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目的:探讨血管紧张素Ⅱ(angiotensinⅡ,AngⅡ)预处理对骨髓间充质干细胞(mesenchymal stem cells,MSCs)耐缺氧能力的影响。方法:分离培养大鼠MSCs,应用CD29、CD11b/c抗体进行细胞鉴定。对细胞以不同浓度AngⅡ预处理3 h,采用缺氧联合无血清(hypoxia/SD)的方法诱导缺氧损伤模型;应用血管紧张素1型受体(AT1)拮抗剂氯沙坦(losartan)、血管紧张素2型受体(AT2)拮抗剂PD123319进行干预。应用MTT和CCK-8法测定细胞活力以确定最佳刺激浓度,采用台盼蓝染色,CCK-8检测和流式细胞术测定各组细胞活力和凋亡率。结果:①MSCs表面抗原CD29阳性率>97%,CD11b/c阳性率<1%;②AngⅡ预处理显著提高MSCs的活力,减少细胞凋亡率;③AngⅡ对MSCs的预适应保护作用呈现一定的浓度依赖性,最佳浓度为10nmol/L;④氯沙坦可抑制AngⅡ的预适应细胞保护作用,而PD123319无此效应。结论:AngⅡ通过AT1受体发挥对MSCs的预适应保护作用。
Objective: To investigate the effect of angiotensin Ⅱ (AngⅡ) pretreatment on hypoxia tolerance of mesenchymal stem cells (MSCs). Methods: Rat MSCs were isolated and cultured, and the cells were identified by CD29 and CD11b / c antibodies. The cells were pretreated with different concentrations of Ang II for 3 h, hypoxic injury was induced by hypoxia combined with hypoxia / SD, and losartan, an angiotensin type 1 receptor (AT1) antagonist, Intervention with angiotensin II receptor (AT2) antagonist PD123319. Cell viability was determined by MTT assay and CCK-8 assay to determine the best stimulation concentration. Cell viability and apoptosis rate were determined by trypan blue staining, CCK-8 assay and flow cytometry. Results: ① The positive rate of CD29 positive surface antigen of MSCs was> 97%, the positive rate of CD11b / c was less than 1%. ② AngⅡ pretreatment significantly increased the viability of MSCs and decreased the apoptosis rate. ③ AngⅡ had a preconditioning protective effect on MSCs in a concentration-dependent manner , The optimal concentration was 10nmol / L; ④ losartan inhibited Ang Ⅱ preconditioning cell protection, while PD123319 had no such effect. Conclusion: AngⅡ exerts its preconditioning protective effect on MSCs through AT1 receptor.