Effects of bone marrow mesenchymal stem cell transplantation on voltage-gated   K+ channel proteins and cytokines in the infarcted myocardium of rats

doi:10.3969/j.issn.2095-4344.0469

Abstract

Abstract:

BACKGROUND: Studies have shown that bone marrow mesenchymal stem cell (BMSC) transplantation can effectively improve cardiac function after myocardial infarction. However, few reports have been issued on myocardial electrophysiology after BMSCs transplantation.
OBJECTIVE: To observe the effects of BMSCs transplantation on voltage-gated K⁺ channel protein and myocardial infarction-related cytokines, thereby providing basic evidence for further exploration on the mechanism underlying arrhythmia in myocardial infarction due to BMSCs transplantation.
METHODS: Forty male Wistar rats, SPF grade, were randomly divided into four groups: sham group, model group, cell culture medium group and BMSCs group. The myocardial infarction model was created in rats by permanent ligation of the left descending coronary artery. At 15-20 minutes after surgery, BMSCs (100 µL, 1×10⁶) or cell culture medium (100 µL) was injected at four sites in the peri-infarct zone. Four weeks after cell therapy, cardiac samples were taken, the pathological morphology of the infarcted myocardium was observed by hematoxylin-eosin staining, and the infarct size was calculated; the expression levels of voltage-gated K⁺ channel proteins Kv1.2 and Kv1.5 and cardiac troponin T (cTnT) were measured by western blot assay; and the expression levels of apoptotic factor (Caspase-3), autophagy factor (Bcl-2), nitric oxide and superoxide dismutase were tested by immunohistochemistry.
RESULTS AND CONCLUSION: Compared with the model group and cell culture medium group, the infarct size decreased in the BMSCs group (P < 0.05); the expression levels of cTnT, Kv1.5 and superoxide dismutase increased (P < 0.05), and the expression levels of Caspase-3, Bcl-2 and Kv1.2 decreased (P < 0.05) in the BMSCs group. In summary, BMSCs transplantation can promote the expression of voltage-gated K⁺ channel proteins, and improve anti-oxidation capacity of the myocardium and decrease apoptosis and autophagy.

中国组织工程研究杂志出版内容重点：干细胞；骨髓干细胞；造血干细胞；脂肪干细胞；肿瘤干细胞；胚胎干细胞；脐带脐血干细胞；干细胞诱导；干细胞分化；组织工程

Key words: Bone Marrow, Mesenchymal Stem Cell Transplantation, Myocardial Infarction, Potassium Channels, Voltage-Gated, Apoptosis, Tissue Engineering

CLC Number:

R394.2

Hu Ji-hong, Zhao Jing-miao, Wang Qiu-ping, Jia Jia, Lu Juan, Jin Hua, Hou Qian. Effects of bone marrow mesenchymal stem cell transplantation on voltage-gated K⁺ channel proteins and cytokines in the infarcted myocardium of rats[J]. Chinese Journal of Tissue Engineering Research, 2018, 22(9): 1389-1394.

Figures/Tables 2

2.1 实验动物数量分析实验开始入选Wistar大鼠100只，造模成功70只，二次开胸注射细胞或细胞培养液后成活40只，干预4周后对存活的40只大鼠的相关实验数据进行统计分析。 2.2 各组心肌组织病理形态和心肌梗死面积假手术组大鼠的心肌纤维横纹清晰，未见有增生、肥大、萎缩及断裂，未见颗粒样变性、脂肪变性，未见液化性肌溶解及凝固性坏死，未见纤维化；心肌细胞核居中，结构清晰，核两端无脂褐素沉着；心肌纤维间质无水肿及炎性细胞浸润，未见明显的病理形态。模型组大鼠心肌梗死区炎症反应明显，大量中性粒细胞浸润；心肌纤维断裂、排列紊乱。细胞培养液组及骨髓间充质干细胞组病理改变与模型组相似，但病变范围较模型组有所减少，部分坏死心肌组织由新生的肉芽组织取代，见图1。模型组、细胞培养液组和骨髓间充质干细胞组心肌梗死面积均高于假手术组(P < 0.05)；骨髓间充质干细胞组梗死面积较模型组和细胞培养液组有所减少(P < 0.05)，见表1。 2.3 心肌钾离子通道蛋白Kv1.2、Kv1.5和心肌特异性蛋白cTnT的表达各组β-actin均有表达。与假手术组比较，模型组、细胞培养液组和骨髓间充质干细胞组Kv1.2和Kv1.5表达水平均降低(P < 0.05)，而cTnT表达水平均升高(P < 0.05)；与模型组及细胞培养液组比较，骨髓间充质干细胞组Kv1.5和cTnT表达水平均提高(P < 0.05)，而Kv1.2表达水平降低(P < 0.05)，见图2和表2。 2.4 细胞凋亡因子Caspase-3和自噬因子Bcl-2的表达由表3和图3可见，模型组、骨髓间充质干细胞组和细胞培养液组心肌梗死组织中细胞凋亡因子Caspase-3和自噬因子Bcl-2的表达高于假手术组(P < 0.05)，骨髓间充质干细胞组低于模型组和细胞培养液组(P < 0.05)。 2.5 梗死区心肌组织中一氧化氮和超氧化物歧化酶表达水平由表4可见，模型组、细胞培养液组和骨髓间充质干细胞组超氧化物歧化酶表达水平均低于假手术组(P < 0.05)，骨髓间充质干细胞组高于模型组及细胞培养液组(P < 0.05)；模型组、细胞培养液组和骨髓间充质干细胞组一氧化氮水平均高于假手术组(P < 0.05)，但模型组、细胞培养液组和骨髓间充质干细胞组间差异无显著性意义(P > 0.05)。"

References

[1] 吕光,雷建华,曹秀云.不同年龄组急性心肌梗死危险因素临床分析[J].中国实用医药,2008,3(28):91-92.

[2] Brunskill SJ, Hyde CJ, Doree CJ, et al. Route of delivery and baseline left ventricular ejection fraction, key factors of bone-marrow-derived cell therapy for ischaemic heart disease. Eur J Heart Fail. 2009;11(9):887-896.

[3] 罗涛,李玉明.心肌梗死后心室电重构的细胞核分子生物学机制[J].中国病理生理杂志,2008,24(11):2276-2281.

[4] 杨琳,黄诒焯. 心肌细胞离子通道与心律失常[J].中国心脏起搏与心电生理杂志,2003,17(2):81-88.

[5] Deng XL, Sun HY, Lau CP, et al. Properties of ion channels in rabbit mesenchymal stem cells from bone marrow. Biochem Biophys Res Commun. 2006;348(1):301-309.

[6] Cai B, Wang G, Chen N, et al. Bone marrow mesenchymal stem cells protected post-infarcted myocardium against arrhythmias via reversing potassium channels remodelling. J Cell Mol Med. 2014;18(7):1407-1416.

[7] Pfeffer MA, Pfeffer JM, Steinberg C, et al. Survival after an experimental myocardial infarction: beneficial effects of long-term therapy with captopril. Circulation. 1985;72(2): 406-412.

[8] Gao L, Bledsoe G, Yin H, et al. Tissue kallikrein-modified mesenchymal stem cells provide enhanced protection against ischemic cardiac injury after myocardial infarction. Circ J. 2013;77(8):2134-2144.

[9] Takagawa J, Zhang Y, Wong ML, et al. Myocardial infarct size measurement in the mouse chronic infarction model: comparison of area- and length-based approaches. J Appl Physiol (1985). 2007;102(6):2104-2111.

[10] 杨国勋,刘唐威,钟国强,等.骨髓间充质干细胞移植对心肌梗死后心功能的影响[J]. 中华危重病急救医学, 2007, 19(7): 428-430.

[11] 梁丽玲,杨庭树,李萍,等. 同种异体骨髓间充质干细胞移植治疗大鼠急性心肌梗死[J]. 中国组织工程研究,2014,18(37): 5983-5987.

[12] Chen H, Xia R, Li Z, et al. Mesenchymal Stem Cells Combined with Hepatocyte Growth Factor Therapy for Attenuating Ischaemic Myocardial Fibrosis: Assessment using Multimodal Molecular Imaging. Sci Rep. 2016;6:33700.

[13] Hua P, Liu LB, Liu JL, et al. Inhibition of apoptosis by knockdown of caspase-3 with siRNA in rat bone marrow mesenchymal stem cells. Exp Biol Med (Maywood). 2013; 238(9):991-998.

[14] Li L, Guan Q, Dai S, et al. Integrin β1 Increases Stem Cell Survival and Cardiac Function after Myocardial Infarction. Front Pharmacol. 2017;8:135.

[15] 李艳菊,刘洋,郭坤元.骨髓间充质干细胞对衰老心脏血管内皮生长因子表达的影响[J].中国组织工程研究,2012,16(41): 7709-7712.

[16] Kheloufi M, Rautou PE, Boulanger CM. Autophagy in the cardiovascular system. Med Sci (Paris). 2017;33(3):283-289.

[17] Lee SI, Kim BG, Hwang DH, et al. Overexpression of Bcl-XL in human neural stem cells promotes graft survival and functional recovery following transplantation in spinal cord injury. J Neurosci Res. 2009;87(14):3186-3197.

[18] De Meyer GR, Martinet W. Autophagy in the cardiovascular system. Biochim Biophys Acta. 2009;1793(9):1485-1495.

[19] Zhong Z, Hu J, Sun Y, et al. Impact of mesenchymal stem cells transplantation on myocardial myocardin-related transcription factor-A and bcl-2 expression in rats with experimental myocardial infarction. Zhonghua Xin Xue Guan Bing Za Zhi. 2015;43(6):531-536.

[20] 吕延杰,蔡本志,艾静,等.骨髓间充质干细胞对心肌细胞K+电流的影响[J].中国药理通讯,2013,30(3):46.

[21] Deng XL, Lau CP, Lai K, et al. Cell cycle-dependent expression of potassium channels and cell proliferation in rat mesenchymal stem cells from bone marrow. Cell Prolif. 2007;40(5):656-670.

[22] Leanza L, Zoratti M, Gulbins E, et al. Induction of apoptosis in macrophages via Kv1.3 and Kv1.5 potassium channels. Curr Med Chem. 2012;19(31):5394-5404.