdG and rOOG1 in rat with focal cerebral ischemia and reperfusion. Methods: Male Wistar rats weighting 250±30g were dividing into 2 groups; ①pure model group,② sham-operated group. The focal cerebral ischemia and reperfusion model was made by thread embolish of middle cerebral artery. Rats were killed at 6, 24 and 48 hours after ischemia/reperfusion. The brain was removed, sliced into 2-mm sections. The third 2-mm section was applied to DNA or RNA extraction. DNA was analyzed by HPLC with the electrochemical (EC) detection. Using a semi-quantitative reverse transcription-polymerase chain reaction assay on total mRNA from the ischemic tissue, OGG1 mRNA was determined in rat brains. Results: ①-The amount of 8-ohdG in pure model group after reperfusion was significantly increasing compared with sham-operated group (P<0. 01). During reperfusion after 2 hours of ischemia with the passage of time, the amount of 8-ohdG in penumbra was reducing gradually. ②2During reperfusion after 2 hours of ischemia with the pasage of time, rOGGlmRNA expression in pure model group was increasing gradually, but there was a significant reduction compared with sham-operated group(P<0. 01). Conclusions: Oxidative DNA damage and repair defect might play a vital role in regulating the neuron death during reperfusion after ischemia.