缓激肽预适应人内皮祖细胞移植治疗裸鼠急性心肌梗死

目的:研究内皮祖细胞(endothelial progenitor cells,EPCs)经缓激肽(bradykinin,BK)预适应后移植对急性心肌梗死(acute myocardial infarction,AMI)的治疗作用?方法:细胞分为未经BK预适应人EPCs组(EPCs组)?BK预适应EPCs组(BK PC组)?BK预适应EPCs + 艾替班特(HOE140)组(BK PC/HOE组),双染色法四甲基偶氮唑蓝(MTT)比色法检测各组增殖情况,予缺氧诱导后Annexin V-FITC/PI双染色法及Hoechst33342染色法检测各组凋亡情况,同时收集上清液予ELISA法检测血管内皮生长因子(VEGF)的表达?体内实验建立裸鼠急性心肌梗死模型,将羰花青荧光染料DiD标记的人EPCs通过心肌局部注射进行细胞移植到实验组,安慰剂组注入等量生理盐水?分为假手术组?MI + 生理盐水组(安慰剂组)?MI + 单纯干细胞治疗组(EPCs组)?MI + BK预适应EPCs治疗组(BK PC组)及MI + BK预适应EPCs + HOE140治疗组(BK PC/HOE组)?细胞移植后10 d行超声心动图检测心功能?移植后10 d处死动物,取心脏标本行相关检查?结果:①与EPCs组相比,BK预适应可促进人EPCs增殖;缺氧诱导凋亡后,BK PC组凋亡率低于EPCs组(P < 0.01),且均可被HOE140阻断(P < 0.01)?同时BK PC组上清液中VEGF的分泌高于EPCs组(P < 0.01),同样可被HOE140所阻断;② 细胞移植10 d后BK PC组左心射血分数(LVEF)明显高于安慰剂组(P < 0.01)及EPCs组(P < 0.01),且可被HOE140阻断(P < 0.01)?与安慰剂组及EPCs组相比,BK PC组梗死周边区移植细胞存活明显增多,而细胞凋亡显著减少;同样可被HOE140所阻断?结论:BK预适应人EPCs后可显著改善裸小鼠AMI后的心功能,其疗效明显优于单纯人EPCs移植治疗?其治疗效应是通过促进移植细胞存活及抗移植细胞及宿主心肌细胞凋亡等而实现的?

Effect of human endothelial progenitor cells preconditioned with bradykinin on ischemic nude mice hearts

Objective:To investigate the effects of intra-myocardial transfer of bradykinin (BK)preconditioning(PC)endothelial progenitor cells (EPCs)in a mouse model of acute myocardial infarction. Methods:Cells were divided into three groups:human EPCs(hEPCs)without BK preconditioning group(EPCs group),BK PC hEPCs group(BK PC group),BK PC hEPCs + HOE140(icatibant)group (BK PC/HOE group). In vitro,proliferative ability was detected by MTT assay. Apoptosis induced by hypoxia was determined by annexin V-fluorescein isothiocyanate/propidium iodide staining and Hoechst 33342 staining. Cytokine concentrations in supernatant were measured. In vivo,hEPCs was labeled with carbocyanine near-infrared dye DiD prior to heart transplantation. Nude mice were allocated to the following treatment groups:sham group,MI + saline group (placebo group),MI + hEPCs group(EPCs group),MI + BK PC hEPCs group(BK PC group),MI + BK PC hEPCs + HOE140 group(BK PC/HOE group). Cardiac function was evaluated by echocardiography and postmortem analysis were assessed 10 days after transplantation. Results:In vitro,BK PC significanty promoted EPCs proliferation compared with the EPCs group(P < 0.01). Cell apoptosis rate post hypoxia was significantly reduced in the BK PC group compared with that in the EPCs group(P < 0.01),and was blocked by HOE140(P < 0.01). VEGF level was also significantly up-regulated in the BK PC group compared with that in the EPCs group(P < 0.01),and was also blocked by HOE140(P < 0.01). The BK PC group showed robust cell apoptosis inhibition,infarct size reduction,and cardiac function improvement in vivo and these effects were abrogated by the HOE140. Conclusion:Efficacy of BK PC hEPCs on improving cardiac function was greater than those by hEPCs in the mouse ischemia model. Its therapeutic effect could be achieved by promoting cell survival and resisting transplanted cells and host cardiomyocytes apoptosis.