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Int J Stem Cells. 2019 Jul 31;12(2):251-264. doi: 10.15283/ijsc18126.

Angiotensin II and TGF-β1 Induce Alterations in Human Amniotic Fluid-Derived Mesenchymal Stem Cells Leading to Cardiomyogenic Differentiation Initiation.

Author information

1
Department of Molecular Cell Biology, Institute of Biochemistry, Life Sciences Center, Vilnius University, Vilnius, Lithuania.
2
Electronic Systems Department, Electronics Faculty, Vilnius Gediminas Technical University, Vilnius, Lithuania.

Abstract

Background and Objectives:

Human amniotic fluid-derived mesenchymal stem cells (AF-MSCs) may be a valuable source for cardiovascular tissue engineering and cell therapy. The aim of this study is to verify angiotensin II and transforming growth factor-beta 1 (TGF-β1) as potential cardiomyogenic differentiation inducers of AF-MSCs.

Methods and Results:

AF-MSCs were obtained from amniocentesis samples from second-trimester pregnant women, isolated and characterized by the expression of cell surface markers (CD44, CD90, CD105 positive; CD34 negative) and pluripotency genes (OCT4, SOX2, NANOG, REX1). Cardiomyogenic differentiation was induced using different concentrations of angiotensin II and TGF-β1. Successful initiation of differentiation was confirmed by alterations in cell morphology, upregulation of cardiac genes-markers NKX2-5, TBX5, GATA4, MYH6, TNNT2, DES and main cardiac ion channels genes (sodium, calcium, potassium) as determined by RT-qPCR. Western blot and immunofluorescence analysis revealed the increased expression of Connexin43, the main component of gap junctions, and Nkx2.5, the early cardiac transcription factor. Induced AF-MSCs switched their phenotype towards more energetic and started utilizing oxidative phosphorylation more than glycolysis for energy production as assessed using Agilent Seahorse XF analyzer. The immune analysis of chromatin-modifying enzymes DNMT1, HDAC1/2 and Polycomb repressive complex 1 and 2 (PRC1/2) proteins BMI1, EZH2 and SUZ12 as well as of modified histones H3 and H4 indicated global chromatin remodeling during the induced differentiation.

Conclusions:

Angiotensin II and TGF-β1 are efficient cardiomyogenic inducers of human AF-MSCs; they initiate alterations at the gene and protein expression, metabolic and epigenetic levels in stem cells leading towards cardiomyocyte- like phenotype formation.

KEYWORDS:

Amniotic fluid; Cardiac; Cell differentiation; Chromatin; Histones; Myocytes

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