心肌裂解液体外诱导人羊膜间充质细胞分化过程中离子通道表达的实验研究
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摘要
研究背景
心肌细胞损伤后不能再生,以干细胞为基础的细胞治疗有望修复损伤心肌。在心肌梗死后干细胞治疗中细胞来源与评价一直是该领域研究热点。近年来,采用羊膜间充质细胞(Amniotic Membrane-human Mesenchymal Stromal Cells AM-hMSCs)进行干细胞治疗受到关注,这些看似有应用前景的细胞在体外诱导分化后是否具有心肌细胞的电生理特点仍缺乏研究。
实验目的
1.明确在诱导分化前,AM-hMSCs上表面离子电流的分布特征以及相应离子通道蛋白表达
2.在心肌裂解液诱导作用下,AM-hMSCs在诱导分化过程中,细胞表面离子通道的变化情况。
实验方法
取羊膜组织,经胶原酶消化,获得AM-hMSCs。根据不同培养条件,将AM-hMSCs分为对照组以及心肌裂解液诱导组。
采用RT-PCR方法检测两组细胞离子通道蛋白mRNA的表达。通过膜片钳技术,记录细胞表面离子电流。比较两组细胞离子电流分布和离子通道蛋白mRNA表达的差异。
实验结果
在诱导分化前的AM-hMSCs中,共可记录到三种外向电流,其中外向延迟整流钾电流(IkDR)占90%,钙激活钾电流(IkCa)占43.3%,瞬时外向电流(Ito)占16.7%。实验中没有记录到内向电流。
经心肌裂解液诱导培养后,AM-hMSCs上Ito电流明显增加(诱导前16.7%与诱导后30.7%,p=0.04),并且有8%的细胞上可以记录到超极化激活的阳离子流(funny current,If)。
对离子通道mRNA的研究显示,在诱导分化前的AM-hMSCs上可以检测到一定水平的MaxIK, Kir2.1, Kvl.4, HminK通道蛋白mRNA的表达。经心肌裂解液诱导分化后,介导If电流的HCN2通道蛋白可表达阳性。
实验结论
本研究首次证实在诱导前的AM-hMSC上存在三种个不同的外向离子通道电流,分别为Ikca,IKDR和Ito,并且可以检测到相应离子通道蛋白mRNA的表达,与记录到的离子流相对应,分别是MaxiK(IkCa),HminK(IKDR),kvl.4(Ito)。
经心肌裂解液体外诱导培养,可促使AM-hMSCs分化,表达起搏通道蛋白HCN2,形成If电流,并且增加细胞表面的Ito电流。
Background:
The Amniotic Membrane-human Mesenchymal Stromal Cells (AM-hMSCs) from amniotic membrane are new ideal cell sources for cardiomyoplasty; however, the property and characteristic of the AM-hMSC electrophysiology were not well understood.
Purpose:1. The study is to determine the distribution of ion channel and property of ion current in AM-hMSCs.
2. To study the development and differentiation of ion current in AM-hMSCs under the cardiac microenvironment by using myocardial cell lysate.
Method:
The amniotic membrane was gathered and digested to get mesenchymal stem cell. The isolated AM-hMSCs were cultured in different medium of DMEM with or without myocardial cell lysate. The mRNA of AM-hMSC was collected after2week, and the Reverse transcriptase-polymerase chain reaction (RT-PCR) were then performed to detect the mRNA expression of ion channel encoding subunits. The ionic current was recorded under the whole cell mode with patch clamp.
Result:
There were3types outward could be recorded in the AM-hMSCs, including the delayed rectifier potassium current (IKDR)(90%), Calcium-activated-potassium current (Ikca)(43.3%), and transient outward current (Ito)(16.7%). The inward currents were absent.
Cultured in the medium with myocardial cell lysate, the induced AM-hMSCs can initiate the If current expression and increase the expression of Ito current (16.7%vs.30.7%, p=0.04).
The expression of mRNA of certain ion channel subunit can be detected in AM-hMSCs, which reveal the molecular evidence of ion current. The ion channel mRNA were detected in the AM-hMSCs, including the MaxIK (for IkCa), Kir2.1, Kv1.4(for Ito), HminK (for IKDR.).The mRNA expressions of the ion channel of HCN2(for If)can be detected in the AM-hMSCs cultured with myocardial cell lysate.
Conclusion:
This is the first study of the electrophysiology property in AM-hMSCs. The AM-hMSC can express a distinct pattern of ion channel mRNA and functional ion channels, including MaxIK (for IkCa), Kir2.1, Kv1.4(for Ito), HminK(for IkDR.).The AM-hMSCs cultured under the microenvironmen by using myocardial lysate can initiate the If current and HCN2expression, and increase the expression of Ito.
心肌细胞损伤后不能再生,以干细胞为基础的细胞治疗有望修复损伤心肌。在心肌梗死后干细胞治疗中细胞来源与评价一直是该领域研究热点。近年来,采用羊膜间充质细胞(Amniotic Membrane-human Mesenchymal Stromal Cells AM-hMSCs)进行干细胞治疗受到关注,这些看似有应用前景的细胞在体外诱导分化后是否具有心肌细胞的电生理特点仍缺乏研究。
实验目的
1.明确在诱导分化前,AM-hMSCs上表面离子电流的分布特征以及相应离子通道蛋白表达
2.在心肌裂解液诱导作用下,AM-hMSCs在诱导分化过程中,细胞表面离子通道的变化情况。
实验方法
取羊膜组织,经胶原酶消化,获得AM-hMSCs。根据不同培养条件,将AM-hMSCs分为对照组以及心肌裂解液诱导组。
采用RT-PCR方法检测两组细胞离子通道蛋白mRNA的表达。通过膜片钳技术,记录细胞表面离子电流。比较两组细胞离子电流分布和离子通道蛋白mRNA表达的差异。
实验结果
在诱导分化前的AM-hMSCs中,共可记录到三种外向电流,其中外向延迟整流钾电流(IkDR)占90%,钙激活钾电流(IkCa)占43.3%,瞬时外向电流(Ito)占16.7%。实验中没有记录到内向电流。
经心肌裂解液诱导培养后,AM-hMSCs上Ito电流明显增加(诱导前16.7%与诱导后30.7%,p=0.04),并且有8%的细胞上可以记录到超极化激活的阳离子流(funny current,If)。
对离子通道mRNA的研究显示,在诱导分化前的AM-hMSCs上可以检测到一定水平的MaxIK, Kir2.1, Kvl.4, HminK通道蛋白mRNA的表达。经心肌裂解液诱导分化后,介导If电流的HCN2通道蛋白可表达阳性。
实验结论
本研究首次证实在诱导前的AM-hMSC上存在三种个不同的外向离子通道电流,分别为Ikca,IKDR和Ito,并且可以检测到相应离子通道蛋白mRNA的表达,与记录到的离子流相对应,分别是MaxiK(IkCa),HminK(IKDR),kvl.4(Ito)。
经心肌裂解液体外诱导培养,可促使AM-hMSCs分化,表达起搏通道蛋白HCN2,形成If电流,并且增加细胞表面的Ito电流。
Background:
The Amniotic Membrane-human Mesenchymal Stromal Cells (AM-hMSCs) from amniotic membrane are new ideal cell sources for cardiomyoplasty; however, the property and characteristic of the AM-hMSC electrophysiology were not well understood.
Purpose:1. The study is to determine the distribution of ion channel and property of ion current in AM-hMSCs.
2. To study the development and differentiation of ion current in AM-hMSCs under the cardiac microenvironment by using myocardial cell lysate.
Method:
The amniotic membrane was gathered and digested to get mesenchymal stem cell. The isolated AM-hMSCs were cultured in different medium of DMEM with or without myocardial cell lysate. The mRNA of AM-hMSC was collected after2week, and the Reverse transcriptase-polymerase chain reaction (RT-PCR) were then performed to detect the mRNA expression of ion channel encoding subunits. The ionic current was recorded under the whole cell mode with patch clamp.
Result:
There were3types outward could be recorded in the AM-hMSCs, including the delayed rectifier potassium current (IKDR)(90%), Calcium-activated-potassium current (Ikca)(43.3%), and transient outward current (Ito)(16.7%). The inward currents were absent.
Cultured in the medium with myocardial cell lysate, the induced AM-hMSCs can initiate the If current expression and increase the expression of Ito current (16.7%vs.30.7%, p=0.04).
The expression of mRNA of certain ion channel subunit can be detected in AM-hMSCs, which reveal the molecular evidence of ion current. The ion channel mRNA were detected in the AM-hMSCs, including the MaxIK (for IkCa), Kir2.1, Kv1.4(for Ito), HminK (for IKDR.).The mRNA expressions of the ion channel of HCN2(for If)can be detected in the AM-hMSCs cultured with myocardial cell lysate.
Conclusion:
This is the first study of the electrophysiology property in AM-hMSCs. The AM-hMSC can express a distinct pattern of ion channel mRNA and functional ion channels, including MaxIK (for IkCa), Kir2.1, Kv1.4(for Ito), HminK(for IkDR.).The AM-hMSCs cultured under the microenvironmen by using myocardial lysate can initiate the If current and HCN2expression, and increase the expression of Ito.
引文
1.Hellermann, J. P.; Goraya, T. Y.; Jacobsen, S. J.; Weston, S. A.; Reeder, G S.; Gersh, B. J.; Redfield, M. M.; Rodeheffer, R. J.; Yawn, B. P. & Roger, V. L. Incidence of heart failure after myocardial infarction: is it changing over time? Am J Epidemiol, 2003; 157: 1101-1107.
2.Jonathan Leor, Stem Cell and Gene-Based Therapy: Frontiers in Regenerative Medicine, Berlin,springer, 2005:3-4.
3.Itskovitz-Eldor, J.; Schuldiner, M.; Karsenti, D.; Eden, A.; Yanuka, O.; Amit, M.; Soreq, H. & Benvenisty, N. Differentiation of human embryonic stem cells into embryoid bodies compromising the three embryonic germ layers. Mol Med, 2000; 6:88-95.
4.Kehat, I.; Kenyagin-Karsenti, D.; Snir, M.; Segev, H.; Amit, M.; Gepstein, A.; Livne, E.; Binah, O.; Itskovitz-Eldor, J. & Gepstein, L. Human embryonic stem cells can differentiate into myocytes with structural and functional properties of cardiomyocytes. J Clin Invest, 2001; 108: 407-414.
5.Xu, C; Police, S.; Rao, N. & Carpenter, M. K. Characterization and enrichment of cardiomyocytes derived from human embryonic stem cells. CircRes,2002;91: 501-508.
6.Xu, C; Lebkowski, J. & Gold, J. D. Growth and differentiation of human embryonic stem cells for cardiac cell replacement therapy. Curr Stem Cell Res Ther, 2006; 1:173-187.
7.He, J.; Ma, Y; Lee, Y; Thomson, J. A. & Kamp, T. J. Human embryonic stem cells develop into multiple types of cardiac myocytes: action potential characterization. Circ Res, 2003; 93: 32-39.
8.Kehat, I.; Khimovich, L.; Caspi, O.; Gepstein, A.; Shofti, R.; Arbel, G; Huber, I.; Satin, J.; Itskovitz-Eldor, J. & Gepstein, L. Electromechanical integration of cardiomyocytes derived from human embryonic stem cells. Nat Biotechnol, 2004; 22: 1282-1289.
9.Xu, W.; Zhang, X.; Qian, H.; Zhu, W.; Sun, X.; Hu, J.; Zhou, H. & Chen, Y. Mesenchymal stem cells from adult human bone marrow differentiate into a cardiomyocyte phenotype in vitro. Exp Biol Med (Maywood), 2004; 229: 623-631.
10. Antonitsis, P.; Ioannidou-Papagiannaki, E.; Kaidoglou, A.; Charokopos, N.; Kalogeridis, A.; Kouzi-Koliakou, K.; Kyriakopoulou, I.; Klonizakis, I. & Papakonstantinou, C. Cardiomyogenic potential of human adult bone marrow mesenchymal stem cells in vitro. Thorac Cardiovasc Surg, 2008;56: 77-82.
11.Antonitsis, P.; Ioannidou-Papagiannaki, E.; Kaidoglou, A. & Papakonstantinou, C. In vitro cardiomyogenic differentiation of adult human bone marrow mesenchymal stem cells. The role of 5-azacytidine. Interact Cardiovasc Thorac Surg, 2007; 6:593-597.
12.Makino, S.; Fukuda, K.; Miyoshi, S.; Konishi, R; Kodama, H.; Pan, J.; Sano, M.; Takahashi, T; Hori, S.; Abe, H.; Hata, J.; Umezawa, A. & Ogawa, S. Cardiomyocytes can be generated from marrow stromal cells in vitro. J Clin Invest, 1999; 103:697-705.
13.Hakuno, D.; Fukuda, K.; Makino, S.; Konishi, R; Tomita, Y.; Manabe, T; Suzuki, Y; Umezawa, A. & Ogawa, S. Bone marrow-derived regenerated cardiomyocytes (CMG Cells) express functional adrenergic and muscarinic receptors. Circulation, 2002; 105: 380-386.
14. Cao, R; Niu, L. L.; Meng, L.; Zhao, L. X.; Zheng, M.; Yue, W.; Bai, C. X.; Jia, G L. & Pei, X. T. [Cardiomyocyte-like differentiation of human bone marrow mesenchymal stem cells after exposure of 5-azacytidine in vitro] Shi Yan Sheng Wu Xue Bao, 2004; 37: 118-124.
15. Liu, Y; Song, J.; Liu, W.; Wan, Y; Chen, X. & Hu, C. Growth and differentiation of rat bone marrow stromal cells: does 5-azacytidine trigger their cardiomyogenic differentiation? Cardiovasc Res, 2003; 58: 460-468.
16. Alviano, R.; Fossati, V; Marchionni, C.; Arpinati, M.; Bonsi, L.; Franchina, M.; Lanzoni, G.; Cantoni, S.; Cavallini, C.; Bianchi, F.; Tazzari, P. L. Pasquinelli, G.; Foroni, L.; Ventura, C.; Grossi, A. & Bagnara, G P. Term Amniotic membrane is a high throughput source for multipotent Mesenchymal Stem Cells with the ability to differentiate into endothelial cells in vitro. BMCDev Biol, 2007; 7:11.
17. Sakuragawa, N.; Thangavel, R.; Mizuguchi, M.; Hirasawa, M. & Kamo, I. Expression of markers for both neuronal and glial cells in human amniotic epithelial cells. Neurosci Lett, 1996; 209: 9-12.
18.Ishii, T.; Ohsugi, K.; Nakamura, S.; Sato, K.; Hashimoto, M.; Mikoshiba, K. & Sakuragawa, N. Gene expression of oligodendrocyte markers in human amniotic epithelial cells using neural cell-type-specific expression system. Neurosci Lett, 1999;268:131-134.
19. Zhang, Y.; Li, C; Jiang, X.; Zhang, S.; Wu, Y.; Liu, B.; Tang, P. & Mao, N. Human placenta-derived mesenchymal progenitor cells support culture expansion of long-term culture-initiating cells from cord blood CD34+ cells. Exp Hematol, 2004; 32: 657-664.
20. Yen, B. L.; Huang, H.; Chien, C; Jui, H.; Ko, B.; Yao, M.; Shun, C.; Yen, M; Lee, M. & Chen, Y. Isolation of multipotent cells from human term placenta. Stem Cells, 2005; 23: 3-9.
21. In't Anker, P. S. I.; Scherjon, S. A.; der Keur, C. K.; de Groot-Swings, G M. J. S.; Claas, F. H. J.; Fibbe, W. E. & Kanhai, H. H. H. Isolation of mesenchymal stem cells of fetal or maternal origin from human placenta. Stem Cells, 2004; 22:1338-1345.
22. Zhao, P.; Ise, H.; Hongo, M; Ota, M.; Konishi, I. & Nikaido, T. Human amniotic mesenchymal cells have some characteristics of cardiomyocytes. Transplantation,2005;79:528-535.
23. Rooney, I. A. & Morgan, B. P. Characterization of the membrane attack complex inhibitory protein CD59 antigen on human amniotic cells and in amniotic fluid. Immunology,1992; 76:541-547.
24. Soncini, M.; Vertua, E.; Gibelli, L.; Zorzi, F.; Denegri, M.; Albertini, A.; Wengler, G S. & Parolini, O. Isolation and characterization of mesenchymal cells from human fetal membranes. J Tissue Eng Regen Med,2007; 1: 296-305.
25. Horwitz, E. M.; Blanc, K. Le; Dominici, M.; Mueller, I.; Slaper-Cortenbach, I.; Marini, F. C.; Deans, R. J.; Krause, D. S.; Keating, A. & the International Society for Cellular Therapy Clarification of the nomenclature for MSC:The International Society for Cellular Therapy position statement. Cytotherapy, 2005; 7:393-395.
26. Cohen, I. S.; Rosen, A. B. & Gaudette, G R. A Caveat Emptor for myocardial regeneration:mechanical without electrical recovery will not suffice. J Mol Cell Cardiol,2007; 42:285-288.
27. Chang, M. G.; Tung, L.; Sekar, R. B.; Chang, C. Y.; Cysyk, J.; Dong, P.; Marban, E. & Abraham, M. R. Proarrhythmic potential of mesenchymal stem cell transplantation revealed in an in vitro coculture model. Circulation,2006; 113:1832-1841.
28. Ben-Dor, I.; Fuchs, S. & Kornowski, R. Potential hazards and technical considerations associated with myocardial cell transplantation protocols for ischemic myocardial syndrome. J Am Coll Cardiol,2006;48:1519-1526
29. Schram, G.; Pourrier, M.; Melnyk, P. & Nattel, S. Differential distribution of cardiac ion channel expression as a basis for regional specialization in electrical function. Circ Res,2002; 90:939-950.
30.李澈.心肌细胞膜上的离子通道.见：李澈主编.窦房结.北京：北京医 科大学出版社,2001.69-102.
31.刘泰槰.离子通道与离子流.见：刘泰槰主编.心肌细胞点生理学—离子通道,离子载体和离子流.北京：人民卫生出版社,2005.30-46.
32.刘泰槰.心肌细胞离子流和离子通道研究简史.见：刘泰槰主编.心肌细胞离子通道和离子通道病.北京：人民卫生出版社,2006.1-5.
33.李泱,程芮细胞电生理学.见：李泱,程芮主编.离子通道学.湖北：湖北科学技术出版社,2007.3-88.
34. Kawano, S.; Otsu, K.; Shoji, S.; Yamagata, K. & Hiraoka, M. Ca(2+) oscillations regulated by Na+-Ca+ exchanger and plasma membrane Ca(2+) pump induce fluctuations of membrane currents and potentials in human mesenchymal stem cells. Cell Calcium,2003;34:145-156.
35. Park, K. S.; Jung, K. H.; Kim, S. H.; Kim, K. S.; Choi, M. R.; Kim, Y. & Chai, Y. G. Functional expression of ion channels in mesenchymal stem cells derived from umbilical cord vein. Stem Cells,2007; 25:2044-2052.
36. J. J. B. Jack, D. Noble, R. W. Tsien. Electric current flow in excitable cells. Oxford Science Publications; 1983:229.
37. Fauza, D. Amniotic fluid and placental stem cells. Best Pract Res Clin Obstet Gynaecol,2004; 18:877-891.
38. Marcus, A. J.; Coyne, T. M.; Rauch, J.; Woodbury, D. & Black, I. B. Isolation, characterization, and differentiation of stem cells derived from the rat amniotic membrane. Differentiation,2008; 76:130-144.
39. Miki, T.; Lehmann, T.; Cai, H.; Stolz, D. B. & Strom, S. C. Stem cell characteristics of amniotic epithelial cells. Stem Cells,2005; 23:1549-1559.
40.林雪.羊膜间质干细胞的基本性质和向心肌细胞诱导分化的立体培养系统研究.北京：中国协和医科大学,2007.
41. Pasquinelli, G.; Tazzari, P.; Ricci, F.; Vaselli, C.; Buzzi, M.; Conte, R.; Orrico, C.; Foroni, L.; Stella, A.; Alviano, F.; Bagnara, G. P. Ultrastructural characteristics of human mesenchymal stromal (stem) cells derived from bone marrow and term placenta. Ultrastruct Pathol,2007; 31:23-31.
42. Kim, J.; Kang, H. M.; Kim, H.; Kim, M. R.; Kwon, H. C.; Gye, M. C.; Kang, S. G.; Yang, H. S. & You, J. Ex vivo characteristics of human amniotic membrane-derived stem cells. Cloning Stem Cells,2007; 9:581-594.
43. Kunisaki, S. M.; Armant, M.; Kao, G. S.; Stevenson, K.; Kim, H. & Fauza, D. O. Tissue engineering from human mesenchymal amniocytes:a prelude to clinical trials. J Pediatr Surg,2007; 42:974-979.
44. Heubach, J. F.; Graf, E. M.; Leutheuser, J.; Bock, M.; Balana, B.; Zahanich, I.; Christ, T.; Boxberger, S.; Wettwer, E. & Ravens, U. Electrophysiological properties of human mesenchymal stem cells. J Physiol,2004; 554:659-672.
45. Li, G.; Sun, H.; Deng, X. & Lau, C. Characterization of ionic currents in human mesenchymal stem cells from bone marrow. Stem Cells,2005; 23: 371-382.
46. Bai, X.; Ma, J.; Pan, Z.; Song, Y; Freyberg, S.; Yan, Y.; Vykoukal, D. & Alt, E. Electrophysiological properties of human adipose tissue-derived stem cells. Am J Physiol Cell Physiol,2007;293:C1539-C1550.
47. Brahmajothi MV, Campbell DL, Rasmusson RL, et al. Distinct transient outward potassium current (Ito) phenotypes and distribution of fast-inactivating potassium channel alpha subunits in ferret left ventricular myocytes. J Gen Physiol.1999; 113:581-600.
48. Campbell, D.L., R.L. Rasmusson, Y Qu, and H.C. Strauss. The calcium-independent transient outward potassium current in isolated ferret right ventricular myocytes. I. Basic characterization and kinetic analysis. J. Gen. Physiol.1993; 101:571-601.
49. Campbell, D.L., Y Qu, R.L. Rasmusson, and H.C. Strauss. The calcium-independent transient outward potassium current in isolated ferret right ventricular myocytes. Ⅱ. Closed state reverse use-dependent block by 4-aminopyridine. J. Gen. Physiol.1993; 101:603-626.
50. Gryschenko, O.; Lu, Z. J.; Fleischmann, B. K. & Hescheler, J. Outwards currents in embryonic stem cell-derived cardiomyocytes. Pflugers Arch, 2000;439:798-807.
51. Wei, A. D.; Gutman, G. A.; Aldrich, R.; Chandy, K. G.; Grissmer, S. & Wulff, H. International Union of Pharmacology. LⅡ. Nomenclature and molecular relationships of calcium-activated potassium channels. Pharmacol Rev, 2005;57:463-472.
52. Cui, D.H. Cox, and R.W. Aldrich. Intrinsic Voltage Dependence and Ca2+ Regulation of mslo Large Conductance Ca-activated K+ Channels J. Gen. Physiol.1997; 109:647-673.
53. Hamada E, Nakajima T, Ota S, Terano A, Omata M, Nakade S, Mikoshiba K, Kurachi Y. Activation of Ca(2+)-dependent K+ current by acetylcholine and histamine in a human gastric epithelial cell line. J Gen Physiol.1993; 102:667-692.
54. Weerapura, M.; Nattel, S.; Chartier, D.; Caballero, R. & Hebert, T. E. A comparison of currents carried by HERG, with and without coexpression of MiRP1, and the native rapid delayed rectifier current. Is MiRP10 the missing link? J Physiol,2002; 540:15-27.
55. Federico Sesti and Steve A.N. Goldstein Single-Channel Characteristics of Wild-Type IKs Channels and Channels formed with Two MinK Mutants that Cause Long QT Syndrome J. Gen. Physiol.1998;112:651-663.
56. Sesti, F. & Goldstein, S. A. Single-channel characteristics of wild-type IKs channels and channels formed with two minK mutants that cause long QT syndrome. J Gen Physiol,1998; 112:651-663.
57. Grissmer S, Nguyen AN, Aiyar J, Hanson DC, Mather RJ, Gutman GA, Karmilowicz MJ, Auperin DD, Chandy KG. Pharmacological characterization of five cloned voltage-gated K+channels, types Kvl.1,1.2, 1.3,1.5, and 3.1, stably expressed in mammalian cell lines. Mol Pharmacol. 1994; 45:1227-1234.
58. Kawano, S.; Shoji, S.; Ichinose, S.; Yamagata, K.; Tagami, M. & Hiraoka, M. Characterization of Ca2+ signaling pathways in human mesenchymal stem cells. Cell Calcium,2002; 32:165-174.
59. Deng, X.; Sun, H.; Lau, C. & Li, G. Properties of ion channels in rabbit mesenchymal stem cells from bone marrow. Biochem Biophys Res Commun, 2006;348:301-309
60. Li, G; Deng, X.; Sun, H.; Chung, S. S. M.; Tse, H. & Lau, C. Ion channels in mesenchymal stem cells from rat bone marrow. Stem Cells, 2006;24:1519-1528.
61. Tao, R.; Lau, C.; Tse, H. & Li, G Functional ion channels in mouse bone marrow mesenchymal stem cells. Am J Physiol Cell Physiol,2007;293: C1561-C1567.
62. Wang, K.; Xue, T.; Tsang, S.; Huizen, R. V.; Wong, C. W.; Lai, K. W.; Ye, Z.; Cheng, L.; Au, K. W.; Zhang, J.; Li, G.; Lau, C.; Tse, H. & Li, R. A. Electrophysiological properties of pluripotent human and mouse embryonic stem cells. Stem Cells,2005;23:1526-1534.
63. Wonderlin, W. F. & Strobl, J. S. Potassium channels, proliferation and G1 progression. J Membr Biol,1996; 154:91-107.
64. Czarnecki, A.; Vaur, S.; Dufy-Barbe, L.; Dufy, B. & Bresson-Bepoldin, L. Cell cycle-related changes in transient K(+) current density in the GH3 pituitary cell line. Am J Physiol Cell Physiol,2000; 279:C1819-C1828.
65. Czarnecki, A.; Dufy-Barbe, L.; Huet, S.; Odessa, M. & Bresson-Bepoldin, L. Potassium channel expression level is dependent on the proliferation state in the GH3 pituitary cell line. Am J Physiol Cell Physiol,2003; 284: C1054-C1064.
66. Boullin, J. & Morgan, J. M. The development of cardiac rhythm. Heart,2005; 91:874-875.
67. Moorman AF, Christoffels VM. Cardiac Chamber Formation:Development, Genes, and Evolution. Physiol Rev.2003;83:1223-1267.
68. Moorman, A. F.; de Jong, F.; Denyn, M. M. & Lamers, W. H. Development of the cardiac conduction system. Circ Res,1998;82:629-644.
69. Pennisi, D. J.; Rentschler, S.; Gourdie, R. G.; Fishman, G I. & Mikawa, T. Induction and patterning of the cardiac conduction system. Int J Dev Biol, 2002; 46:765-775.
70. Cheng, G.; Litchenberg, W. H.; Cole, G J.; Mikawa, T.; Thompson, R. P. & Gourdie, R. G Development of the cardiac conduction system involves recruitment within a multipotent cardiomyogenic lineage. Development, 1999,126,5041-5049.
71. Tomita, Y.; Makino, S.; Hakuno, D.; Hattan, N.; Kimura, K.; Miyoshi, S.; Murata, M.; Ieda, M. & Fukuda, K. Application of mesenchymal stem cell-derived cardiomyocytes as bio-pacemakers:current status and problems to be solved. Med Biol Eng Comput,2007;45:209-220.
72. Donmez, A.; Zoghi, M.; Cagirgan, S.; Acarlar, C. & Tombuloglu, M. The effect of hematopoietic progenitor cells'temperature on cardiac arrhythmias in patients given peripheral blood progenitor cells. Transfus Apher Sci,2006, 34,245-251.
73. J. J. B. Jack, D. Noble, R. W. Tsien. Electric current flow in excitable cells. Oxford Science Publications; 1983:225-227.
74.袁岩,陈连凤,张抒扬,吴炜,陈浩,严晓伟心肌微环境对骨髓间充质干细胞的诱导分化作用.中国介入心脏病学杂志2005；13：315-318.
75.袁岩,陈连凤,张抒扬,吴炜,陈浩,严晓伟心肌裂解液对对骨髓间充质干细胞向心肌细胞分化诱导作用的研究.中国心血管病杂志2005；33：170-173.
76. Hirota, A.; Fujii, S. & Kamino, K. Optical monitoring of spontaneous electrical activity of 8-somite embryonic chick heart. Jpn J Physiol,1979;29: 635-639.
77. Kamino, K.; Hirota, A. & Fujii, S. Localization of pacemaking activity in early embryonic heart monitored using voltage-sensitive dye. Nature,1981; 290:595-597.
78. Viragh, S. & Challice, C. E. The development of the early atrioventricular conduction system in the embryonic heart. Can J Physiol Pharmacol, 1983;61:775-792.
79. Viragh, S. & Challice, C. E. The development of the conduction system in the mouse embryo heart. Dev Biol,1982;89:25-40.
80.李澈.心肌细胞膜上的离子通道,钾通道和钾电流.见：李澈主编.窦房结.北京：北京医科大学出版社,2001.79-88.
81.刘泰槰.瞬时外向电流Ito.见：刘泰槰主编.心肌细胞点生理学—离子通道,离子载体和离子流.北京：人民卫生出版社,2005.101-103.
82.李泱,程芮瞬时外向钾电流.见：李泱,程芮主编.离子通道学.湖北：湖北科学技术出版社,2007.49-54.
83. Crumb, W. J.; Pigott, J. D. & Clarkson, C. W. Comparison of Ito in young and adult human atrial myocytes:evidence for developmental changes. Am J Physiol,1995; 268:H1335-H134.
84. Sartiani, L.; Bettiol, E.; Stillitano, F.; Mugelli, A.; Cerbai, E. & Jaconi, M. E. Developmental changes in cardiomyocytes differentiated from human embryonic stem cells:a molecular and electrophysiological approach. Stem Cells,2007; 25:1136-1144.
85. Mills, W. R.; Mai, N.; Kiedrowski, M. J.; Unger, R.; Forudi, F.; Popovic, Z. B.; Penn, M. S. & Laurita, K. R. Stem cell therapy enhances electrical viability in myocardial infarction. J Mol Cell Cardiol,2007; 42:304-314.
86. Tang, Y. L.; Zhao, Q.; Qin, X.; Shen, L.; Cheng, L.; Ge, J. & Phillips, M. I. Paracrine action enhances the effects of autologous mesenchymal stem cell transplantation on vascular regeneration in rat model of myocardial infarction. Ann Thorac Surg,2005;80:229-236; discussion 236-237.
87. Cowan, D. B. & McGowan, F. X. A paradigm shift in cardiac pacing therapy? Circulation,2006; 114:986-988.
88. Rosen, M. R.; Brink, P. R.; Cohen, I. S. & Robinson, R. B. Genes, stem cells and biological pacemakers. Cardiovasc Res,2004; 64:12-23.
89. Boink, G. J. J.; Seppen, J.; de Bakker, J. M. T. & Tan, H. L. Gene therapy to create biological pacemakers. Med Biol Eng Comput,2007;45:167-176.
90. Anghel, T. M. & Pogwizd, S. M. Creating a cardiac pacemaker by gene therapy. Med Biol Eng Comput,2007;45:145-155.
91. Edelberg, J. M.; Aird, W. C. & Rosenberg, R. D. Enhancement of murine cardiac chronotropy by the molecular transfer of the human beta2 adrenergic receptor cDNA. J Clin Invest,1998; 101:337-343.
92. Miake, J.; Marban, E. & Nuss, H. B. Biological pacemaker created by gene transfer. Nature,2002;419:132-133.
93. Tan, H. L.; Bardai, A.; Shimizu, W.; Moss, A. J.; Schulze-Bahr, E.; Noda, T. & Wilde, A. A. M. Genotype-specific onset of arrhythmias in congenital long-QT syndrome:possible therapy implications. Circulation,2006; 114: 2096-2103.
94. Rosen, M. R.; Brink, P. R.; Cohen, I. S. & Robinson, R. B. Biological pacemakers based on If. Med Biol Eng Comput,2007;45:157-166.
95. Qu, J.; Barbuti, A.; Protas, L.; Santoro, B.; Cohen, I. S. & Robinson, R. B. HCN2 overexpression in newborn and adult ventricular myocytes:distinct effects on gating and excitability. Circ Res,2001;89:E8-14.
96. Qu, J.; Plotnikov, A. N.; Danilo, P.; Shlapakova, I.; Cohen, I. S.; Robinson, R. B. & Rosen, M. R. Expression and function of a biological pacemaker in canine heart. Circulation,2003,107,1106-1109.
97. Potapova I, Plotnikov A, Lu Z, Danilo P Jr, Valiunas V, Qu J, Doronin S, Zuckerman J, Shlapakova IN, Gao J, Pan Z, Herron AJ, Robinson RB, Brink PR, Rosen MR, Cohen IS. Human mesenchymal stem cells as a gene delivery system to create cardiac pacemakers. Circ Res.2004; 94:952-959.
98. Tse H-F, Xue T, Lau C-P, Siu C-W, Wang K, Zhang Q-Y, Tomaselli GF, Akar FG, Li RA. Bioartificial sinus node constructed via in vivo gene transfer of an engineered pacemaker HCN channel reduces the dependence on electronic pacemaker in a sick-sinus syndrome model. Circulation.2006; 114:1000-1011.
99. Gepstein, L. Stem cells as biological heart pacemakers. Expert Opin Biol Ther,2005;5:1531-153.
100. Lin G, Cai J, Jiang H, Shen H, Jiang X, Yu Q, Song J. Biological pacemaker created by fetal cardiomyocyte transplantation. J Biomed Sci.2005; 12:513-519.
101. Ruhparwar A, Tebbenjohanns J, Niehaus M, Mengel M, Irtel T, Kofidis T, Pichlmaier AM, Haverich A. Transplanted fetal cardiomyocytes as cardiac pacemaker. Eur J Cardiothorac Surg.2002; 21:853-857.
102. Cai J, Lin G, Jiang H, Yang B, Jiang X, Yu Q, Song J. Transplanted neonatal cardiomyocytes as a potential biological pacemaker in pigs with complete atrioventricular block. Transplantation.2006; 81:1022-1026.
103. Kehat I, Khimovich L, Caspi O, Gepstein A, Shofti R, Arbel G, Huber I, Satin J, Itskovitz-Eldor J, Gepstein L. Electromechanical integration of cardiomyocytes derived from human embryonic stem cells. Nat Biotechnol. 2004; 22:1282-1289.
104. Xue T, Cho HC, Akar FG, Tsang SY, Jones SP, Marban E, Tomaselli GF, Li RA. Functional integration of electrically active cardiac derivatives from genetically engineered human embryonic stem cells with quiescent recipient ventricular cardiomyocytes:insights into the development of cell-based pacemakers. Circulation.2005; 111:11-20.
105. Choi YH, Stamm C, Hammer PE, Kwaku KF, Marler JJ, Friehs I, Jones M, Rader CM, Roy N, Eddy MT, Triedman JK, Walsh EP, McGowan FX Jr, del Nido PJ, Cowan DB. Cardiac conduction through engineered tissue. Am J Pathol.2006; 1691:72-85.
106. Zhou, Y.; Yang, X.; Li, H.; Han, L. & Jiang, W. Mesenchymal stem cells transfected with HCN2 genes by LentiV can be modified to be cardiac pacemaker cells. Med Hypotheses,2007; 69:1093-1097.
107. Zhou, Y.; Yang, X. & Li, H. Hyperpolarization-activated cyclic nucleotide-gated channel gene:the most possible therapeutic applications in the field of cardiac biological pacemakers. Med Hypotheses,2007; 69: 541-544.
108. DiFrancesco, D.; Ferroni, A.; Mazzanti, M. & Tromba, C. Properties of the hyperpolarizing-activated current (If) in cells isolated from the rabbit sino-atrial node. J Physiol,1986; 377:61-88.
109. DiFrancesco, D. The contribution of the 'pacemaker' current (If) to generation of spontaneous activity in rabbit sino-atrial node myocytes. J Physiol,1991; 434:23-40.
1. Rosen, M. R.; Brink, P. R.; Cohen, I. S. & Robinson, R. B. Genes, stem cells and biological pacemakers. Cardiovasc Res,2004; 64:12-23.
2. Rosen, M. R.15th annual Gordon K. Moe Lecture. Biological pacemaking: in our lifetime? Heart Rhythm,2005; 2:418-428.
3. Gepatein L, Feld Y, Yankelson L. Somatic gene and cell therapy strategies for the treament of cardiac. Am J Physiol Heart Cir Physiol 2004;286:H815-H822.
4.李澈.窦房结的组织学和发生.见李澈主编.窦房结.北京：北京医科大学出版社,2001.9-24.
5.刘泰槰.心肌细胞的动作电位.见刘泰槰主编.心肌细胞点生理学—离子通道,离子载体和离子流.北京：人民卫生出版社,2005.17-27.
6. Brown, H. & Difrancesco, D. Voltage-clamp investigations of membrane currents underlying pace-maker activity in rabbit sino-atrial node. J Physiol, 1980; 308:331-351.
7. DiFrancesco, D. & Ojeda, C. Properties of the current If in the sino-atrial node of the rabbit compared with those of the current iK, in Purkinje fibres. J Physiol,1980;308:353-367.
8. DiFrancesco, D. Properties of the cardiac pacemaker (If current. Boll Soc Ital Biol Sper,1984; 60 Suppl 4:29-33.
9. Di Francesco D. The pacemaker current (If) Plays an important role in regulating SA node pacemaker activity.Cardiovasc Res,1995;30:307-308.
10. DiFrancesco, D. Serious workings of the funny current. Prog Biophys Mol Biol,2006; 90:13-25.
11. Santoro, B.; Grant, S. G.; Bartsch, D. & Kandel, E. R. Interactive cloning with the SH3 domain of N-src identifies a new brain specific ion channel protein, with homology to eag and cyclic nucleotide-gated channels. Proc Natl Acad Sci U S A,1997; 94:14815-1482.
12. Boullin, J. & Morgan, J. M. The development of cardiac rhythm. Heart, 2005;91:874-875.
13. Moorman AF, Christoffels VM. Cardiac Chamber Formation: Development, Genes, and Evolution. Physiol Rev.2003;83:1223-1267
14. Gorza L, Schiaffino S, Vitadello M. Heart conduction system:a neural crest derivative? Brain Res 1988;457:360-366.
15. Cheng, G.; Litchenberg, W. H.; Cole, G. J.; Mikawa, T.; Thompson, R. P. & Gourdie, R. G. Development of the cardiac conduction system involves recruitment within a multipotent cardiomyogenic lineage. Development, 1999; 126:5041-5049.
16. Gourdie RG, Mima T, Thompson RP, Mikawa T. Terminal diversification of the myocyte lineage generates Purkinje fibers of the cardiac conduction system. Development.1995; 121:1423-1431
17. Hirota, A.; Fujii, S. & Kamino, K. Optical monitoring of spontaneous electrical activity of 8-somite embryonic chick heart. Jpn J Physiol,1979; 29: 635-639.
18. Kamino, K.; Hirota, A. & Fujii, S. Localization of pacemaking activity in early embryonic heart monitored using voltage-sensitive dye. Nature,1981; 290:595-597.
19. Viragh, S. & Challice, C. E. The development of the conduction system in the mouse embryo heart. Dev Biol,1980; 80:28-45.
20. Pennisi, D. J.; Rentschler, S.; Gourdie, R. G.; Fishman, G. I. & Mikawa, T. Induction and patterning of the cardiac conduction system. Int J Dev Biol, 2002; 46:765-775.
21. Gourdie, R. G.; Wei, Y.; Kim, D.; Klatt, S. C. & Mikawa, T. Endothelin-induced conversion of embryonic heart muscle cells into impulse-conducting Purkinje fibers. Proc Natl Acad Sci U S A,1998; 95: 6815-6818.
22. Rentschler, S.; Vaidya, D. M.; Tamaddon, H.; Degenhardt, K.; Sassoon, D.; Morley, G. E.; Jalife, J. & Fishman, G. I. Visualization and functional characterization of the developing murine cardiac conduction system. Development,2001; 128:1785-1792.
23. Cowan, D. B. & McGowan, F. X. A paradigm shift in cardiac pacing therapy? Circulation,2006; 114:986-988.
24. Boink, G. J. J.; Seppen, J.; de Bakker, J. M. T. & Tan, H. L. Gene therapy to create biological pacemakers. Med Biol Eng Comput,2007; 45:167-176.
25. Anghel, T. M. & Pogwizd, S. M. Creating a cardiac pacemaker by gene therapy. Med Biol Eng Comput,2007;45:145-155.
26. Edelberg, J. M.; Aird, W. C. & Rosenberg, R. D. Enhancement of murine cardiac chronotropy by the molecular transfer of the human beta2 adrenergic receptor cDNA. J Clin Invest,1998; 101:337-343.
27. Miake, J.; Marban, E. & Nuss, H. B. Biological pacemaker created by gene transfer. Nature,2002; 419:132-133.
28. Tan, H. L.; Bardai, A.; Shimizu, W.; Moss, A. J.; Schulze-Bahr, E.; Noda, T. & Wilde, A. A. M. Genotype-specific onset of arrhythmias in congenital long-QT syndrome:possible therapy implications. Circulation,2006; 114: 2096-2103
29. Qu, J.; Plotnikov, A. N.; Danilo, P.; Shlapakova, I.; Cohen, I. S.; Robinson, R. B. & Rosen, M. R. Expression and function of a biological pacemaker in canine heart. Circulation,2003; 107:1106-1109.
30. Potapova I, Plotnikov A, Lu Z, Danilo P Jr, Valiunas V, Qu J, Doronin S, Zuckerman J, Shlapakova IN, Gao J, Pan Z, Herron AJ, Robinson RB, Brink PR, Rosen MR, Cohen IS. Human mesenchymal stem cells as a gene delivery system to create cardiac pacemakers. Circ Res.2004;94:952-959.
31.Tse H-F, Xue T, Lau C-P, Siu C-W, Wang K, Zhang Q-Y, Tomaselli GF, Akar FG, Li RA. Bioartificial sinus node constructed via in vivo gene transfer of an engineered pacemaker HCN channel reduces the dependence on electronic pacemaker in a sick-sinus syndrome model. Circulation. 2006;114:1000-1011.
32.Xiao, J.; Yang, B.; Lin, H.; Lu, Y.; Luo, X. & Wang, Z. Novel approaches for gene-specific interference via manipulating actions of microRNAs: examination on the pacemaker channel genes HCN2 and HCN4. J Cell Physiol,2007; 212:285-292.
33.Lin G, Cai J, Jiang H, Shen H, Jiang X, Yu Q, Song J. Biological pacemaker created by fetal cardiomyocyte transplantation. J Biomed Sci. 2005; 12:513-519.
34.Ruhparwar A, Tebbenjohanns J, Niehaus M, Mengel M, Irtel T, Kofidis T, Pichlmaier AM, Haverich A. Transplanted fetal cardiomyocytes as cardiac pacemaker. Eur J Cardiothorac Surg.2002;21:853-857.
35.Cai J, Lin G, Jiang H, Yang B, Jiang X, Yu Q, Song J. Transplanted neonatal cardiomyocytes as a potential biological pacemaker in pigs with complete atrioventricular block. Transplantation.2006;81:1022-1026.
36.Kehat I, Khimovich L, Caspi O, Gepstein A, Shofti R, Arbel G, Huber I, Satin J, Itskovitz-Eldor J, Gepstein L. Electromechanical integration of cardiomyocytes derived from human embryonic stem cells. Nat Biotechnol. 2004;22:1282-1289.
37.Xue T, Cho HC, Akar FG, Tsang SY, Jones SP, Marban E, Tomaselli GF, Li RA. Functional integration of electrically active cardiac derivatives from genetically engineered human embryonic stem cells with quiescent recipient ventricular cardiomyocytes:insights into the development of cell-based pacemakers. Circulation.2005; 111:11-20.
38.Choi YH, Stamm C, Hammer PE, Kwaku KF, Marler JJ, Friehs I, Jones M, Rader CM, Roy N, Eddy MT, Triedman JK, Walsh EP, McGowan FX Jr, del Nido PJ, Cowan DB. Cardiac conduction through engineered tissue. Am J Pathol. 2006; 169:72-85.
39.Gepstein, L. Stem cells as biological heart pacemakers. Expert Opin Biol Ther,2005;5: 1531-1537.
40.Plotnikov, A. N.; Shlapakova, I.; Szabolcs, M. J.; Danilo, P.; Lorell, B. H.; Potapova, I. A.; Lu, Z.; Rosen, A. B.; Mathias, R. T.; Brink, P. R.; Robinson, R. B.; Cohen, I. S. & Rosen, M. R. Xenografted adult human mesenchymal stem cells provide a platform for sustained biological pacemaker function in canine heart. Circulation, 2007; 116: 706-713.
41.Zhou, Y.; Yang, X.; Li, H.; Han, L. & Jiang, W. Mesenchymal stem cells transfected with HCN2 genes by LentiV can be modified to be cardiac pacemaker cells. Med Hypotheses, 2007; 69:1093-1097.
42.Li, H.; Yang, X.; Zhao, X.; Jiang, B.; Cheng, X.; Chen, T.; Han, L.; Song, J.; Liu, Z. & Jiang, W. [Pacemaker current gene expression of rat mesenchymal stem cells and identification of mesenchymal stem cells expressing human pacemaker current gene (hHCN2)]Zhonghua Xin Xue Guan Bing Za Zhi, 2006; 34: 917-921.
43.Potapova, I.; Plotnikov, A.; Lu, Z.; Danilo, P.; Valiunas, V.; Qu, J.; Doronin, S.; Zuckerman, J.; Shlapakova, I. N.; Gao, J.; Pan, Z.; Herron, A. J.; Robinson, R. B.; Brink, P. R.; Rosen, M. R. & Cohen, I. S. Human mesenchymal stem cells as a gene delivery system to create cardiac pacemakers. Circ Res, 2004; 94: 952-959.
44.Tomita, Y.; Makino, S.; Hakuno, D.; Hattan, N.; Kimura, K.; Miyoshi, S.; Murata, M.; Ieda, M. & Fukuda, K. Application of mesenchymal stem cell-derived cardiomyocytes as bio-pacemakers: current status and problems to be solved. Med Biol Eng Comput, 2007; 45: 209-220.
45.Gepstein, L. Cardiovascular therapeutic aspects of cell therapy and stem cells. Ann N Y Acad Sci, 2006; 1080: 415-425.
2.Jonathan Leor, Stem Cell and Gene-Based Therapy: Frontiers in Regenerative Medicine, Berlin,springer, 2005:3-4.
3.Itskovitz-Eldor, J.; Schuldiner, M.; Karsenti, D.; Eden, A.; Yanuka, O.; Amit, M.; Soreq, H. & Benvenisty, N. Differentiation of human embryonic stem cells into embryoid bodies compromising the three embryonic germ layers. Mol Med, 2000; 6:88-95.
4.Kehat, I.; Kenyagin-Karsenti, D.; Snir, M.; Segev, H.; Amit, M.; Gepstein, A.; Livne, E.; Binah, O.; Itskovitz-Eldor, J. & Gepstein, L. Human embryonic stem cells can differentiate into myocytes with structural and functional properties of cardiomyocytes. J Clin Invest, 2001; 108: 407-414.
5.Xu, C; Police, S.; Rao, N. & Carpenter, M. K. Characterization and enrichment of cardiomyocytes derived from human embryonic stem cells. CircRes,2002;91: 501-508.
6.Xu, C; Lebkowski, J. & Gold, J. D. Growth and differentiation of human embryonic stem cells for cardiac cell replacement therapy. Curr Stem Cell Res Ther, 2006; 1:173-187.
7.He, J.; Ma, Y; Lee, Y; Thomson, J. A. & Kamp, T. J. Human embryonic stem cells develop into multiple types of cardiac myocytes: action potential characterization. Circ Res, 2003; 93: 32-39.
8.Kehat, I.; Khimovich, L.; Caspi, O.; Gepstein, A.; Shofti, R.; Arbel, G; Huber, I.; Satin, J.; Itskovitz-Eldor, J. & Gepstein, L. Electromechanical integration of cardiomyocytes derived from human embryonic stem cells. Nat Biotechnol, 2004; 22: 1282-1289.
9.Xu, W.; Zhang, X.; Qian, H.; Zhu, W.; Sun, X.; Hu, J.; Zhou, H. & Chen, Y. Mesenchymal stem cells from adult human bone marrow differentiate into a cardiomyocyte phenotype in vitro. Exp Biol Med (Maywood), 2004; 229: 623-631.
10. Antonitsis, P.; Ioannidou-Papagiannaki, E.; Kaidoglou, A.; Charokopos, N.; Kalogeridis, A.; Kouzi-Koliakou, K.; Kyriakopoulou, I.; Klonizakis, I. & Papakonstantinou, C. Cardiomyogenic potential of human adult bone marrow mesenchymal stem cells in vitro. Thorac Cardiovasc Surg, 2008;56: 77-82.
11.Antonitsis, P.; Ioannidou-Papagiannaki, E.; Kaidoglou, A. & Papakonstantinou, C. In vitro cardiomyogenic differentiation of adult human bone marrow mesenchymal stem cells. The role of 5-azacytidine. Interact Cardiovasc Thorac Surg, 2007; 6:593-597.
12.Makino, S.; Fukuda, K.; Miyoshi, S.; Konishi, R; Kodama, H.; Pan, J.; Sano, M.; Takahashi, T; Hori, S.; Abe, H.; Hata, J.; Umezawa, A. & Ogawa, S. Cardiomyocytes can be generated from marrow stromal cells in vitro. J Clin Invest, 1999; 103:697-705.
13.Hakuno, D.; Fukuda, K.; Makino, S.; Konishi, R; Tomita, Y.; Manabe, T; Suzuki, Y; Umezawa, A. & Ogawa, S. Bone marrow-derived regenerated cardiomyocytes (CMG Cells) express functional adrenergic and muscarinic receptors. Circulation, 2002; 105: 380-386.
14. Cao, R; Niu, L. L.; Meng, L.; Zhao, L. X.; Zheng, M.; Yue, W.; Bai, C. X.; Jia, G L. & Pei, X. T. [Cardiomyocyte-like differentiation of human bone marrow mesenchymal stem cells after exposure of 5-azacytidine in vitro] Shi Yan Sheng Wu Xue Bao, 2004; 37: 118-124.
15. Liu, Y; Song, J.; Liu, W.; Wan, Y; Chen, X. & Hu, C. Growth and differentiation of rat bone marrow stromal cells: does 5-azacytidine trigger their cardiomyogenic differentiation? Cardiovasc Res, 2003; 58: 460-468.
16. Alviano, R.; Fossati, V; Marchionni, C.; Arpinati, M.; Bonsi, L.; Franchina, M.; Lanzoni, G.; Cantoni, S.; Cavallini, C.; Bianchi, F.; Tazzari, P. L. Pasquinelli, G.; Foroni, L.; Ventura, C.; Grossi, A. & Bagnara, G P. Term Amniotic membrane is a high throughput source for multipotent Mesenchymal Stem Cells with the ability to differentiate into endothelial cells in vitro. BMCDev Biol, 2007; 7:11.
17. Sakuragawa, N.; Thangavel, R.; Mizuguchi, M.; Hirasawa, M. & Kamo, I. Expression of markers for both neuronal and glial cells in human amniotic epithelial cells. Neurosci Lett, 1996; 209: 9-12.
18.Ishii, T.; Ohsugi, K.; Nakamura, S.; Sato, K.; Hashimoto, M.; Mikoshiba, K. & Sakuragawa, N. Gene expression of oligodendrocyte markers in human amniotic epithelial cells using neural cell-type-specific expression system. Neurosci Lett, 1999;268:131-134.
19. Zhang, Y.; Li, C; Jiang, X.; Zhang, S.; Wu, Y.; Liu, B.; Tang, P. & Mao, N. Human placenta-derived mesenchymal progenitor cells support culture expansion of long-term culture-initiating cells from cord blood CD34+ cells. Exp Hematol, 2004; 32: 657-664.
20. Yen, B. L.; Huang, H.; Chien, C; Jui, H.; Ko, B.; Yao, M.; Shun, C.; Yen, M; Lee, M. & Chen, Y. Isolation of multipotent cells from human term placenta. Stem Cells, 2005; 23: 3-9.
21. In't Anker, P. S. I.; Scherjon, S. A.; der Keur, C. K.; de Groot-Swings, G M. J. S.; Claas, F. H. J.; Fibbe, W. E. & Kanhai, H. H. H. Isolation of mesenchymal stem cells of fetal or maternal origin from human placenta. Stem Cells, 2004; 22:1338-1345.
22. Zhao, P.; Ise, H.; Hongo, M; Ota, M.; Konishi, I. & Nikaido, T. Human amniotic mesenchymal cells have some characteristics of cardiomyocytes. Transplantation,2005;79:528-535.
23. Rooney, I. A. & Morgan, B. P. Characterization of the membrane attack complex inhibitory protein CD59 antigen on human amniotic cells and in amniotic fluid. Immunology,1992; 76:541-547.
24. Soncini, M.; Vertua, E.; Gibelli, L.; Zorzi, F.; Denegri, M.; Albertini, A.; Wengler, G S. & Parolini, O. Isolation and characterization of mesenchymal cells from human fetal membranes. J Tissue Eng Regen Med,2007; 1: 296-305.
25. Horwitz, E. M.; Blanc, K. Le; Dominici, M.; Mueller, I.; Slaper-Cortenbach, I.; Marini, F. C.; Deans, R. J.; Krause, D. S.; Keating, A. & the International Society for Cellular Therapy Clarification of the nomenclature for MSC:The International Society for Cellular Therapy position statement. Cytotherapy, 2005; 7:393-395.
26. Cohen, I. S.; Rosen, A. B. & Gaudette, G R. A Caveat Emptor for myocardial regeneration:mechanical without electrical recovery will not suffice. J Mol Cell Cardiol,2007; 42:285-288.
27. Chang, M. G.; Tung, L.; Sekar, R. B.; Chang, C. Y.; Cysyk, J.; Dong, P.; Marban, E. & Abraham, M. R. Proarrhythmic potential of mesenchymal stem cell transplantation revealed in an in vitro coculture model. Circulation,2006; 113:1832-1841.
28. Ben-Dor, I.; Fuchs, S. & Kornowski, R. Potential hazards and technical considerations associated with myocardial cell transplantation protocols for ischemic myocardial syndrome. J Am Coll Cardiol,2006;48:1519-1526
29. Schram, G.; Pourrier, M.; Melnyk, P. & Nattel, S. Differential distribution of cardiac ion channel expression as a basis for regional specialization in electrical function. Circ Res,2002; 90:939-950.
30.李澈.心肌细胞膜上的离子通道.见：李澈主编.窦房结.北京：北京医 科大学出版社,2001.69-102.
31.刘泰槰.离子通道与离子流.见：刘泰槰主编.心肌细胞点生理学—离子通道,离子载体和离子流.北京：人民卫生出版社,2005.30-46.
32.刘泰槰.心肌细胞离子流和离子通道研究简史.见：刘泰槰主编.心肌细胞离子通道和离子通道病.北京：人民卫生出版社,2006.1-5.
33.李泱,程芮细胞电生理学.见：李泱,程芮主编.离子通道学.湖北：湖北科学技术出版社,2007.3-88.
34. Kawano, S.; Otsu, K.; Shoji, S.; Yamagata, K. & Hiraoka, M. Ca(2+) oscillations regulated by Na+-Ca+ exchanger and plasma membrane Ca(2+) pump induce fluctuations of membrane currents and potentials in human mesenchymal stem cells. Cell Calcium,2003;34:145-156.
35. Park, K. S.; Jung, K. H.; Kim, S. H.; Kim, K. S.; Choi, M. R.; Kim, Y. & Chai, Y. G. Functional expression of ion channels in mesenchymal stem cells derived from umbilical cord vein. Stem Cells,2007; 25:2044-2052.
36. J. J. B. Jack, D. Noble, R. W. Tsien. Electric current flow in excitable cells. Oxford Science Publications; 1983:229.
37. Fauza, D. Amniotic fluid and placental stem cells. Best Pract Res Clin Obstet Gynaecol,2004; 18:877-891.
38. Marcus, A. J.; Coyne, T. M.; Rauch, J.; Woodbury, D. & Black, I. B. Isolation, characterization, and differentiation of stem cells derived from the rat amniotic membrane. Differentiation,2008; 76:130-144.
39. Miki, T.; Lehmann, T.; Cai, H.; Stolz, D. B. & Strom, S. C. Stem cell characteristics of amniotic epithelial cells. Stem Cells,2005; 23:1549-1559.
40.林雪.羊膜间质干细胞的基本性质和向心肌细胞诱导分化的立体培养系统研究.北京：中国协和医科大学,2007.
41. Pasquinelli, G.; Tazzari, P.; Ricci, F.; Vaselli, C.; Buzzi, M.; Conte, R.; Orrico, C.; Foroni, L.; Stella, A.; Alviano, F.; Bagnara, G. P. Ultrastructural characteristics of human mesenchymal stromal (stem) cells derived from bone marrow and term placenta. Ultrastruct Pathol,2007; 31:23-31.
42. Kim, J.; Kang, H. M.; Kim, H.; Kim, M. R.; Kwon, H. C.; Gye, M. C.; Kang, S. G.; Yang, H. S. & You, J. Ex vivo characteristics of human amniotic membrane-derived stem cells. Cloning Stem Cells,2007; 9:581-594.
43. Kunisaki, S. M.; Armant, M.; Kao, G. S.; Stevenson, K.; Kim, H. & Fauza, D. O. Tissue engineering from human mesenchymal amniocytes:a prelude to clinical trials. J Pediatr Surg,2007; 42:974-979.
44. Heubach, J. F.; Graf, E. M.; Leutheuser, J.; Bock, M.; Balana, B.; Zahanich, I.; Christ, T.; Boxberger, S.; Wettwer, E. & Ravens, U. Electrophysiological properties of human mesenchymal stem cells. J Physiol,2004; 554:659-672.
45. Li, G.; Sun, H.; Deng, X. & Lau, C. Characterization of ionic currents in human mesenchymal stem cells from bone marrow. Stem Cells,2005; 23: 371-382.
46. Bai, X.; Ma, J.; Pan, Z.; Song, Y; Freyberg, S.; Yan, Y.; Vykoukal, D. & Alt, E. Electrophysiological properties of human adipose tissue-derived stem cells. Am J Physiol Cell Physiol,2007;293:C1539-C1550.
47. Brahmajothi MV, Campbell DL, Rasmusson RL, et al. Distinct transient outward potassium current (Ito) phenotypes and distribution of fast-inactivating potassium channel alpha subunits in ferret left ventricular myocytes. J Gen Physiol.1999; 113:581-600.
48. Campbell, D.L., R.L. Rasmusson, Y Qu, and H.C. Strauss. The calcium-independent transient outward potassium current in isolated ferret right ventricular myocytes. I. Basic characterization and kinetic analysis. J. Gen. Physiol.1993; 101:571-601.
49. Campbell, D.L., Y Qu, R.L. Rasmusson, and H.C. Strauss. The calcium-independent transient outward potassium current in isolated ferret right ventricular myocytes. Ⅱ. Closed state reverse use-dependent block by 4-aminopyridine. J. Gen. Physiol.1993; 101:603-626.
50. Gryschenko, O.; Lu, Z. J.; Fleischmann, B. K. & Hescheler, J. Outwards currents in embryonic stem cell-derived cardiomyocytes. Pflugers Arch, 2000;439:798-807.
51. Wei, A. D.; Gutman, G. A.; Aldrich, R.; Chandy, K. G.; Grissmer, S. & Wulff, H. International Union of Pharmacology. LⅡ. Nomenclature and molecular relationships of calcium-activated potassium channels. Pharmacol Rev, 2005;57:463-472.
52. Cui, D.H. Cox, and R.W. Aldrich. Intrinsic Voltage Dependence and Ca2+ Regulation of mslo Large Conductance Ca-activated K+ Channels J. Gen. Physiol.1997; 109:647-673.
53. Hamada E, Nakajima T, Ota S, Terano A, Omata M, Nakade S, Mikoshiba K, Kurachi Y. Activation of Ca(2+)-dependent K+ current by acetylcholine and histamine in a human gastric epithelial cell line. J Gen Physiol.1993; 102:667-692.
54. Weerapura, M.; Nattel, S.; Chartier, D.; Caballero, R. & Hebert, T. E. A comparison of currents carried by HERG, with and without coexpression of MiRP1, and the native rapid delayed rectifier current. Is MiRP10 the missing link? J Physiol,2002; 540:15-27.
55. Federico Sesti and Steve A.N. Goldstein Single-Channel Characteristics of Wild-Type IKs Channels and Channels formed with Two MinK Mutants that Cause Long QT Syndrome J. Gen. Physiol.1998;112:651-663.
56. Sesti, F. & Goldstein, S. A. Single-channel characteristics of wild-type IKs channels and channels formed with two minK mutants that cause long QT syndrome. J Gen Physiol,1998; 112:651-663.
57. Grissmer S, Nguyen AN, Aiyar J, Hanson DC, Mather RJ, Gutman GA, Karmilowicz MJ, Auperin DD, Chandy KG. Pharmacological characterization of five cloned voltage-gated K+channels, types Kvl.1,1.2, 1.3,1.5, and 3.1, stably expressed in mammalian cell lines. Mol Pharmacol. 1994; 45:1227-1234.
58. Kawano, S.; Shoji, S.; Ichinose, S.; Yamagata, K.; Tagami, M. & Hiraoka, M. Characterization of Ca2+ signaling pathways in human mesenchymal stem cells. Cell Calcium,2002; 32:165-174.
59. Deng, X.; Sun, H.; Lau, C. & Li, G. Properties of ion channels in rabbit mesenchymal stem cells from bone marrow. Biochem Biophys Res Commun, 2006;348:301-309
60. Li, G; Deng, X.; Sun, H.; Chung, S. S. M.; Tse, H. & Lau, C. Ion channels in mesenchymal stem cells from rat bone marrow. Stem Cells, 2006;24:1519-1528.
61. Tao, R.; Lau, C.; Tse, H. & Li, G Functional ion channels in mouse bone marrow mesenchymal stem cells. Am J Physiol Cell Physiol,2007;293: C1561-C1567.
62. Wang, K.; Xue, T.; Tsang, S.; Huizen, R. V.; Wong, C. W.; Lai, K. W.; Ye, Z.; Cheng, L.; Au, K. W.; Zhang, J.; Li, G.; Lau, C.; Tse, H. & Li, R. A. Electrophysiological properties of pluripotent human and mouse embryonic stem cells. Stem Cells,2005;23:1526-1534.
63. Wonderlin, W. F. & Strobl, J. S. Potassium channels, proliferation and G1 progression. J Membr Biol,1996; 154:91-107.
64. Czarnecki, A.; Vaur, S.; Dufy-Barbe, L.; Dufy, B. & Bresson-Bepoldin, L. Cell cycle-related changes in transient K(+) current density in the GH3 pituitary cell line. Am J Physiol Cell Physiol,2000; 279:C1819-C1828.
65. Czarnecki, A.; Dufy-Barbe, L.; Huet, S.; Odessa, M. & Bresson-Bepoldin, L. Potassium channel expression level is dependent on the proliferation state in the GH3 pituitary cell line. Am J Physiol Cell Physiol,2003; 284: C1054-C1064.
66. Boullin, J. & Morgan, J. M. The development of cardiac rhythm. Heart,2005; 91:874-875.
67. Moorman AF, Christoffels VM. Cardiac Chamber Formation:Development, Genes, and Evolution. Physiol Rev.2003;83:1223-1267.
68. Moorman, A. F.; de Jong, F.; Denyn, M. M. & Lamers, W. H. Development of the cardiac conduction system. Circ Res,1998;82:629-644.
69. Pennisi, D. J.; Rentschler, S.; Gourdie, R. G.; Fishman, G I. & Mikawa, T. Induction and patterning of the cardiac conduction system. Int J Dev Biol, 2002; 46:765-775.
70. Cheng, G.; Litchenberg, W. H.; Cole, G J.; Mikawa, T.; Thompson, R. P. & Gourdie, R. G Development of the cardiac conduction system involves recruitment within a multipotent cardiomyogenic lineage. Development, 1999,126,5041-5049.
71. Tomita, Y.; Makino, S.; Hakuno, D.; Hattan, N.; Kimura, K.; Miyoshi, S.; Murata, M.; Ieda, M. & Fukuda, K. Application of mesenchymal stem cell-derived cardiomyocytes as bio-pacemakers:current status and problems to be solved. Med Biol Eng Comput,2007;45:209-220.
72. Donmez, A.; Zoghi, M.; Cagirgan, S.; Acarlar, C. & Tombuloglu, M. The effect of hematopoietic progenitor cells'temperature on cardiac arrhythmias in patients given peripheral blood progenitor cells. Transfus Apher Sci,2006, 34,245-251.
73. J. J. B. Jack, D. Noble, R. W. Tsien. Electric current flow in excitable cells. Oxford Science Publications; 1983:225-227.
74.袁岩,陈连凤,张抒扬,吴炜,陈浩,严晓伟心肌微环境对骨髓间充质干细胞的诱导分化作用.中国介入心脏病学杂志2005；13：315-318.
75.袁岩,陈连凤,张抒扬,吴炜,陈浩,严晓伟心肌裂解液对对骨髓间充质干细胞向心肌细胞分化诱导作用的研究.中国心血管病杂志2005；33：170-173.
76. Hirota, A.; Fujii, S. & Kamino, K. Optical monitoring of spontaneous electrical activity of 8-somite embryonic chick heart. Jpn J Physiol,1979;29: 635-639.
77. Kamino, K.; Hirota, A. & Fujii, S. Localization of pacemaking activity in early embryonic heart monitored using voltage-sensitive dye. Nature,1981; 290:595-597.
78. Viragh, S. & Challice, C. E. The development of the early atrioventricular conduction system in the embryonic heart. Can J Physiol Pharmacol, 1983;61:775-792.
79. Viragh, S. & Challice, C. E. The development of the conduction system in the mouse embryo heart. Dev Biol,1982;89:25-40.
80.李澈.心肌细胞膜上的离子通道,钾通道和钾电流.见：李澈主编.窦房结.北京：北京医科大学出版社,2001.79-88.
81.刘泰槰.瞬时外向电流Ito.见：刘泰槰主编.心肌细胞点生理学—离子通道,离子载体和离子流.北京：人民卫生出版社,2005.101-103.
82.李泱,程芮瞬时外向钾电流.见：李泱,程芮主编.离子通道学.湖北：湖北科学技术出版社,2007.49-54.
83. Crumb, W. J.; Pigott, J. D. & Clarkson, C. W. Comparison of Ito in young and adult human atrial myocytes:evidence for developmental changes. Am J Physiol,1995; 268:H1335-H134.
84. Sartiani, L.; Bettiol, E.; Stillitano, F.; Mugelli, A.; Cerbai, E. & Jaconi, M. E. Developmental changes in cardiomyocytes differentiated from human embryonic stem cells:a molecular and electrophysiological approach. Stem Cells,2007; 25:1136-1144.
85. Mills, W. R.; Mai, N.; Kiedrowski, M. J.; Unger, R.; Forudi, F.; Popovic, Z. B.; Penn, M. S. & Laurita, K. R. Stem cell therapy enhances electrical viability in myocardial infarction. J Mol Cell Cardiol,2007; 42:304-314.
86. Tang, Y. L.; Zhao, Q.; Qin, X.; Shen, L.; Cheng, L.; Ge, J. & Phillips, M. I. Paracrine action enhances the effects of autologous mesenchymal stem cell transplantation on vascular regeneration in rat model of myocardial infarction. Ann Thorac Surg,2005;80:229-236; discussion 236-237.
87. Cowan, D. B. & McGowan, F. X. A paradigm shift in cardiac pacing therapy? Circulation,2006; 114:986-988.
88. Rosen, M. R.; Brink, P. R.; Cohen, I. S. & Robinson, R. B. Genes, stem cells and biological pacemakers. Cardiovasc Res,2004; 64:12-23.
89. Boink, G. J. J.; Seppen, J.; de Bakker, J. M. T. & Tan, H. L. Gene therapy to create biological pacemakers. Med Biol Eng Comput,2007;45:167-176.
90. Anghel, T. M. & Pogwizd, S. M. Creating a cardiac pacemaker by gene therapy. Med Biol Eng Comput,2007;45:145-155.
91. Edelberg, J. M.; Aird, W. C. & Rosenberg, R. D. Enhancement of murine cardiac chronotropy by the molecular transfer of the human beta2 adrenergic receptor cDNA. J Clin Invest,1998; 101:337-343.
92. Miake, J.; Marban, E. & Nuss, H. B. Biological pacemaker created by gene transfer. Nature,2002;419:132-133.
93. Tan, H. L.; Bardai, A.; Shimizu, W.; Moss, A. J.; Schulze-Bahr, E.; Noda, T. & Wilde, A. A. M. Genotype-specific onset of arrhythmias in congenital long-QT syndrome:possible therapy implications. Circulation,2006; 114: 2096-2103.
94. Rosen, M. R.; Brink, P. R.; Cohen, I. S. & Robinson, R. B. Biological pacemakers based on If. Med Biol Eng Comput,2007;45:157-166.
95. Qu, J.; Barbuti, A.; Protas, L.; Santoro, B.; Cohen, I. S. & Robinson, R. B. HCN2 overexpression in newborn and adult ventricular myocytes:distinct effects on gating and excitability. Circ Res,2001;89:E8-14.
96. Qu, J.; Plotnikov, A. N.; Danilo, P.; Shlapakova, I.; Cohen, I. S.; Robinson, R. B. & Rosen, M. R. Expression and function of a biological pacemaker in canine heart. Circulation,2003,107,1106-1109.
97. Potapova I, Plotnikov A, Lu Z, Danilo P Jr, Valiunas V, Qu J, Doronin S, Zuckerman J, Shlapakova IN, Gao J, Pan Z, Herron AJ, Robinson RB, Brink PR, Rosen MR, Cohen IS. Human mesenchymal stem cells as a gene delivery system to create cardiac pacemakers. Circ Res.2004; 94:952-959.
98. Tse H-F, Xue T, Lau C-P, Siu C-W, Wang K, Zhang Q-Y, Tomaselli GF, Akar FG, Li RA. Bioartificial sinus node constructed via in vivo gene transfer of an engineered pacemaker HCN channel reduces the dependence on electronic pacemaker in a sick-sinus syndrome model. Circulation.2006; 114:1000-1011.
99. Gepstein, L. Stem cells as biological heart pacemakers. Expert Opin Biol Ther,2005;5:1531-153.
100. Lin G, Cai J, Jiang H, Shen H, Jiang X, Yu Q, Song J. Biological pacemaker created by fetal cardiomyocyte transplantation. J Biomed Sci.2005; 12:513-519.
101. Ruhparwar A, Tebbenjohanns J, Niehaus M, Mengel M, Irtel T, Kofidis T, Pichlmaier AM, Haverich A. Transplanted fetal cardiomyocytes as cardiac pacemaker. Eur J Cardiothorac Surg.2002; 21:853-857.
102. Cai J, Lin G, Jiang H, Yang B, Jiang X, Yu Q, Song J. Transplanted neonatal cardiomyocytes as a potential biological pacemaker in pigs with complete atrioventricular block. Transplantation.2006; 81:1022-1026.
103. Kehat I, Khimovich L, Caspi O, Gepstein A, Shofti R, Arbel G, Huber I, Satin J, Itskovitz-Eldor J, Gepstein L. Electromechanical integration of cardiomyocytes derived from human embryonic stem cells. Nat Biotechnol. 2004; 22:1282-1289.
104. Xue T, Cho HC, Akar FG, Tsang SY, Jones SP, Marban E, Tomaselli GF, Li RA. Functional integration of electrically active cardiac derivatives from genetically engineered human embryonic stem cells with quiescent recipient ventricular cardiomyocytes:insights into the development of cell-based pacemakers. Circulation.2005; 111:11-20.
105. Choi YH, Stamm C, Hammer PE, Kwaku KF, Marler JJ, Friehs I, Jones M, Rader CM, Roy N, Eddy MT, Triedman JK, Walsh EP, McGowan FX Jr, del Nido PJ, Cowan DB. Cardiac conduction through engineered tissue. Am J Pathol.2006; 1691:72-85.
106. Zhou, Y.; Yang, X.; Li, H.; Han, L. & Jiang, W. Mesenchymal stem cells transfected with HCN2 genes by LentiV can be modified to be cardiac pacemaker cells. Med Hypotheses,2007; 69:1093-1097.
107. Zhou, Y.; Yang, X. & Li, H. Hyperpolarization-activated cyclic nucleotide-gated channel gene:the most possible therapeutic applications in the field of cardiac biological pacemakers. Med Hypotheses,2007; 69: 541-544.
108. DiFrancesco, D.; Ferroni, A.; Mazzanti, M. & Tromba, C. Properties of the hyperpolarizing-activated current (If) in cells isolated from the rabbit sino-atrial node. J Physiol,1986; 377:61-88.
109. DiFrancesco, D. The contribution of the 'pacemaker' current (If) to generation of spontaneous activity in rabbit sino-atrial node myocytes. J Physiol,1991; 434:23-40.
1. Rosen, M. R.; Brink, P. R.; Cohen, I. S. & Robinson, R. B. Genes, stem cells and biological pacemakers. Cardiovasc Res,2004; 64:12-23.
2. Rosen, M. R.15th annual Gordon K. Moe Lecture. Biological pacemaking: in our lifetime? Heart Rhythm,2005; 2:418-428.
3. Gepatein L, Feld Y, Yankelson L. Somatic gene and cell therapy strategies for the treament of cardiac. Am J Physiol Heart Cir Physiol 2004;286:H815-H822.
4.李澈.窦房结的组织学和发生.见李澈主编.窦房结.北京：北京医科大学出版社,2001.9-24.
5.刘泰槰.心肌细胞的动作电位.见刘泰槰主编.心肌细胞点生理学—离子通道,离子载体和离子流.北京：人民卫生出版社,2005.17-27.
6. Brown, H. & Difrancesco, D. Voltage-clamp investigations of membrane currents underlying pace-maker activity in rabbit sino-atrial node. J Physiol, 1980; 308:331-351.
7. DiFrancesco, D. & Ojeda, C. Properties of the current If in the sino-atrial node of the rabbit compared with those of the current iK, in Purkinje fibres. J Physiol,1980;308:353-367.
8. DiFrancesco, D. Properties of the cardiac pacemaker (If current. Boll Soc Ital Biol Sper,1984; 60 Suppl 4:29-33.
9. Di Francesco D. The pacemaker current (If) Plays an important role in regulating SA node pacemaker activity.Cardiovasc Res,1995;30:307-308.
10. DiFrancesco, D. Serious workings of the funny current. Prog Biophys Mol Biol,2006; 90:13-25.
11. Santoro, B.; Grant, S. G.; Bartsch, D. & Kandel, E. R. Interactive cloning with the SH3 domain of N-src identifies a new brain specific ion channel protein, with homology to eag and cyclic nucleotide-gated channels. Proc Natl Acad Sci U S A,1997; 94:14815-1482.
12. Boullin, J. & Morgan, J. M. The development of cardiac rhythm. Heart, 2005;91:874-875.
13. Moorman AF, Christoffels VM. Cardiac Chamber Formation: Development, Genes, and Evolution. Physiol Rev.2003;83:1223-1267
14. Gorza L, Schiaffino S, Vitadello M. Heart conduction system:a neural crest derivative? Brain Res 1988;457:360-366.
15. Cheng, G.; Litchenberg, W. H.; Cole, G. J.; Mikawa, T.; Thompson, R. P. & Gourdie, R. G. Development of the cardiac conduction system involves recruitment within a multipotent cardiomyogenic lineage. Development, 1999; 126:5041-5049.
16. Gourdie RG, Mima T, Thompson RP, Mikawa T. Terminal diversification of the myocyte lineage generates Purkinje fibers of the cardiac conduction system. Development.1995; 121:1423-1431
17. Hirota, A.; Fujii, S. & Kamino, K. Optical monitoring of spontaneous electrical activity of 8-somite embryonic chick heart. Jpn J Physiol,1979; 29: 635-639.
18. Kamino, K.; Hirota, A. & Fujii, S. Localization of pacemaking activity in early embryonic heart monitored using voltage-sensitive dye. Nature,1981; 290:595-597.
19. Viragh, S. & Challice, C. E. The development of the conduction system in the mouse embryo heart. Dev Biol,1980; 80:28-45.
20. Pennisi, D. J.; Rentschler, S.; Gourdie, R. G.; Fishman, G. I. & Mikawa, T. Induction and patterning of the cardiac conduction system. Int J Dev Biol, 2002; 46:765-775.
21. Gourdie, R. G.; Wei, Y.; Kim, D.; Klatt, S. C. & Mikawa, T. Endothelin-induced conversion of embryonic heart muscle cells into impulse-conducting Purkinje fibers. Proc Natl Acad Sci U S A,1998; 95: 6815-6818.
22. Rentschler, S.; Vaidya, D. M.; Tamaddon, H.; Degenhardt, K.; Sassoon, D.; Morley, G. E.; Jalife, J. & Fishman, G. I. Visualization and functional characterization of the developing murine cardiac conduction system. Development,2001; 128:1785-1792.
23. Cowan, D. B. & McGowan, F. X. A paradigm shift in cardiac pacing therapy? Circulation,2006; 114:986-988.
24. Boink, G. J. J.; Seppen, J.; de Bakker, J. M. T. & Tan, H. L. Gene therapy to create biological pacemakers. Med Biol Eng Comput,2007; 45:167-176.
25. Anghel, T. M. & Pogwizd, S. M. Creating a cardiac pacemaker by gene therapy. Med Biol Eng Comput,2007;45:145-155.
26. Edelberg, J. M.; Aird, W. C. & Rosenberg, R. D. Enhancement of murine cardiac chronotropy by the molecular transfer of the human beta2 adrenergic receptor cDNA. J Clin Invest,1998; 101:337-343.
27. Miake, J.; Marban, E. & Nuss, H. B. Biological pacemaker created by gene transfer. Nature,2002; 419:132-133.
28. Tan, H. L.; Bardai, A.; Shimizu, W.; Moss, A. J.; Schulze-Bahr, E.; Noda, T. & Wilde, A. A. M. Genotype-specific onset of arrhythmias in congenital long-QT syndrome:possible therapy implications. Circulation,2006; 114: 2096-2103
29. Qu, J.; Plotnikov, A. N.; Danilo, P.; Shlapakova, I.; Cohen, I. S.; Robinson, R. B. & Rosen, M. R. Expression and function of a biological pacemaker in canine heart. Circulation,2003; 107:1106-1109.
30. Potapova I, Plotnikov A, Lu Z, Danilo P Jr, Valiunas V, Qu J, Doronin S, Zuckerman J, Shlapakova IN, Gao J, Pan Z, Herron AJ, Robinson RB, Brink PR, Rosen MR, Cohen IS. Human mesenchymal stem cells as a gene delivery system to create cardiac pacemakers. Circ Res.2004;94:952-959.
31.Tse H-F, Xue T, Lau C-P, Siu C-W, Wang K, Zhang Q-Y, Tomaselli GF, Akar FG, Li RA. Bioartificial sinus node constructed via in vivo gene transfer of an engineered pacemaker HCN channel reduces the dependence on electronic pacemaker in a sick-sinus syndrome model. Circulation. 2006;114:1000-1011.
32.Xiao, J.; Yang, B.; Lin, H.; Lu, Y.; Luo, X. & Wang, Z. Novel approaches for gene-specific interference via manipulating actions of microRNAs: examination on the pacemaker channel genes HCN2 and HCN4. J Cell Physiol,2007; 212:285-292.
33.Lin G, Cai J, Jiang H, Shen H, Jiang X, Yu Q, Song J. Biological pacemaker created by fetal cardiomyocyte transplantation. J Biomed Sci. 2005; 12:513-519.
34.Ruhparwar A, Tebbenjohanns J, Niehaus M, Mengel M, Irtel T, Kofidis T, Pichlmaier AM, Haverich A. Transplanted fetal cardiomyocytes as cardiac pacemaker. Eur J Cardiothorac Surg.2002;21:853-857.
35.Cai J, Lin G, Jiang H, Yang B, Jiang X, Yu Q, Song J. Transplanted neonatal cardiomyocytes as a potential biological pacemaker in pigs with complete atrioventricular block. Transplantation.2006;81:1022-1026.
36.Kehat I, Khimovich L, Caspi O, Gepstein A, Shofti R, Arbel G, Huber I, Satin J, Itskovitz-Eldor J, Gepstein L. Electromechanical integration of cardiomyocytes derived from human embryonic stem cells. Nat Biotechnol. 2004;22:1282-1289.
37.Xue T, Cho HC, Akar FG, Tsang SY, Jones SP, Marban E, Tomaselli GF, Li RA. Functional integration of electrically active cardiac derivatives from genetically engineered human embryonic stem cells with quiescent recipient ventricular cardiomyocytes:insights into the development of cell-based pacemakers. Circulation.2005; 111:11-20.
38.Choi YH, Stamm C, Hammer PE, Kwaku KF, Marler JJ, Friehs I, Jones M, Rader CM, Roy N, Eddy MT, Triedman JK, Walsh EP, McGowan FX Jr, del Nido PJ, Cowan DB. Cardiac conduction through engineered tissue. Am J Pathol. 2006; 169:72-85.
39.Gepstein, L. Stem cells as biological heart pacemakers. Expert Opin Biol Ther,2005;5: 1531-1537.
40.Plotnikov, A. N.; Shlapakova, I.; Szabolcs, M. J.; Danilo, P.; Lorell, B. H.; Potapova, I. A.; Lu, Z.; Rosen, A. B.; Mathias, R. T.; Brink, P. R.; Robinson, R. B.; Cohen, I. S. & Rosen, M. R. Xenografted adult human mesenchymal stem cells provide a platform for sustained biological pacemaker function in canine heart. Circulation, 2007; 116: 706-713.
41.Zhou, Y.; Yang, X.; Li, H.; Han, L. & Jiang, W. Mesenchymal stem cells transfected with HCN2 genes by LentiV can be modified to be cardiac pacemaker cells. Med Hypotheses, 2007; 69:1093-1097.
42.Li, H.; Yang, X.; Zhao, X.; Jiang, B.; Cheng, X.; Chen, T.; Han, L.; Song, J.; Liu, Z. & Jiang, W. [Pacemaker current gene expression of rat mesenchymal stem cells and identification of mesenchymal stem cells expressing human pacemaker current gene (hHCN2)]Zhonghua Xin Xue Guan Bing Za Zhi, 2006; 34: 917-921.
43.Potapova, I.; Plotnikov, A.; Lu, Z.; Danilo, P.; Valiunas, V.; Qu, J.; Doronin, S.; Zuckerman, J.; Shlapakova, I. N.; Gao, J.; Pan, Z.; Herron, A. J.; Robinson, R. B.; Brink, P. R.; Rosen, M. R. & Cohen, I. S. Human mesenchymal stem cells as a gene delivery system to create cardiac pacemakers. Circ Res, 2004; 94: 952-959.
44.Tomita, Y.; Makino, S.; Hakuno, D.; Hattan, N.; Kimura, K.; Miyoshi, S.; Murata, M.; Ieda, M. & Fukuda, K. Application of mesenchymal stem cell-derived cardiomyocytes as bio-pacemakers: current status and problems to be solved. Med Biol Eng Comput, 2007; 45: 209-220.
45.Gepstein, L. Cardiovascular therapeutic aspects of cell therapy and stem cells. Ann N Y Acad Sci, 2006; 1080: 415-425.