麻黄碱及阿托品对人诱导多能干细胞来源心肌细胞动作电位的影响  

Effects of Ephedrine and Atropine on Action Potential of Human Induced Pluripotent Stem Cell Derived Cardiomyocytes

作　　者：宫艺其 杨礼 艾雪峰 颜冰倩 谭瑶 王会景 徐徐 付炜 王伟[1] GONG Yiqi;YANG Li;AI Xuefeng;YAN Bingqian;TAN Yao;WANG Huijing;XU Xu;FU Wei;WANG Wei(Department of Pediatric Cardiothoracic Surgery,Shanghai Children's Medical Center,Shanghai Jiao Tong University School of Medicine,Shanghai 200127,China;Department of Anesthesiology,Shanghai Cancer Center,Shanghai Medical College,Fudan University,Shanghai 200032,China;Institute of Pediatric Translational Medicine,Shanghai Children's Medical Center,Shanghai Jiao Tong University School of Medicine,Shanghai 200127,China;Shanghai Key Laboratory of Tissue Engineering,Shanghai 200011,China.)

机构地区：[1]上海交通大学医学院附属上海儿童医学中心心胸外科,上海市200127 [2]复旦大学附属肿瘤医院麻醉科,上海市200032 [3]上海交通大学医学院附属上海儿童医学中心儿科转化医学研究所,上海市200127 [4]上海市组织工程研究重点实验室,上海市200011

出　　处：《组织工程与重建外科杂志》2020年第2期75-80,共6页Journal of Tissue Engineering and Reconstructive Surgery

基　　金：国家自然科学基金项目(81670372);上海市浦江人才计划(18PJD031);上海交通大学医学院转化医学协同创新中心合作研究项目(TM201821);上海交通大学医工交叉项目(YG2016MS28);上海市组织工程重点实验室开放课题;上海交通大学医学院儿科硕士研究生临床科研能力提升计划(EKKY2018003DGD)。

摘　　要：目的探讨麻黄碱及阿托品对人诱导多能干细胞来源心肌细胞动作电位的影响,为人诱导多能干细胞来源心肌细胞的临床应用提供依据。方法利用免疫荧光鉴定人诱导多能干细胞多能性,并利用CHIR99021及IWP-2小分子在体外将其分化为心肌细胞。利用免疫荧光及膜片钳对获得的心肌细胞进行形态学及电生理鉴定。随后通过灌流系统添加500μmol/L麻黄碱及100μmol/L阿托品,并用膜片钳记录动作电位的变化。结果人诱导多能干细胞高表达多能性标志物oct4、sox2、nanog及tra-1-60。分化得到的心肌细胞高表达ctnt、α-actinin、mlc2v等心肌特异性标志物,同时具有自发节律的动作电位。麻黄碱及阿托品能够显著加快自发动作电位的频率,但对动作电位去极化幅度、最大去极化速率及90%动作电位时程无明显影响。结论麻黄碱及阿托品能够加快人诱导多能干细胞来源心肌细胞自发动作电位的频率,证明其具有良好的药物反应性。Objective To explore the effects of ephedrine and atropine on the action potential of human induced pluripotent stem cell-derived cardiomyocytes,and provide a basis for the clinical application of human induced pluripotent stem cell-derived cardiomyocytes in the future.Methods Immunofluorescence was used to identify the pluripotency of human induced pluripotent stem cells,and CHIR99021 and IWP-2 were used to implement the cardiac differentiation process in vitro.Morphological and electrophysiological identification of the obtained cardiomyocytes were performed by immunofluorescence and patch clamp.Then 500μmol/L ephedrine and 100μmol/L atropine were subsequently added through a perfusion system,and changes in action potential were recorded with patch clamp.Results Human induced pluripotent stem cells highly expressed the pluripotency markers oct4,sox2,nanog and tra-1-60.The obtained cardiomyocytes highly expressed the myocardial specific markers ctnt,α-actinin and mlc2v,as well as possessed spontaneously rhythmic action potential.Ephedrine and atropine significantly speeded up the frequency of spontaneous action potential,but had no effect on APA,d V/d tmax and APD 90.Conclusion Ephedrine and atropine can accelerate the frequency of spontaneous action potential of human induced pluripotent stem cell-derived cardiomyocytes,which proves that these kinds of cardiomyocytes have good drug reactivity.

关 键 词：人诱导多能干细胞 心肌细胞 电生理 动作电位 麻黄碱 阿托品 

分 类 号：R318.11[医药卫生—生物医学工程]