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

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

• GasiÅ«nienÄ— M
• Petkus G
• Matuzevičius D
• Navakauskas D
• NavakauskienÄ— R

• KMID: 2465896
• DOI: http://doi.org/10.15283/ijsc18126

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.

Keyword

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

MeSH Terms

Amniocentesis
Amniotic Fluid
Angiotensin II*
Angiotensins*
Blotting, Western
Calcium
Cell Differentiation
Cell- and Tissue-Based Therapy
Chromatin
Chromatin Assembly and Disassembly
Connexin 43
Epigenomics
Female
Fluorescent Antibody Technique
Gap Junctions
Glycolysis
Histones
Humans*
Ion Channels
Mesenchymal Stromal Cells*
Muscle Cells
Oxidative Phosphorylation
Phenotype
Polycomb Repressive Complex 1
Pregnant Women
Smegmamorpha
Stem Cells
Tissue Engineering
Transcription Factors
Up-Regulation
Angiotensin II
Angiotensins
Calcium
Chromatin
Connexin 43
Histones
Ion Channels
Polycomb Repressive Complex 1
Transcription Factors