丹参酮ⅡA对急性心肌缺血大鼠S100A1蛋白表达的影响

摘要

ObjectiveTo investigate the protective effect of tanshinoneⅡA on the expression of S100A1 protein after acute myocardial ischemia injury in rats.Methods Sixty Wistar rats were randomly divided into sham operation group, acute myocardial ischemia model group and tanshinoneⅡA pretreatment group by random number table. The acute myocardial ischemia model was established by thoracotomy and penetration of a thread and occlusion around the root part of the left anterior descending coronary artery, while the sham operation group was established only by thoracotomy and penetration of a thread around the root part of that artery but without occlusion; 3 days before the operation, in the tanshinoneⅡA pretreatment group, intraperitoneal injection of tanshinoneⅡA solution(at a dose of 1.5 mg/kg) was applied, while in the sham and acute myocardial ischemia groups, intraperitoneal injection of an equal volume of saline was given. Myocardial cell apoptosis was detected by the terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling(TUNEL), the levels of serum superoxide dismutase (SOD), malondialdehyde(MDA), creatine kinase(CK), lactate dehydrogenase(LDH) and S100A1 protein were examined and the level of expression of S100A1 protein in myocardial tissue was assayed by immunohistochemical staining and Western Blot.Results Compared with the sham operation group, the myocardial cell apoptosis rate, the contents of MDA, CK, LDH, S100A1 and the level of S100A1 expression in myocardial ischemia group and tanshinoneⅡA pretreated group were significantly increased, while SOD activity was decreased obviously; compared with the myocardial ischemia model group, the myocardial cell apoptosis rate, the contents of MDA, CK, LDH, S100A1 and the level of S100A1 protein expression were significantly reduced〔apoptosis rate:(32.1±4.2)% vs.(72.4±5.4)%, MDA(μmol/L): 9.1±2.2 vs. 17.3±5.2, CK(U/L): 83.3±12.2 vs. 107.5±12.4, LDH (μmol·s-1·L-1): 84.0±16.4 vs. 114.4±16.0, S100A1(μg/L): 37.6±6.0 vs. 78.4±8.6,P<0.05 orP<0.01〕, while the activity of SOD was increased markedly in tanshinoneⅡA pretreated group(kU/L:72.8±10.2 vs. 49.6±8.8,P<0.01). TUNEL staining showed that in the myocardial ischemia model group and tanshinoneⅡA pretreated group, the myocardial cells represented positive staining(brown-yellow in color), irregular in shape with nuclear pyknosis, cell detachment from the surrounding tissue and other characteristics. And in sham operation group,the staining of majority of cells was negative. The results of immunohistochemistry showed that S100A1 protein staining was relatively deep in the myocardial ischemia model group and tanshinoneⅡA pretreated group, and in the latter group, the color of S100A1 protein positive staining was not as deep as that in the former group. Western Blot showed that the S100A1 protein expression in myocardial ischemia model group was 2.8 folds of that of the sham operation group, while the S100A1 protein expression in tanshinoneⅡA pretreated group was significantly decreased compared with that of myocardial ischemia model group(bothP<0.05),which was 1.5 folds of that of the sham operation group.ConclusionTanshinoneⅡA may play a role in inhibiting the expression of S100A1 protein to protect against acute myocardial ischemia injury, suggesting that this agent have a potential effect for treatment of myocardial ischemia.%目的：探讨丹参酮ⅡA对大鼠急性心肌缺血损伤后S100A1蛋白表达的影响。方法选择Wistar大鼠60只，按随机数字表法分为假手术组、急性心肌缺血模型组、丹参酮ⅡA预处理组。采用左冠状动脉（冠脉）前降支根部穿线结扎的方法建立大鼠急性心肌缺血模型；假手术组仅开胸于大鼠左冠脉前降支根部穿线但不结扎；丹参酮ⅡA预处理组于术前3 d腹腔注射丹参酮ⅡA注射液（剂量为1.5 mg/kg），假手术组和急性心肌缺血模型组以等体积生理盐水腹腔注射。采用原位末端缺刻标记法（TUNEL）检测心肌细胞凋亡情况；检测血清超氧化物歧化酶（SOD）、丙二醛（MDA）、肌酸激酶（CK）、乳酸脱氢酶（LDH）及S100A1蛋白水平；采用免疫组化染色和蛋白质免疫印迹试验（Western Blot）检测心肌组织S100A1蛋白的表达水平。结果与假手术组比较，心肌缺血模型组和丹参酮ⅡA预处理组心肌细胞凋亡率及MDA、CK、LDH、S100A1表达水平均明显升高，SOD活性明显降低；与心肌缺血模型组比较，丹参酮ⅡA预处理组心肌细胞凋亡率及MDA、CK、LDH、S100A1蛋白表达水平均明显降低〔细胞凋亡率：（32.1±4.2）%比（72.4±5.4）%，MDA（μmol/L）：9.1±2.2比17.3±5.2，CK（U/L）：83.3±12.2比107.5±12.4，LDH（μmol · s-1· L-1）：84.0±16.4比114.4±16.0，S100A1蛋白（μg/L）：37.6±6.0比78.4±8.6，P＜0.05或P＜0.01〕，SOD活性明显升高（kU/L：72.8±10.2比49.6±8.8，P＜0.01）。TUNEL染色结果显示，心肌缺血模型组和丹参酮ⅡA预处理组心肌细胞部分染色呈阳性（棕黄色），呈现形态不规则、与周围组织脱离和核固缩等特征，假手术组大部分细胞染色呈阴性。免疫组化结果显示，心肌缺血模型组和丹参酮ⅡA预处理组S100A1蛋白染色较深，丹参酮ⅡA预处理组阳性蛋白染色较浅。Western Blot结果显示，心肌缺血模型组S100A1蛋白表达是假手术组的2.8倍，丹参酮ⅡA预处理组较心肌缺血模型组明显降低（均P＜0.05），是假手术组的1.5倍。结论丹参酮ⅡA可能是通过抑制S100A1蛋白的表达对急性心肌缺血损伤起到保护作用，提示丹参酮ⅡA具有治疗心肌缺血的潜能。