接受骨髓间充质干细胞移植的心肌梗死后慢性心力衰竭大鼠心肌组织基质金属蛋白酶及抑制剂的变化

背景:Ⅰ型及Ⅲ型胶原含量与比例的变化是促使心脏几何形态改变、心肌收缩和舒展功能的重要因素,基质金属蛋白酶2及抑制剂是胶原代谢的主要调节物质。心肌梗死后心力衰竭大鼠接受骨髓干细胞移植后,心肌组织中这些指标会发生哪些变化?目的:观察接受骨髓干细胞移植治疗的心肌梗死后慢性心力衰竭大鼠,心肌组织中胶原含量与比例、基质金属蛋白酶2及抑制剂的变化。设计:随机对照实验。单位:解放军总医院老年心血管病科和北京医科大学生物化学系。材料:实验于2004-07/2005-12在北京医科大学生物化学系周春燕实验室完成。实验动物由北京医科大学动物实验中心提供,选取4周龄清洁级雄性SD大鼠用于骨髓间充质干细胞的制备,200~250g清洁级雌性SD大鼠14只用于心肌梗死后心力衰竭模型的制备,实验过程中对动物处置符合动物伦理学标准。方法:采用梯度离心法与贴壁培养法分离纯化、扩增骨髓间充质干细胞。结扎雌性SD大鼠左冠状动脉制作急性心肌梗死后心力衰竭模型。造模成功4周后将14只大鼠随机分为2组:移植组大鼠梗死后心肌瘢痕区接受雄性SD大鼠来源的5×106骨髓间充质干细胞。对照组大鼠梗死后心肌瘢痕区接受等体积磷酸盐缓冲液。每组7只。主要观察指标:移植治疗后21d:①以苏木精-伊红染色和Masson染色评价左心室形态。②以免疫组织化学分析心肌基质金属蛋白酶2及抑制剂与瘢痕区Ⅰ型及Ⅲ型胶原的变化。③以Western杂交检测基质金属蛋白酶2及抑制剂蛋白的变化。结果:14只大鼠均进入结果分析,无脱失值。与对照组大鼠比较,骨髓间充质干细胞移植大鼠心肌梗死瘢痕区Ⅰ型胶原增加,Ⅲ型胶原降低,心肌组织基质金属蛋白酶抑制剂表达增加,基质金属蛋白酶2表达降低,心室壁增厚,心室腔缩小,心功能增强。结论:骨髓间充质干细胞移植心力衰竭大鼠后,基质金属蛋白酶2及抑制剂发生了动态变化,进而影响瘢痕区胶原比例重建,逆转心力衰竭心脏异常的胶原平衡。

Expression levels of matrix metalloproteinases and tissue inhibitor of metalloproteinase in rats with myocardial infarction-caused chronic heart failure following bone marrow mesenchymal stem cell transplantation

BACKGROUND: The content of type Ⅰ and Ⅲ collagen and the ratio of both are crucial factors to promote heart geometric morphology change,and ventricular systolic and diastolic myocardial performance.Matrix metalloproteinase 2 (MMP-2) and tissue inhibitor of metalloproteinase 1 (TIMP1) are primary regulatory substances of collagen metabolism.After myocardial infarction,chronic heart failure rats were subjected to bone marrow meseuchymal stem cell transplantation.What changes in MMP-2 and TIMPI would occur?OBJECTIVE: To observe the content of type Ⅰ and Ⅲcollagen and the ratio between both,as well as expression levels of MMP-2 and TIMP1 in the left ventricular tissue of rats with myocardial infarction-caused chronic heart failure following bone marrow mesenchymal stem cell (MSC) transplantation.DESIGN: A randomized controlled experiment.SETTING: Department of Senile Angiocardiopathy,General Hospital of Chinese PLA & Department of Biochemistry,Peking University Health Science Center.MATERIALS: This study was performed at the laboratory of Department of Biochemistry,Peking University Health Science Center between July 2004 and December 2005.Male Sprage Dawley rats of clean grade,aged 4 weeks old,were provided by the Laboratory Animal Center,Beijing Medical University and used for preparation of MSCs.Fourteen female rats,weighing 200-250g,were developed into models of heart failure-caused by myocardial infarction.METHODS: MSCs were isolated and,purified by gradient centrifugation and adherent cells were allowed to proliferate.Female rats underwent coronary artery ligation to induce chronic ischemic heart failure.Four weeks later,the rats were randomly divided into 2 groups: (1) experimental group (n=7),rats received transplantation of MSCs harvested from male rats 5×106 in 50 μL phosphate buffered saline(PBS)]by injection into the ischemic myocardium; (2) control group (n=7),rats received the same volume of PBS.MAIN OUTCOME MEASURES: Twenty-one days after therapy,(1) left ventricular fusion was tested by hematoxylin-eosinstaining and Masson staining; (2) Expression of MMP-2 and 1TMPI as well as contents of type I and llI collagen was analyzed by immunohistochemistry; (3) MMP-2 and TIMP1 expression levels were examined by Western blot.RESULTS: Fourteen rats were included in the final analysis.Type Ⅰ collagen expression in the scar area was much higher in theexperimental group than in the control group,while type Ⅲ collagen expression was much lower in the experimental group.MMP-2 expression was reduced and TIMPI expression was increased in the experimental group compared with the control group.Together,ventricular wall was thickened,ventricular chamber was reduced,and heart function was strengthened in the experimental group compared with the control group.CONCLUSION: MSC transplantation alleviated left ventricular remodeling in chronic ischemie heart failure,which results from dynamic regulation of MMP-2/TIMP1.