我们的网站为什么显示成这样？

可能因为您的浏览器不支持样式，您可以更新您的浏览器到最新版本，以获取对此功能的支持，访问下面的网站，获取关于浏览器的信息：

|本期目录/Table of Contents|

间接接触共培养下骨髓间充质干细胞分化为心肌样细胞及相关调控基因的时序表达

《心脏杂志》[ISSN:1009-7236/CN:61-1268/R]

期数:
2010年第6期
页码:
801-806
栏目:
基础研究
出版日期:
2010-08-23

文章信息/Info

Title:
Transdifferentiation of bone marrow mesenchymal stem cells into cardiomyocyte-like cells by indirect contact with cardiomyocytes and temporal expression of regulatory genes
作者:
曾俊义1; 魏云峰1; 汪泱2; 娄远蕾2; 张梅1; 应国秋1; 徐信群1
南昌大学第一附属医院：1.心血管内科，2.泌尿外科研究所，江西 南昌 330006
Author(s):
ZENG Jun-yi1;  WEI Yun-feng1;  WANG Yang2;  LOU Yuan-lei2;  ZHANG Mei1;  YING Guo-qiu1;  XU Xin-qun1
1.Department of Cardiology, 2.Institute of Urinary Surgery, First Affiliated Hospital, Nanchang University, Nanchang 330006, Jiangxi, China
关键词:
骨髓间充质干细胞; 间接接触; 分化; 调控基因; 心肌细胞
Keywords:
bone marrow mesenchymal stem cells;  indirect contact;  differentiation;  genes;  cardiomyocytes
分类号:
R329.21
DOI:
-
文献标识码:
A
摘要:
目的: 研究间接接触共培养条件下骨髓间充质干细胞(mesenchymal stem cells,MSCs)向心肌细胞(myocadium-lkce cells，CMs)的分化及相关调控基因的时序表达；筛选MSCs定向分化为CMs的重要调控基因。方法： 将MSCs与CMs按1∶5的比例进行间接接触共培养,连续观察两周,在相差显微镜下观察MSCs的形态变化。采用免疫荧光染色法检测心肌特征性肌动蛋白α(α-actin)和心脏肌钙蛋白T(cTnT)的表达。应用半定量RT-PCR分析TGF-β、Nkx-2.5、GATA-4、MEF-2C及TEF-1等相关调控基因在分化过程中的时序表达。结果: 共培养后，MSCs的体积增大,其梭形形态逐渐变短变粗近似棒状或椭圆形,细胞之间形成连接,排列方向趋于一致。共培养两周时，α-actin及cTnT阳性细胞的比例分别为29.63%和27.38%。TGF-β、Nkx-2.5、GATA-4和MEF-2C基因在共培养后1 d表达开始增强,诱导后7 d达高峰,以后虽有所下降但仍维持在较高水平；TEF-1基因在诱导过程中表达无明显变化。结论: 间接接触共培养条件下，MSCs可分化为心肌细胞。在此过程中,TGF-β、Nkx-2.5、GATA-4和MEF-2C基因可能是调控MSCs定向分化为心肌样细胞的重要调控基因。
Abstract:
AIM: To investigate the transdifferentiation of bone marrow mesenchymal stem cells (MSCs) into cardiomyocytes and the temporal expression of regulatory genes in the simulated myocardial microenvironment in which MSCs were co-cultured with cardiomyocytes by indirect contact in vitro and to screen some important genes in this differentiation. METHODS: MSCs isolated from adult rats were cocultured with CMs obtained from neonatal rat ventricles at a 1∶5 ratio in a dual-chamber dish separated by a semipermeable membrane for 2 weeks. During this indirect contact coculture procedure, cardiomyogenic differentiation was confirmed by phase contrast microscope and immunostaining against α-actin and cardiac troponin-T (cTnT), and reverse transcriptase-polymerase chain reaction was performed for temporal expression of regulatory genes TGF-β, Nkx-2.5, GATA-4, MEF-2C and TEF-1. RESULTS: After coculture with CMs, MSCs showed a stick-like or elliptical morphology. Immunofluorescent stain results revealed that α-actin- and cardiac troponin T (cTnT)-positive cells were found in MSCs at day 5 after coculture with CMs, and 29.63% of MSCs were positive for α-actin and 27.38% for cTnT at 14 days. RT-PCR demonstrated that the expression level of TGF-β, Nkx-2.5, GATA-4 and MEF-2C genes began to increase at day 1 after coculture, reached their peak at day 7 and remained relatively high afterwards, even though there was a slight decrease. Only insignificant changes were observed in the expression level of TEF-1 gene in the coculture. CONCLUSION: Indirect contact with CMs (1∶5) is conducive for MSCs to differentiate into CMs in vitro. TGF-β, Nkx-2.5, GATA-4 and MEF-2C genes may play a crucial role during transdifferentiation.

参考文献/References

［1］Wang JS, Shum-Tim D, Chedrawy E, et al. The coronary delivery of marrow stromal cells for myocardial regeneration: Pathophysiologic and therapeutic implications［J］. J Thorac Cardiovasc Surg, 2001, 122(4):699-705.

［2］Mummery C, Ward-van Oostwaard D, Doevendans P, et al. Differentiation of human embryonic stem cells to cardiomyocytes: role of coculture with visceral endoderm-Like cells［J］. Circulation, 2003, 107(21):2733-2740.

［3］Zaffran S, Frasch M. Early signals in cardiac development［J］. Circ Res, 2002, 91(6):457-469.

［4］Bruneau BG. Transcriptional regulation of vertebrate cardiac morphogenesis［J］. Circ Res, 2002, 90(5):509-519.

［5］Pittenger MF, Mackay AM, Beck SC, et al. Multilineage potential of adult human mesenchymal stem cells［J］. Science, 1999, 284(5411):143-147.

［6］Toma C, Pittenger MF, Cahill KS, et al. Human mesenchymal Stem cells differentiate to a cardiomyocyte phenotype in the adult murine heart［J］. Circulation, 2002, 105(1):93-98.

［7］Wang T, Xu Z, Jiang W, et al.Cell-to-cell contact induces mesenchymal stem cell to differentiate into cardiomyocyte and smooth muscle cell［J］. Int J Cardiol, 2006, 109(1):74-81.

［8］Rangappa S, Entwistle JW, Wechsler AS, et al. Cardiomyocyte-mediated contact programs human mesenchymal stem cells to express cardiogenic phenotype［J］. J Thorac Cardiovasc Surg, 2003, 126(1):124-132.

［9］ Yoon J, Shim WJ, Ro YM, et al. Transdifferentiation of mesenchymal stem cells into cardiomyocytes by direct cell-to-cell contact with neonatal cardiomyocyte but not adult cardiomyocytes［J］. Ann Hematol, 2005, 84(11):715-721.

［10］Li H, Yu B, Zhang Y, et al. Jagged1 protein enhances the differentiation of mesenchymal stem cells into cardiomyocytes［J］. Biochem Biophys Res Commun, 2006, 341(2):320-325.

［11］Lokuta A, Kirby MS, Gaa ST, et al. On establishing primary cultures of neonatal rat ventricular myocytes for analysis over long periods［J］. J Cardiovasc Electrophysiol, 1994, 5(1):50-62.

［12］Banyasz T, Lozinskiy I, Payne CE, et al. Transformation of adult rat cardiac myocytes in primary culture［J］. Exp Physiol, 2008, 93(3):370-382.

［13］Li X, Yu X, Lin Q, et al. Bone marrow mesenchymal stem cells differentiate into functional cardiac phenotypes by cardiac microenvironment［J］. J Mol Cell Cardiol, 2007, 42(2):295-303.

[14]Spradling A, Drummond-Barbosa D, Kai T. Stem cells find their niche［J］. Nature, 2001, 414(6859):98-104.

[15]Song YS, Lee HJ, Park IH, et al. Potential differentiation of human mesenchymal stem cell transplanted in rat corpus cavernosum toward endothelial or smooth muscle cells［J］. Int J Impot Res, 2007, 19(4):378-385.

[16]Hwang NS, Varghese S, Puleo C, et al. Morphogenetic signals from chondrocytes promote chondrogenic and osteogenic differentiation of mesenchymal stem cellsv［J］. J Cell Physiol, 2007, 212(2):281-284.

[17]Matsushita K, Wu Y, Okamoto Y, et al. Local renin angiotensin expression regulates human mesenchymal stem cell differentiation to adipocytes［J］. Hypertension, 2006, 48(6):1095-1102.

[18]Kawakami Y, Rodriguez-León J, Belmonte JC. The role of TGF betas and Sox9 during limb chondrogenesis［J］. Curr Opin Cell Biol, 2006, 18(6):723-729.

[19]Burch JB. Regulation of GATA gene expression during vertebrate development［J］. Semin Cell Dev Bio, 2005, 16(1):71-81.

[20]Wang Y, Liu L, Xia Z. Brain-derived neurotrophic factor stimulates the transcriptional and neuroprotective activity of myocyte-enhancer factor 2C through an ERK1/2-RSK2 signaling cascade［J］. J Neurochem, 2007, 102(3):957-966.

[21]Toko H, Zhu W, Takimoto E, et al. Csx/Nkx2-5 is required for homeostasis and survival of cardiac myocytes in the adult heart［J］. J Biol Chem, 2002, 277(27):24735-24743.

[22]Iida K, Hidaka K, Takeuchi M, et al. Expression of MEF2 genes during human cardiac development［J］. Tohoku J Exp Med, 1999, 187(1):15-23.

[23]Morkin E. Control of cardiac myosin heavy chain gene expression［J］. Microsc Res Tech, 2000, 50(6):522-531.

备注/Memo

备注/Memo:
收稿日期：2009-11-02.基金项目：江西省卫生厅重大招标项目资助（200602） 通讯作者：魏云峰，主任医师，主要从事干细胞与缺血性心脏病的研究Email:weiyunfeng2007@126.com 作者简介：曾俊义，医师，硕士Email:zjy2005052@163.com
更新日期/Last Update: 2010-08-22