缺氧对骨髓间充质干细胞肾素血管紧张素系统表达的影响  

Upregulating the renin-angiotensin system in bone marrow mesenchymal stem cells by hypoxia

作　　者：肖容容 高景红[1] 范越[1] 周露[1] 施睿臻[1] 李庆平[1] 

机构地区：[1]南京医科大学基础医学院药理系,南京医学硕士研究生210029

出　　处：《医学研究生学报》2015年第2期123-126,共4页Journal of Medical Postgraduates

基　　金：国家自然科学基金(30973534;81173052);南京医科大学科技发展基金(2013NJMU004)

摘　　要：目的肾素-血管紧张素系统(renin-angiotensin system,RAS)参与缺氧心肌损伤过程。文中旨在观察骨髓间充质干细胞(mesenchymal stem cells,MSCs)自身主要反应元件血管紧张素受体1(angiotensin II type 1 receptor,AT1-R)、AT2-R和血管紧张素转换酶(angiotensin-converting enzyme,ACE)在缺氧条件下的表达情况。方法分离培养大鼠的MSCs,用倒置相差显微镜观察细胞形态,应用CD29、CD11b/c抗体进行细胞鉴定。实验分为对照组和Hypoxia/SD组:对照组用含10%FBS的L-DMEM培养MSCs,Hypoxia/SD组采用缺氧联合无血清处理24 h诱导缺氧损伤模型。2组分别用锥虫蓝染色、CCK-8检测及流式细胞术,测定细胞活力和凋亡率。应用Real-time Quantitative PCR法检测细胞内AT1-R、AT2-R和ACE的mRNA表达水平;用Western blot检测细胞内AT1-R、AT2-R和ACE蛋白表达的变化。结果 MSCs特异性抗原CD29阳性率>97%,非特异性抗原CD11b/c阳性率<1%,分离培养成功;与对照组相比,Hypoxia/SD组24 h可显著增加MSCs凋亡率[(6.73±0.78)%vs(19.93±4.92)%,P<0.01],降低细胞活力[(78.49±4.94)%vs(37.33±2.91)%,P<0.01]。Hypoxia/SD环境下MSCs自身的RAS主要反应元件AT1-R、AT2-R及ACE的mRNA和蛋白表达量均增加。结论推断缺氧导致MSCs自身RAS激活,从而提高MSCs对缺氧损伤区的心肌修复能力。Objective The renin-angiotensin system（ RAS） is involved in myocardial anoxic injury. This study aimed to investigate the expressions of AT1-R,AT2-R,and angiotensin-converting enzyme（ ACE） in bone marrow mesenchymal stem cells（ MSCs） under hypoxia. Methods Rat MSCs were isolated,cultured,and identified with CD29 and CD11b/c antibodies. The ischemic injury model was established by exposing the MSCs to hypoxia and serum deprivation（ Hypoxia/SD） for 24 hours,while the control cells were cultured in L-DMEM with 10% FBS. The vitality and apoptosis of the cells were detected by trypan blue staining,CCK8 assay,and Annexin V-FITC staining. The mRNA and protein expressions of AT1-R,AT2-R,and ACE were determined by realtime quantitative PCR and Western blot,respectively. Results The positive rate of CD29 was 〉97% and that of CD11 b/c was 1% in the MSCs. Compared with the control group,Hypoxia/SD significantly increased the rate of cell apoptosis（ [6. 73 ± 0. 78]% vs[19. 93 ± 4. 92]%,P〈0. 01）,decreased the rate of cell viability（ [78. 49 ± 4. 94]% vs [37. 33 ± 2. 91]%,P〈0. 01）,and upregulated the mRNA and protein expressions of AT1-R,AT2-R,and ACE. Conclusion Hypoxia/SD activates the RAS in MSCs and improves the protective function of the cells against myocardial anoxic injury.

关 键 词：肾素血管经张素系统 骨髓间充质干细胞 缺氧/无血清 

分 类 号：R363[医药卫生—病理学]