人牙源性iPSCs分化心肌细胞促进急性心肌梗死小鼠心肌修复的研究

doi:10.11958/20220106

天津医药 ›› 2022, Vol. 50 ›› Issue (8): 822-826.doi: 10.11958/20220106

人牙源性iPSCs分化心肌细胞促进急性心肌梗死小鼠心肌修复的研究

谭小兵¹(), 王儒仙², 吕美荣², 孙显凤², 吕娜英², 戴青原²^,^Δ()

1云南省第一人民医院口腔医学中心,昆明理工大学附属医院（邮编650032）
2昆明医科大学第一附属医院心内科

收稿日期:2022-01-20 修回日期:2022-04-26 出版日期:2022-08-15 发布日期:2022-08-12
通讯作者: 戴青原 E-mail:txb942005@163.com;dqy0823@163.com
作者简介:谭小兵（1974）,男,副主任医师,主要从事牙源性干细胞与再生方面研究。E-mail： txb942005@163.com
基金资助:
国家自然科学基金资助项目(82060074);云南省科技厅-昆明医科大学应用基础研究联合专项资助(2019FE001-(113));云南省高层次卫生计生技术人才培养经费资助(D-2018045)

The effect of cardiomyocytes derived from human dental-origin iPSCs on cardiac repair in murine model of acute myocardial infarction

TAN Xiaobing¹(), WANG Ruxian², LYU Meirong², SUN Xianfeng², LYU Naying², DAI Qingyuan²^,^Δ()

1 Center of Stomatology, the First People’s Hospital of Yunnan Province, the Affiliated Hospital of Kunming University of Science and Technology, Kunming 650032, China
2 Department of Cardiology, the First Affiliated Hospital of Kunming Medical University

Received:2022-01-20 Revised:2022-04-26 Published:2022-08-15 Online:2022-08-12
Contact: DAI Qingyuan E-mail:txb942005@163.com;dqy0823@163.com

谭小兵, 王儒仙, 吕美荣, 孙显凤, 吕娜英, 戴青原. 人牙源性iPSCs分化心肌细胞促进急性心肌梗死小鼠心肌修复的研究[J]. 天津医药, 2022, 50(8): 822-826.

TAN Xiaobing, WANG Ruxian, LYU Meirong, SUN Xianfeng, LYU Naying, DAI Qingyuan. The effect of cardiomyocytes derived from human dental-origin iPSCs on cardiac repair in murine model of acute myocardial infarction[J]. Tianjin Medical Journal, 2022, 50(8): 822-826.

摘要/Abstract

摘要：

目的探讨人牙源性诱导性多能干细胞（iPSCs）分化心肌细胞对急性心肌梗死（AMI）小鼠心功能的影响。方法采用仙台病毒诱导、单克隆挑选和无饲养层体系将人牙根尖乳头干细胞（SCAP）重编程为iPSCs,心肌分化培养基定向诱导。9只BALB/C雄性小鼠按照随机数字表法分为正常组（不干预）、AMI组（AMI造模）和实验组（AMI造模+iPSCs-CMs注射）,每组3只。通过冠状动脉结扎法建立急性心肌梗死小鼠模型,iPS分化心肌细胞行心肌内局部注射,超声心动图评估心脏功能。结果人SCAP-iPSCs呈现典型胚胎干细胞样克隆形态,体内具备多向分化潜能,分化心肌细胞具有自主搏动性,表达特异性标志物α-辅肌动蛋白（α-actinin）和肌钙蛋白T（TNNT-2）。术后即刻心电图和超声心动图证实AMI模型成功。与正常组相比,AMI组左心室短轴缩短率（LVFS）和左心室射血分数（LVEF）明显下降,实验组LVFS和LVEF显著高于AMI组（P<0.05）。结论人牙源性iPSCs分化心肌细胞局部注射可以明显改善AMI小鼠的心功能,促进心肌修复。

关键词: 心肌梗死, 诱导多能干细胞, 肌细胞,心脏, 小鼠,近交BALB C, 心功能, 心肌修复, 人根尖乳头干细胞

Abstract:

Objective To investigate the effect of myocardial cells differentiated from human SCAP-induced pluripotent stem cells (iPSCs) on the myocardial function of acute myocardial infarction (AMI) mice. Methods Human stem cells from apical papilla (SCAP) was reprogrammed to iPSCs with Sendai reprogramming kit and passed using single colony subcloning under feeder-free condition. Specific myocardial differentiation medium was utilized to induce iPSCs. Nine BALB/C male mice were divided into the normal group (no intervention), the AMI group (AMI modeling) and the experimental group (AMI modeling + iPSCs-CMs injection) using random number table method. AMI mice model was established by coronary artery ligation. The derived cardiomyocytes (CMs) were injected into peri-infarct region of mice, and myocardial function was accessed by echocardiography. Results The derived iPSCs exhibited morphology resembling with embryonic stem cells (ESCs) and formed three-germ layers in nude mice. The CMs were able to beat independently and expressed specific markers, α-actinin and TNNT-2. The success of AMI model was confirmed by immediate electrocardiogram after operation. Compared with the normal group, LVFS and LVEF decreased significantly in the AMI group, and LVFS and LVEF were significantly higher in the experimental group than those in the AMI group (P<0.05). Conclusion Local injection of human odontogenic iPSCs differentiated cardiomyocytes can significantly improve the cardiac function and promote myocardial repair in AMI mice.

Key words: myocardial infarction, induced pluripotent stem cells, myocytes, cardiac, mice, inbred BALB C, cardiac function, cardiac repair, stem cells from apical papilla

中图分类号:

R541.4
R781

图/表 4

图1 人SCAP-iPSCs重编程过程及畸胎瘤实验 A：人SCAP（×100）;B：重编程小克隆（×50）;C：hSCAP-iPSCs克隆（×100）;D、E、F分别为内胚层、中胚层及外胚层：畸胎瘤HE染色（×100）。

Fig.1 Reprogramming process of human SCAP-iPSCs and teratoma assay

图2 人SCAP-iPSCs分化心肌细胞标志物免疫荧光染色（×400） A：TNNT-2;B：细胞核;C：重合;D：α-actinin;E：细胞核;F：重合。

Fig.2 Immunofluorescence staining of differentiated hiPSCs-CMs (×400)

图3 AMI动物模型及细胞注射后3组超声心动图检查结果 A：AMI造模即刻心电图;B：正常组超声心动图;C：AMI组超声心动图;D：实验组超声心动图。

Fig.3 AMI model and echocardiography results after CMs injection of the three groups

图4 细胞注射后3组超声心动图检查结果比较 n=3,a与正常组比较,b与AMI组比较,P<0.05。

Fig.4 Comparison of echocardiography results after CMs injection between the three groups

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人牙源性iPSCs分化心肌细胞促进急性心肌梗死小鼠心肌修复的研究

The effect of cardiomyocytes derived from human dental-origin iPSCs on cardiac repair in murine model of acute myocardial infarction

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