AT2R基因在体可调控表达对大鼠血管损伤后MMP-2表达的影响  

Effect of regulatable expression of AT2R on MMP-2 expression in neointima after vascular injury in rats

作　　者：景涛[1] 刘建平[1] 苗莉[1] 冯健[2] 冉擘力[1] 王海东[3] 何国祥[1] 

机构地区：[1]第三军医大学西南医院心内科,重庆400038 [2]第三军医大学西南医院胸心外科,重庆400038 [3]泸州医学院附属医院心内科,泸州646000

出　　处：《中华老年多器官疾病杂志》2013年第3期213-217,共5页Chinese Journal of Multiple Organ Diseases in the Elderly

基　　金：国家自然科学基金（No.30400180）

摘　　要：目的研究间充质干细胞（MSCs）移植实现血管紧张素Ⅱ2型受体（AT2R）基因在体可调控表达对大鼠颈动脉损伤后新生内膜中基质金属蛋白酶2（MMP-2）表达的影响。方法采用常规分子生物学方法连续两个回合转染体外培养的MSCs，获得受到强力霉素（Dox）调控的低背景、高诱导表达AT2R基因的双重稳定MSCs系；建立大鼠颈动脉球囊损伤动物模型，将双重稳定MSCs系种植于动脉损伤局部，通过尾静脉注射Dox，分别于术后14、28d行病理切片，采用免疫印迹、免疫组化法及RT-PCR等技术检测AT2R基因在新生内膜中的表达及其对MMP．2表达的影响。结果成功建立了低背景、高诱导表达AT2R基因的双重稳定MSCs系。该MSCs系受到Dox给予／去除的紧密调控，诱导后48h即可使AT2R显著表达，在Dox干预72h后AT2R表达进一步增强；这种表达的可诱导性至少在8周内维持稳定。免疫印迹、免疫组化及RT-PCR检测显示：球囊损伤大鼠血管后14d及28d，Dox组新生内膜中AT2R的表达显著高于对照组、MSC组、MSC转染组（P〈0．01）；同时，Dox组新生内膜中MMP．2阳性染色较对照组、MSC组和MSC转染组显著减弱（P〈0．01）。结论血管损伤局部导人双重稳定MSCs系后AT2R表达受到Dox的良好调控。Dox诱导AT2R表达后，新生内膜中MMP-2表达减少，可能是MSCs细胞移植实现AT2R基因在体可调控表达有效抑制新生内膜增生的机制之一。Objective To evaluate the role of regulatable expression of AT2R gene in vivo on the expression of matrix metalloproteinase 2 （MMP-2） in the neointima after vascular injury in rats by deoxycycline （Dox）-on mesenchymal stem cells （MSCs） transplantation. Methods Two successive rounds of transfection of cultured MSCs were performed with conventional molecular biological methods. MSCs with low background expression and high Dox-induced expression of AT2R gene were deemed double-stable MSCs. Rat models of balloon-induced carotid injury were established. Double-stable MSCs were then transplanted into the site of carotid injury, and Dox was injected via vena caudalis into rats. Pathological study was carried out at 14d and 28d after cell transplantation. AT2R gene expression in the neointima and its effect on the mRNA and protein expression of MMP-2 were analyzed by immunohistochemistry, immunoblotting and RT-PCR. Results Double stable MSCs line with low background expression and high Dox induced expression of AT2R gene was established successfully. Significant AT2R expression was observed after 48h of induction and further increased after 72h and remained stable over 8 weeks. Immunohistochemistry, immunoblotting and RT-PCR indicated that AT2R expression in the neointima was significantly upregulated in Dox group than in control group, MSC group and MSC transfection group （P 〈 0.01）, but the expression of MMP-2 was significantly decreased in Dox group than in other groups （P 〈 0.01）. Conclusion AT2R expression at the site of carotid injury after transfection of our double-stable MSCs were well controlled by Dox. After Dox induces the high expression of AT2R, the expression of MMP-2 in neointima is decreased, which may be one of the mechanisms of regulating AT2R expression in injured vessels to prevent vascular restenosis.

关 键 词：血管紧张素Ⅱ2型受体 可调控表达 间充质干细胞 新生内膜增生 基质金属蛋白酶 

分 类 号：R543.3[医药卫生—心血管疾病]