Pitavastatin Pretreatment On Myocardial Ischemia-Reperfusion Oxidative Stress In Rats

Objective:Coronary heart disease is the leading cause of death across the world, which is characterized by myocardial ischemia (MI) as the consequences of coronary atherosclerosis. At present, the treatment used widely for acute or chronic MI is to restore the blood reperfusion to the ischemic regions, i.e. reperfusion therapy. However, reperfusion therapy can also induce myocardial ischemia reperfusion injury (MIRI). Nowadays, MIRI has become a hot issue among studies of cardiovascular diseases. Statins, 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitors, are not only a class of drugs used widely in clinical practice to lower lipid levels, but also show a strong cardioprotective effect, especially for prevention and treatment of MIRI. This study aimed to investigate whether pitavastatin, one of the third generation statins, could reduce the reperfusion injury in the myocardial ischemia-reperfusion(I/R) rat model. We also explored the mechanism of antioxidative stress of pitavastatin, and evaluated the efficacy of statins on the MIRI.Methods: Forty-two adult male and clean grade Sprague Dawley(SD)rats were randomized into three groups: the sham operation group (sham group, n=14) without coronary artery ligation was given saline 2ml/d by gastric irrigation for 4 weeks, and no ischemia-reperfusion happened; the ischemia-reperfusion group (IR group, n=14) with Left Anterior Descending Artery (LAD) ligation (LAD was ligated for 30 min, and then was loosed for 120 min) was given saline 2ml/d by gastric irrigation for 4 weeks; the test group(statin group n=14, pitavastatin, 0.42mg/kg/d, dissolved with saline into 2ml) was given pitavastatin by gastric irrigation for four weeks. After-operation, the bloods were collected from the rat hearts, and serum was extracted after centrifugation for the measurement of creatine kinase isoenzyme(CK-MB). We also obtained the myocardium tissue specimens from of the heart below the ligature to detect the activity of total superoxide dismutase (T-SOD) and the malondialdehyde (MDA) levels by the method of myocardial xanthine oxidase and thiobarbituric acid (TBA). The myocardial infarct size and risk area of each group were analyzed through comparing the differentiation of myocardium tissue stained by triphenyl tetrazolium chloride(TTC) and Evans blue by computer software system. Using this double stain method, normal myocardium tissue can be stained into blue, ischemic myocardium tissue can be stained into light red, and myocardial infarction region can be stained into gray. The percentage of myocardial infarct area can be calculated by equation: AN/AAR, where AN and AAR represents the area of necrosis and area at risk. The percentage of risk area was calculated by equation: AAR/LV, where LV represents the area of left ventricle.Results:1 All the rats in the study were adult male and clean grade SD rats, whose body weight were about 250±20g. Before ligation, all electrocardiograms (ECG) were normal. When LAD artery was ligated successfully, synchronized electrocardiogram showed widened QRS wave, elevated ST segment≥0.2mV in an arch shape, or an elevation of T wave voltage. In addition, pathologic q waves were observed or original q waves became deeper than before. When perfusion was established, synchronized electrocardiogram showed rapidly dropped T wave, ST segment decreased by 50%. The reperfusion-related arrhythmias, such as atrioventricular block, ventricular tachycardia, ventricular fibrillation etc, were also appeared.2 Before ligation, the heart of rats beat strongly, meanwhile, myocardium is bloom. Gross specimen morphology changes during operation: after ligation of the LAD, ischemic myocardial wall immediately turn into purple, or cyanosis, with a pale epicardium and Hypokinetic left ventricular anterior wall. When reperfusion of ischemic myocardium was restored, the ischemic region turned into red again. 3 The CK-MB levels of sham group, IR group and the statin group were (19.88±3.60)IU/L, (84.25±9.25)IU/L and (76.88±5.82)IU/L, respectively. Compared with the Sham group, the serum CK-MB was significantly elevated in IR group and the Statin group (each P less than 0.01). Compared with IR group, there was a dramatic reduction serum CK-MB in Statin group (P <0.01).4 For Sham group, IR group and Statin group, the levels of T-SOD were as follows: (134.43±8.18)U/mgprot, (94.07±6.45)U/mgprot and (113.52±5.73) U/mgport, respectively. Compared with the Sham group, the levels of myocardial T-SOD in IR group and Statin groups were significantly lower than that in Sham group (each P were less than 0.01). The T-SOD levels in Statin group was not significantly higher than that in IR group(P<0.01).5 The levels of MDA in Sham group, IR group and Statin group, were (2.12±0.44) nmol/mgprot, (4.52±0.35) nmol/mgprot, (3.86±0.60)nm -mol/mgport, respectively. The MDA levels in IR group and statin group were significantly higher than that in sham group (all P<0.01). There was a statistical significant increase in MDA in IR group compared with Statin group (P <0.05).6 Myocardial infarct area: Sham group no ischemia-reperfusion happened, and the colour were no difference. For IR group and Statin group, the percentages of AAR/LV was (37.31±5.05)% and (36.49±3.65)%, respectively. There was no significant diference between two groups (P>0.05), indicating the risk of myocardial ischemia in each group were similar. For AN/AAR, infarct size, the percentages in IR Group and Statin group were (65.44±7.61)% and (48.57±9.14)%, respectively. Statin group significantly reduced infarct size (P <0.01) compared with IR group.Conclusions:1 Our data showed Statin could significantly lower the levels of MDA, CK-MB and significantly increase the levels of T-SOD when compared with untreated rats, suggesting the effects of pitavastatin are attributed to increasing antioxidant capacity, alleviating myocardium necrosis, and inhibition of lipid peroxidation.2 Pitavastatin could reduce the levels of serum creatine kinase and the cardiac infarct area, thereby reduce myocardial ischemia-reperfusion injury. The reduction in infarct area might be associated with the effects of antioxidant.