糖尿病大鼠心肌缺血再灌注时DJ-1蛋白表达的变化

Changes in expression of DJ-1 protein during myocardial ischemia-reperfusion in diabetic rats

目的评价糖尿病大鼠心肌缺血再灌注时DJ—1蛋白表达的变化。方法成年雄性sD大鼠50只，体重220～280g，以腹腔注射1％链脲佐菌素-柠檬酸盐缓冲液60mg／kg的方法制备1型糖尿病模型。取糖尿病模型制备成功的大鼠40只，采用随机数字表法，将其分为3组：糖尿病组（D组，n=10）、糖尿病假手术组（DS组，n=15）和糖尿病心肌缺血再灌注损伤组（DIR组，n=15）。另取正常大鼠10只，单次腹腔注射檬酸盐缓冲液6ml／kg，作为对照组（c组）。DIR组采用结扎左冠状动脉前降支30min，再灌注120min的方法建立大鼠心肌缺血再灌注损伤模型。于再灌注120min时，Ds组和DIR组随机处死5只大鼠，取心肌组织，测定心肌梗死面积，计算心肌梗死体积百分比；各组处死10只大鼠，取心肌组织，光镜下观察病理学变化，采用比色法测定MDA含量，采用羟胺法测定SOD活力，采用TUNEL法检测细胞凋亡，计算细胞凋亡指数（AI），采用免疫组化SP法测定DJ-1和磷酸酶-张力蛋白酶基因（PTEN）的蛋白表达。DJ-1蛋白表达分别与MDA含量、SOD活性、AI和PTEN蛋白表达进行直线相关分析。结果与Ds组比较，DIR组心肌梗死体积百分比升高（P〈0．05）。与c组比较，D组、DS组和DIR组MDA含量和AI升高，SOD活性降低，DJ．1蛋白表达下调，PTEN蛋白表达上调（P〈0．05）；与D组和Ds组比较，DIR组MDA含量和AI升高，SOD活性降低，DJ-1蛋白表达下调，PTEN蛋白表达上调（P〈0．05）；D组和DS组上述各指标比较差异无统计学意义（P〉0．05）。MDA含量、SOD活性、AI、PTEN蛋白表达均与DJ-1蛋白表达具有相关性，r依次为-0．734、0．593、-0．818、-0．812（P〈0．05）。结论DJ-1蛋白表达下调可能通过减弱抗氧化应激反应，上调PTEN蛋白表达参与了糖尿病大鼠心肌缺血再灌注损伤。

Objective To evaluate the changes in the expression of DJ-1 protein during myocardial iseh- emia-reperfusion （I/R） in diabetic rats. Methods Fifty male Sprague-Dawley rats, weighing 220-280 g, were used in this study. Type 1 diabetes mellitus was induced by intraperitoneal streptozotoein 65 mg/kg and confirmed by fasting blood glucose 〉 16.7 mmol/L. Forty animals with type 1 diabetes mellitus were randomly divided into 3 groups： diabetes group （group D, n = 10）, diabetic sham operation group （group DS, n = 15） and diabetic I/R group （group DIR, n = 15）. Another 10 non-diabetic rats in which citrate buffer 6 ml/kg was injected intraperito- neally were served as control group （group C） . Myocardial I/R was produced by occlusion of the anterior descend- ing branch of left coronary artery for 30 min followed by 120 min reperfusion in group I/R. At 120 min of reperfu- sion, 5 rats were sacrificed and myocardial specimens were obtained for determination of infarct size in groups DS and DIR, and 10 rats were sacrificed and myocardial specimens were obtained for microscopic examination and for determination of cell apoptosis, malondialdehyde （MDA） content, superoxide dismutase （SOD） activity and expression of D J- 1 and phosphatase and tensin homologue （PTEN） protein. Apoptotic index （AI） was calculated. Linear correlation between the expression of D J-1 protein and MDA content, SOD activity, AI and expression of PTEN protein was analyzed. Results Compared with group DS, the myocardial infraet size was signifieandy in- creased in group DIR （P 〈 0.05 ）. Compared with group C, MDA content and AI were significantly increased, SOD activity was decreased, the expression of DJ-1 was down-regulated, and the expression of PTEN protein was up-regulated in groups D, DS and DIR （P 〈 0.05） . Compared with groups D and DS, MDA content and AI were significantly increased, SOD activity was decreased, the expression of DJ-1 was down-regulated, and the expres- sion of PTEN protein was up-regulated in group DIR （P 〈 0.05） . There was no significant difference in the param- eters mentioned above between groups D and DS （ P 〉 0.05） . There was linear correlation between the expression of DJ-1 protein and MDA content, SOD activity, AI and expression of PTEN protein and the correlation coefficients （r） were -0.734, 0.593, -0.818, and -0.812 in turn. Conclusion Down-regulation of DJ-1 protein expression is involved in myocardial I/R injury in diabetic rats via decreasing anti-oxidative stress responses and upregulating PTEN protein expression.