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Title:: Effect of different concentrations of 5azacytidine on differentiation of human bone marrow mesenchymal stem cells towards cardiomyocytes

作者:: 付国伟; 赵文增; 文冰; 舒礼良; 刘春明; 刘新生; 郑州大学第一附属医院心血管外科，河南郑州 450052

Author(s):: FU Guowei; ZHAO Wenzeng; WEN Bing; SHU Liliang; LIU Chunming; LIU Xinsheng; Department of Cardiovascular Surgery, First Affiliated Hospital, Zhengzhou University, Zhengzhou 450052, Henan, China

关键词:: 人; 骨髓间充质干细胞; 心肌细胞; 5氮胞苷; 分化

Keywords:: human; mesenchymal stem cells; cardiomyocytes; 5azacytidine; differentiation

分类号:: R329.21

DOI:: -

文献标识码:: A

摘要:: 目的探讨不同浓度的5-氮胞苷（5-aza）对人骨髓间充质干细胞（hMSCs）向心肌细胞分化的影响。方法抽取人胸骨骨髓3～5 ml，采用密度梯度离心法和贴壁法进行hMSCs的分离培养。于第3代生长状态良好的hMSCs中，分别加入3、5、10、20和40 μmol/L5aza处理24 h,更换新鲜的培养基继续培养；对照组的细胞始终用完全培养基培养。光镜下动态观察细胞形态的变化，4周后用免疫组化染色法检测心肌肌钙蛋白I（cTnI）、连接蛋白43（Cx43）的表达。结果经5-aza诱导处理后，hMSCs的形态变长、增大，排列更加紧密、整齐。免疫组化染色法检测表明，对照组与3 μmol/L5aza组cTnI、Cx43的表达呈阴性，5、10、20和40 μmol/L5aza组cTnI和Cx43的表达均呈阳性。在一定范围内（<10 μmol/L的5aza），随着5-aza浓度的增加，hMSCs向心肌细胞的转化率逐渐增高；但高浓度（>10 μmol/L） 5-aza的毒性可造成细胞大量死亡。结论 hMSCs经适当浓度的5aza处理后，可向心肌细胞分化，10 μmol/L的5-aza为最适诱导浓度。

Abstract:: AIM To explore the effect of different concentrations of 5-azacytidine (5-aza) on the differentiation of human bone marrow mesenchymal stem cells (hMSCs) towards cardiomyocytes. METHODS Bone marrow samples were derived from human sternum, and hMSCs were isolated and cultivated by density gradient centrifugation (Percoll, 1.077 g/ml) and adherent filtration. The third passage hMSCs were exposed to the medium containing different concentrations of 5-aza (3, 5, 10, 20 and 40 μmol/L 5-aza) for 24 h for induction cellular differentiation and then cultured in new medium. Control group was cultured in complete medium. The differentiation of hMSCs was observed by light microscope and myocardiumspecific cardiac troponin I (cTnI) and connexin43(Cx43) were examined with immunohistochemistry after 4 weeks. RESULTS The hMSCs were enlarged, lengthened and closely arranged after chemical inducement. Immunohistochemical eramination revealed negative expression of cTnI and Cx43 in 3 μmol/L group and control group and positive expression in other groups. With the increasing 5-aza concentration in extents (<10 μmol/L5aza), the differentiation rates of hMSCs increased gradually. There were massive death cells in high concentration groups (>10 μmol/L5aza）. CONCLUSION hMSC can be induced to differentiate into cardiomyocytes by 5-aza in suitable concentration and 10 μmol/L is the optimal concentration.

参考文献/References

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不同浓度的5-氮胞苷对人骨髓间充质干细胞向心肌细胞分化的影响(PDF)

《心脏杂志》[ISSN:1009-7236/CN:61-1268/R]

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