Cardiac Differentiation of Adipose Tissue-Derived Stem Cells Is Driven by BMP4 and bFGF but Counteracted by 5-Azacytidine and Valproic Acid

Sanaz Hasani; Arash Javeri; Asadollah Asadi; Masoumeh Fakhr Taha

doi:10.22074/cellj.2020.6582

Cardiac Differentiation of Adipose Tissue-Derived Stem Cells Is Driven by BMP4 and bFGF but Counteracted by 5-Azacytidine and Valproic Acid

Cell J. 2020 Oct;22(3):273-282. doi: 10.22074/cellj.2020.6582. Epub 2019 Dec 15.

Authors

Sanaz Hasani^{1

2}, Arash Javeri¹, Asadollah Asadi², Masoumeh Fakhr Taha³

Affiliations

¹ Department of Stem Cells and Regenerative Medicine, Institute for Medical Biotechnology, National Institute of Genetic Engineering and Biotechnology (NIGEB), Tehran, Iran.
² Department of Biology, Faculty of Science, University of Mohaghegh Ardabili, Ardabil, Iran.
³ Department of Stem Cells and Regenerative Medicine, Institute for Medical Biotechnology, National Institute of Genetic Engineering and Biotechnology (NIGEB), Tehran, Iran. Elrctronic Address:mftaha@nigeb.ac.ir.

Abstract

Objective: Bone morphogenetic protein 4 (BMP4) and basic fibroblast growth factor (bFGF) play important roles in embryonic heart development. Also, two epigenetic modifying molecules, 5'-azacytidine (5'-Aza) and valproic acid (VPA) induce cardiomyogenesis in the infarcted heart. In this study, we first evaluated the role of BMP4 and bFGF in cardiac trans-differentiation and then the effectiveness of 5´-Aza and VPA in reprogramming and cardiac differentiation of human adipose tissue-derived stem cells (ADSCs).

Materials and methods: In this experimental study, human ADSCs were isolated by collagenase I digestion. For cardiac differentiation, third to fifth-passaged ADSCs were treated with BMP4 alone or a combination of BMP4 and bFGF with or without 5'-Aza and VPA pre-treatment. After 21 days, the expression of cardiac-specific markers was evaluated by reverse transcription polymerase chain reaction (RT-PCR), quantitative real-time PCR, immunocytochemistry, flow cytometry and western blot analyses.

Results: BMP4 and more prominently a combination of BMP4 and bFGF induced cardiac differentiation of human ADSCs. Epigenetic modification of the ADSCs by 5'-Aza and VPA significantly upregulated the expression of OCT4A, SOX2, NANOG, Brachyury/T and GATA4 but downregulated GSC and NES mRNAs. Furthermore, pre-treatment with 5'-Aza and VPA upregulated the expression of TBX5, ANF, CX43 and CXCR4 mRNAs in three-week differentiated ADSCs but downregulated the expression of some cardiac-specific genes and decreased the population of cardiac troponin I-expressing cells.

Conclusion: Our findings demonstrated the inductive role of BMP4 and especially BMP4 and bFGF combination in cardiac trans-differentiation of human ADSCs. Treatment with 5'-Aza and VPA reprogrammed ADSCs toward a more pluripotent state and increased tendency of the ADSCs for mesodermal differentiation. Although pre-treatment with 5'-Aza and VPA counteracted the cardiogenic effects of BMP4 and bFGF, it may be in favor of migration, engraftment and survival of the ADSCs after transplantation.

Keywords: Adipose Tissue-Derived Stem Cells; BMP4; Basic Fibroblast Growth Factor; Cardiomyocyte; Small Molecules.