脐血间充质干细胞移植对盐敏感性高血压大鼠心肌纤维化及细胞凋亡的影响

Effects of umbilical cord blood mesenchymal stem cells transplantation on myocardial fibrosis and cell apoptosis in salt-sensitive hypertensive rats

目的探讨脐血间充质干细胞(UCMSCs)移植对盐敏感性高血压大鼠心肌纤维化和心肌细胞凋亡的影响。方法 60只Dahl大鼠,采用皮下注射辣椒辣素构建盐敏感性高血压实验动物模型,按随机数字表法随机分为正盐组、高盐组及UCMSCs移植组,每组20只。正盐组给予0.3%氯化钠饮食,高盐组给予8%氯化钠饮食,UCMSCs移植组予8%氯化钠饮食。8周之后,UCMSCs移植组大鼠尾静脉注射UCMSCs 1 ml(3×106个/ml PKH-26标记的UCMSCs),连续治疗3 d。实验2周末对大鼠进行超声心动图检测左室射血分数(LVEF)及左室短轴缩短率(FS),计算左心指数。HE染色观察心肌细胞形态,进行Van Gieson(VG)染色应用计算机图像分析系统测算胶原容积积分(CVF);荧光显微镜观察PKH-26标记的UCMSCs在心肌组织中分布情况,TUNEL法检测各组大鼠心肌细胞凋亡情况。结果 (1)正盐组大鼠的心肌细胞排列有序,心肌间质无炎症细胞浸润及纤维化瘢痕形成;高盐组心肌细胞排列紊乱,巨噬细胞浸润明显增多同时伴有明显的纤维化重构;与高盐组相比较,UCMSCs移植组尽管有炎性细胞浸润但心肌细胞排列有序,且无纤维化瘢痕组织形成,心肌的纤维化重构较轻。(2)与正盐组相比,高盐组大鼠的LVEF和FS显著降低(P均<0.05),其左心指数明显上升(P<0.05)。与高盐组相比,UCMSCs移植干预之后大鼠的LVEF和FS明显的提高(P均<0.05),而左心指数降低(P<0.05)。(3)心脏CVF正盐组最低,UCMSCs移植组次之,而高盐组最高,各组组间差异有统计学意义(P<0.05)。(4)仅在UCMSCs移植组的实验大鼠心肌组织中可见较多的PKH-26标记的UCMSCs。(5)正盐组、高盐组、UCMSCs移植组心肌细胞凋亡率分别为0、(28.2±6.3)%、(12.7±2.6)%,与高盐组比较,UCMSCs移植组大鼠心肌细胞凋亡率明显降低(P<0.05)。结论UCMSCs移植能减轻盐敏感性高血压大鼠心脏纤维化,减少胶原沉积,减少心肌细胞凋亡,改善大鼠心脏功能。

Objective To investigate the effects of transplantation of umbilical cord blood mesenchymal stem cells （ UCM- SCs） on myocardial fibrosis and cell apoptosis in rats with salt-sensitive hypertension. Methods The salt-sensitive hyper- tension experimental animal models were constructed by subcutaneous injection of capsaicin in 60 Dahl rats, and the rats were randomly divided into normal salt group, high salt group and UCMSCs transplantation group （n = 20 each） according to the random number table method. 0.3% sodium chloride diet was given in normal salt group;8% sodium chloride diet was given in high salt group and UCMSCs transplantation group. After 8 weeks of feeding, UCMSCs of 1 ml （3×10^6 PKH- 26-labeled UCMSCs/ml） were injected via rat tail vein for 3 days in UCMSCs transplantation group. At the end of 2 weeks of experiment, left ventricular ejection fraction （LVEF） and left ventricular short axis fractional shortening （FS） were meas- ured by echocardiography; left cardiac index was calculated by left ventricular weight/body weight; morphology of myocardial cells was observed after HE staining;collagen volume fraction（CVF） was calculated after Van Gieson（VG） staining by computer image analysis system;distribution of PKH-26 labeled UCMSCs in myocardial tissues was observed under fluores- cence microscope;myocardial cell apoptosis was detected by terminal-deoxynucleoitidyl transferase mediated nick end labe-ling（TUNEL） assay. Results In normal salt group, the myocardial cells were arranged in an orderly manner, and there were no inflammatory cell infiltration and fibrosis scar formation in myocardial interstitium. In high salt group, the myocardial ceils presented disorder arrangement, and macrophage infiltration increased obviously and combined with obvious fibrosis remodeling. In UCMSCs transplantation group, despite the inflammatory cell infiltration, the myocardial ceils arranged orderly,and fibrosis remodeling was lighter without fibrous scar formation compared with high salt group. In high salt group, LVEF and FS decreased significantly（ all P 〈 0.05 ）, and left cardiac index increased significantly （P 〈 0.05 ）compared with normal salt group. In UCMSCs transplantation group, LVEF and FS increased significantly（ all P 〈 0.05 ）, and left cardiac index decreased significantly（P 〈0.05 ）compared with high salt group. CVF rose in turn in the order of normal salt group, UCMSCs transplantation group, high salt group （P 〈 0.05 ）. More PKH-26 labeled UCMSCs were found only in myo- cardial tissues of experimental rats in UCMSCs transplantation group. The myocardial cell apoptosis rates in normal salt group, high salt group and UCMSCs transplantation group were 0%, （28.2 ± 6.3 ）% and （12.7±2.6 ）% , respectively, and the myocardial cell apoptosis rate in UCMSCs transplantation group decreased significantly compared with high salt group （P 〈 0.05）. Conclusions The transplantation of UCMSCs could alleviate myocardial fibrosis of salt-sensitive hypertensive rats, decrease collagen deposition and myocardial cell apoptosis, improve the rat heart function.