Ghrelin对糖尿病大鼠心肌梗死后心肌重塑基质金属蛋白酶表达的影响

Effects of ghrelin on myocardial remodeling matrix metalloproteinases expression in post-myocardial infarction rats with diabetes mellitus

目的：探讨生长激素促分泌物受体（growth hormone secretagogue receptor,GHSR）的内源性配体ghrelin对糖尿病大鼠心肌梗死后心室重塑基质金属蛋白酶（matrix metalloptoteinases,MMPs）表达的影响。方法：选取7~8周龄的SD大鼠随机分为4组：1单纯心肌梗死组;2糖尿病合并心肌梗死组;3糖尿病合并心肌梗死＋ghrelin干预组;4糖尿病合并心肌梗死＋ghrelin＋GHSR1a特异性拮抗剂[D-Lys3]-GHRP-6干预组。建立链脲佐菌素糖尿病大鼠模型,12周后所有大鼠均结扎冠状动脉左前降支建立急性心肌梗死模型。Ghrelin干预组大鼠腹腔注射ghrelin[200μg/（kg·d）],ghrelin＋[D-Lys3]-GHRP-6干预组大鼠分别腹腔注射ghrelin[200μg/（kg·d）]＋[D-Lys3]-GHRP-6[50μg/（kg·d）],其余2组分别腹腔注射等量生理盐水。4周后苦味酸天狼猩红染色检测其心肌纤维化情况,Masson染色检测心肌梗死面积,免疫组织化学和Western blot分别检测左心室心肌组织基质金属蛋白酶2（matrix metalloptoteinase-2,MMP-2）、基质金属蛋白酶9（matrix metalloptoteinase-9,MMP-9）和金属蛋白酶组织抑制因子1（tissue inhibitor of metalloprotroteinase-1,TIMP-1）分布与表达变化。结果：与单纯心肌梗死组相比,糖尿病合并心肌梗死组心肌胶原容量分数（P=0.003）、心肌梗死面积明显增加（P=0.001）,MMP-2（P=0.000）和MMP-9（P=0.000）明显增高,心肌间质纤维增生。Ghrelin腹腔注射干预后可明显降低糖尿病心肌梗死大鼠胶原容量分数（P=0.001）、心肌梗死面积（P=0.005）,MMP-2（P=0.001）和MMP-9（P=0.001）表达减少,心肌间质纤维增生减轻。TIMP-1在合并糖尿病的组别中均明显升高（P=0.000）,但3组间无统计学意义（P〉0.05）。Ghrelin的上述效应均被[D-Lys3]-GHRP-6阻滞。结论：Ghrelin可通过抑制糖尿病心梗大鼠心肌组织MMP-2和MMP-9的表达,从而抑制糖尿病心梗大鼠心肌间质纤维化、减少心肌梗死面积,抑制心肌细胞外基质重塑。

Objective：To investigate the effects of ghrelin on myocardial remodeling matrix metalloproteinases expression in post-myocardial infarction diabetic rats. Methods：Adult male SD rats were divided into four groups ：myocardiac infarction（MI）,diabetes mellitus ＋myocardiac infarction（DM＋MI）,DM＋MI＋ghrelin,DM＋MI＋ghrelin＋GHSR1a inhibitor[D-Lys3]-GHRP-6. Diabetes was induced by injection of streptozotocin（STZ,60 mg/kg）. After three months,left anterior descending artery（LAD）ligation was performed in all groups. DM＋MI＋ghrelin group received ghrelin,200 μg/（kg·d）. DM＋MI＋ghrelin＋[D-Lys3]-GHRP-6 group received ghrelin,200 μg/（kg·d）and [D-Lys3]-GHRP-6,50 mg/（kg·d）. The other two groups received the same amount sterile normal saline only.Four weeks later,Sirius red staining was used to detect myocardial fibrosis. Myocardial infarct size was detected by Masson staining.The distribution and expressions of matrix metalloptoteinase-2（MMP-2）,matrix metalloptoteinase-9（MMP-9）and tissue inhibitor of metalloprotroteinase-1（TIMP-1）were detected by immunohistochemistry and Western blot,separately. Results ：Myocardial collagen volume fraction（P=0.003）,myocardial infarct size（P=0.001）,the expression of MMP-2（P =0.000） and MMP-9（P =0.000）were significantly increased in DM＋MI group than in MI group. After ghrelin administration,myocardial collagen volume fraction（P=0.001）,myocardial infarct size（P=0.005）,the expression of MMP-2（P=0.001）and MMP-9（P=0.001）were significantly decreased in diabetic rats complicated with MI. TIMP-1 was significantly higher in diabetic groups than in MI group（P=0.000）,without significant difference among these groups（P 0.05）. However,[D-Lys3]-GHRP-6 blocked the above effects of ghrelin. Conclusion：Ghrelin can inhibit myocardial extracellular matrix remodeling and reduce myocardial infarct size by suppressing the expressions of MMP-2 and MMP-9.