| BackgroundsIt has been widely accepted that cardiac myocyte hypertrophy is the most important compensation mechanism of the heart to satisfy increased physiological demands and or to adapt to pathological cardiac conditions. On the other hand, there has been a strong belief that the muscle cells of the adult heart cannot divide or replicate under the same conditions~[1].However, recent reports have highlighted evidence on the existence of cardiac cell renewal in either the normal or diseased heart but the clinical significance of this cell renewal in several pathological conditions in humans remains uncertain~[2-4].In the setting of myocardial infarction, for instance, the compensatory replication of myocytes does not repair the infarct area itself, but occurs at the border and at a distance of the infarcted myocardium, leaving behind a scar lesion whose size is a major determinant of morbidity and mortality~[5]. Although we might be able to enhance this proliferative response in the future using pharmacological modulation or genetic manipulations of the cardiac cell cycle, the cardiac regeneration process is likely to be more complex than replicative enhancement as the sole means of obtaining a scarless heart~[6]. In addition, cell cycle manipulations are likely to induce apoptosis, tumor genesis and even cardiac myopathies~[7,8].Stem cells could provide an alternative biological strategy to repair myocardial infarctions and prevent postinfarct congestive heart failure, because there is a growing body of evidence that bone marrow stem cells can generate new cardiomyocytes in animals and humans[9-12]. However, before we can claim that this regenerative strategy in cardiovascularmedicine is truly applicable in humans, many hurdles must be overcome first.Transplantation of both skeletal myoblasts and stem cells into the region of infarcted myocardium results in improved myocardial function in both the murine and porcine infarct models. In order for cell therapy to be widely clinically applicable, the optimal cell has to be compatible both mechanically and electrically with the host myocardium. For this reasons ion channels can be important functional markers of cardiac differentiation. Up to now, enhanced incidence of arrhythmias in patients who received cardiac implantationof bone marrow stem cells has not been noted, although such risk cannot be excluded.The study reported by Pak et al~[13] was designed to explore the hypothesis that another possible mechanism of "proarrhythmia" might be the result of stimulation of sympathetic nerve sprouting and overexpression of neurotransmitters as a consequence of mesenchymal stem cell transplantation to the heart. The notion that sympathetic stimulation and hyperinnervation can be arrhythmogenic in a heart with a prior myocardial infarction has been studied previously~[14,15] and provides a rationale for this study. The investigators injected cultures of BrdU-labeled mesenchymal stem cells, together with unfractonated bone marrow cells, into the peri-infarct zone of swine 1 month after induction of anterior myocardial infarction. They then performed histologic analysis of peri-infarct and remote myocardium after an additional month. Using growth-associated protein 43(GAP43) as a marker for nerve generation, they found a strong increase in nerve sprouting in the hearts that received mesenchymal stem cell implants compared with hearts eceiving bone marrow cells alone or cell culturemedia. Nerve sproutingwas prominent in the remote myocardium, including the atria. No transplanted cells could be identified outside of the original injection sites, suggesting that the nerve sprouting was largely a paracrine or inductive effect of the donor cells. Unfortunately, they were not able to document that these were actual functioning sympathetic nerves, as the increase in tyrosine hydroxylase in the same sections was modest, possibly reflection immaturity. Similarly, the present study does not provide evidence that the nerve sprouts are destined to become hyperfunctional, even after maturing.Currently, the realistic prospects for bone marrow stem cell therapy on a large scale for cardiac repair in humans appears to be remote and one of the reasons could be the electorphysiological properties of the engrafted stem cells. When cultured in vitro, cardiomyocytes derived form mouse embryonic stem cells reveal arrhythmogenic properties withy action potential heterogeneity, protracted automaticity, reentry and frequent spontaneous and easily inducible triggered activity~[16]. In addition, the ischemic milieu that surrounds the engrafted cells could exacerbate arrrhthymogenesis. This developmental heterogeneity has also been demonstrated in mesenchymal stem cell cultures with cardiomyogenic commitment~[17]. It seems very likely that during the transdifFerentiation process, the engrafted stem cells in the infarct area not only recapitulated the morphological characteristics but also the different elctrophysiological stages of cardiac embryogenesis, creating a substrate for arrhythmogenesis in the regeneration heart. ObjectiveAt present there is little information about the electrophysiological behaviors of the bone marrow mesenchymal stem cells. This study is toexplore the biological properties as well as the intracellular calcium concentration of natural or undifferentiated bone marrow mesenchymal stem cells. In addition the change of sinus node function and arrhythmic potential caused by transplanted BMSC were observed. Methods1, Isolate BMSC by Ficoll2 , Induce BMSC by 5-azacytidine, use MTT to measure the cytotoxitycaused by the drug, chose the proper concentration3 Observe the microstructure of BMSC by electron microscopy4 , Measure cTnl by ELISA and Western5 , RT-PCR for connexin 43 expression6 , Use Fura 2/AM to measure intracellular calcium concentration7 , RT-PCR for ion channel gene expression8 , Establishment myocardial infarction model by intracoronary balloonocclusion9 , EPS program stimulate to establish sustained ventricular tachycardiaanimal model 10, Transplant the BMSC into infarcted myocardium and discuss thearrhythmic potential caused by these cells Results1 , The bone marrow mesenchymal stem cells can easily and quickly beobtained by Fi-coll density grads centrifugal method2 , The reasonable cell implant density is 5-10 x 105/cm2, the properfirst medium change time is 48 hours, and the best PH for cell culture is 7.2.3 , Bone marrow mesenchymal stem cells posses the properultrastructure basis to act as seed cell in tissue engineering.4, Induced by appropriate concentration of 5-aza, BMSC can differentiate into cardiomyocytes-like cells and express cardiac special protein troponin I express cardiac special gap junction connexin 43.5, The intracellular calcium concentration of differential group is higher than control group and the mechanism of calcium release is different from that of control group6, Differentiated BMSC can express of SCN5A, KCNE1, which undifferentiated BMSC can't express, and after induced by 5-aza the expression of L-type calcium channel and Kir 2.1 is increased whereas the expression for Kvl.2 is unchanged.7, The technique of LAD occlusion presents a less invasive alternative to open chest models. Proper prevention and management of the complications result from the operation could reduce the mortality rate.8, The mini intracoronary balloon occlusion can not only lead myocardial infarction but also induce monomorphic ventricular tarchycardia.9, Sustained ventricular tachycardia can't be induced in cell treatment group, otherwise it can be induced in control group.Conclusion1, BMSC can be quickly and abundant obtained at proper culture condition.2, After induced by 5-azacytidine, BMSC can differentiated into myocardialcyte-like cells.3, The arrhythmic possibility of BMSC is low, and maybe BMSC posses the possibility which can treat arrhythmia caused bymyocardial infarction.
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