中国组织工程研究

中国组织工程研究 ›› 2010, Vol. 14 ›› Issue (36): 6828-6832.doi: 10.3969/j.issn.1673-8225.2010.36.041

• 干细胞基础实验 basic experiments of stem cells • 上一篇    下一篇

磷酸鞘胺醇1协同条件培养液促进人脐带间充质干细胞向心肌样细胞的分化

赵振强，陈志斌，蔡美华，王淑荣，陈  蓉，王  埮，袁昆雄，容琼文   

  1. 海南医学院附属医院神经内科，海南省海口市  570102
  • 出版日期:2010-09-03 发布日期:2010-09-03
  • 通讯作者: 陈志斌，博士，教授，主任医师，海南医学院附属医院神经内科，海南省海口市 570102
  • 作者简介:赵振强★，男，1975年生，山西省阳泉市人，汉族，2004年重庆医科大学毕业，硕士，讲师，主治医师，主要从事干细胞与脑血管病治疗方面的研究。
  • 基金资助:

    海南省科技厅重点项目(20061003)，课题名称：人脐带间质干细胞分化的研究。

Cardiomyogenesis of human umbilical cord mesenchymal stem cells induced by conditional culture in the presence of sphingosine-1-phosphate

Zhao Zhen-qiang, Chen Zhi-bin, Cai Mei-hua, Wang Shu-rong, Chen Rong, Wang Tan，Yuan Kun-xiong, Rong Qiong-wen   

  1. Department of Neurology, Affiliated Hospital of Hainan Medical College, Haikou   570102, Hainan Province, China
  • Online:2010-09-03 Published:2010-09-03
  • Contact: Chen Zhi-bin, Doctor, Professor, Chief physician, Department of Neurology, Affiliated Hospital of Hainan Medical College, Haikou 570102, Hainan Province, China mumu1202001@yahoo.com.cn
  • About author:Zhao Zhen-qiang★, Master, Lecturer, Attending physician, Department of Neurology, Affiliated Hospital of Hainan Medical College, Haikou 570102, Hainan Province, China
  • Supported by:

    Key Program of Science and Technology Bureau of Hainan Province, No. 20061003*

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摘要：

背景: 研究显示磷酸鞘胺醇1(sphingosine- 1-phosphate，S1P)可诱导脂肪干细胞分化成平滑肌细胞。S1P是否可替代5-氮杂胞苷作为间充质细胞分化为心肌细胞的诱导剂尚不清楚。
目的：探讨S1P促进在不同培养液诱导下的人脐带间充质干细胞向心肌样细胞分化的可能性。
方法：收集培养人心肌细胞的条件培养基(cardiomyocytes   condition   medium, CMCM)，分别用CMCM和(或)S1P培养人
脐带间充质干细胞，在培养的第1，5，10天，观察人脐带间充质干细胞的形态学变化。培养10 d后，应用免疫细胞化学和膜片钳鉴定细胞表型及细胞功能。
结果与结论：随着培养时间的延长，CMCM组和CMCM+S1P组的一些细胞逐渐增大，拉长，与邻近细胞连接并形成肌管样结构，其中一些细胞聚集成簇。在CMCM+S1P组中，细胞出现特殊的垂直对齐梯田状，类闰盘样排列。同时，免疫细胞化学染色结果显示，CMCM组和CMCM+S1P组中一些细胞强烈表达心肌特异性抗体(心肌肌球蛋白重链和横纹肌辅肌动蛋白α)，说明人脐带间充质干细胞在CMCM诱导下可分化为心肌样细胞。膜片钳仅在CMCM+S1P组的部分细胞记录到一个快速上行，但无平台期的动作电位，以及一个电压依赖性内向电流和一个电压依赖性外向电流。说明S1P在促进人脐带间充质干细胞向心肌样细胞分化和功能整合中发挥关键作用。

关键词: 磷酸鞘胺醇1, 人脐带间充质干细胞, 条件培养液, 心肌细胞, 分化

Abstract:

BACKGROUND: Several studies have demonstrated that sphingosine-1-phosphate (S1P) can induce the differentiation of adipose-derived stem cells differentiation into smooth muscle cells. Whether S1P, rather than 5-azacytidine, can be used as an inducer of mesenchymal stem cells differentiation into cardiomyocytes remains unclear. 
OBJECTIVE: To investigate the possibility that S1P promotes human umbilical cord mesenchymal stem cells (HUMSCs) differentiation into cardiomyocytes in the presence of different culture media.
METHODS: HUMSCs were cultured with cardiomyocyte conditional medium (CMCM) and/or S1P media. At 1, 5, and 10 days of culture, morphological changes of HUMSCs were observed. After culture for 10 days, the induced cells were confirmed by immunocytochemical analysis and patch clamp in terms of cell phenotype and function.
RESULTS AND CONCLUSION: During the induction, some cells gradually enlarged, elongated, connected with adjacent cells, and formed myotube-like structures, and some cells congregated into cell clusters in the CMCM and CMCM+S1P groups. In the CMCM+S1P group, cells exhibited special perpendicular terrace-shaped, intercalated disc-like arrangement. Immunohistochemistry results revealed that some cells strongly express specific antibodies against sarcomeric myosin and α-actinin in the CMCM and CMCM+S1P groups. These findings suggest that HUMSCs can be induced to differentiate into cardiomyocytes. Through the use of patch clamp technique, a rapid ascending, but without plateau phase, action potential, a voltage dependent inward current, and a voltage dependent outward current were recorded in some cells from the CMCM+S1P group. These findings indicate that S1P plays a key role in promoting cardiomyogenic differentiation of HUMSCs and functional integration.

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