Differentiation of human adipose-derived stem cells into beating cardiomyocytes

被引:164
作者
Choi, Yu Suk [1 ,2 ]
Dusting, Gregory J. [1 ,2 ]
Stubbs, Samantha [1 ,2 ]
Arunothayaraj, Sandeep [1 ]
Han, Xiao Lian [1 ]
Collas, Philippe [3 ]
Morrison, Wayne A. [1 ,2 ]
Dilley, Rodney J. [1 ,2 ,4 ]
机构
[1] OBrien Inst, Melbourne, Vic, Australia
[2] Univ Melbourne, Dept Surg, St Vincents Hosp, Melbourne, Vic, Australia
[3] Univ Oslo, Inst Basic Med Sci, Fac Med, Oslo, Norway
[4] Australian Tissue Engn Ctr, Melbourne, Vic, Australia
基金
英国医学研究理事会;
关键词
adipose-derived stem cell; cardiomyocyte; cardiomyogenic differentiation; epigenetic modification; co-culture; HISTONE DEACETYLASE ACTIVITY; MARROW STROMAL CELLS; IN-VITRO; TISSUE;
D O I
10.1111/j.1582-4934.2010.01009.x
中图分类号
Q2 [细胞生物学];
学科分类号
071009 ; 090102 ;
摘要
Human adipose-derived stem cells (ASCs) may differentiate into cardiomyocytes and this provides a source of donor cells for tissue engineering. In this study, we evaluated cardiomyogenic differentiation protocols using a DNA demethylating agent 5-azacytidine (5-aza), a modified cardiomyogenic medium (MCM), a histone deacetylase inhibitor trichostatin A (TSA) and co-culture with neonatal rat cardiomyocytes. 5-aza treatment reduced both cardiac actin and TropT mRNA expression. Incubation in MCM only slightly increased gene expression (1.5- to 1.9-fold) and the number of cells co-expressing nkx2.5/sarcomeric alpha-actin (27.2% versus 0.2% in control). TSA treatment increased cardiac actin mRNA expression 11-fold after 1 week, which could be sustained for 2 weeks by culturing cells in cardiomyocyte culture medium. TSA-treated cells also stained positively for cardiac myosin heavy chain, alpha-actin, TropI and connexin43; however, none of these treatments produced beating cells. ASCs in non-contact co-culture showed no cardiac differentiation; however, ASCs co-cultured in direct contact co-culture exhibited a time-dependent increase in cardiac actin mRNA expression (up to 33-fold) between days 3 and 14. Immunocytochemistry revealed co-expression of GATA4 and Nkx2.5, alpha-actin, TropI and cardiac myosin heavy chain in CM-DiI labelled ASCs. Most importantly, many of these cells showed spontaneous contractions accompanied by calcium transients in culture. Human ASC (hASC) showed synchronous Ca2+ transient and contraction synchronous with surrounding rat cardiomyocytes (106 beats/min.). Gap junctions also formed between them as observed by dye transfer. In conclusion, cell-to-cell interaction was identified as a key inducer for cardiomyogenic differentiation of hASCs. This method was optimized by co-culture with contracting cardiomyocytes and provides a potential cardiac differentiation system to progress applications for cardiac cell therapy or tissue engineering.
引用
收藏
页码:878 / 889
页数:12
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