MiR-21, MiR-29a, GATA4, and MEF2c Expression Changes in Endothelin-1 and Angiotensin II Cardiac Hypertrophy Stimulated Isl-1+Sca-1+c-kit+ Porcine Cardiac Progenitor Cells In Vitro.

Katrin Zlabinger,Mariann Gyöngyösi,Julia Mester-Tonczar,Johannes Winkler,Denise Traxler,Bruno Podesser,Andreas Spannbauer,Dominika Lukovic,Alfred Gugerell

doi:10.3390/cells8111416

Abstract

Cost- and time-intensive porcine translational disease models offer great opportunities to test drugs and therapies for pathological cardiac hypertrophy and can be supported by porcine cell culture models that provide further insights into basic disease mechanisms. Cardiac progenitor cells (CPCs) residing in the adult heart have been shown to differentiate in vitro into cardiomyocytes and could contribute to cardiac regeneration. Therefore, it is important to evaluate their changes on the cellular level caused by disease. We successfully isolated Isl1+Sca1+cKit+ porcine CPCs (pCPCs) from pig hearts and stimulated them with endothelin-1 (ET-1) and angiotensin II (Ang II) in vitro. We also performed a cardiac reprogramming transfection and tested the same conditions. Our results show that undifferentiated Isl1+Sca1+cKit+ pCPCs were significantly upregulated in GATA4, MEF2c, and miR-29a gene expressions and in BNP and MCP-1 protein expressions with Ang II stimulation, but they showed no significant changes in miR-29a and MCP-1 when stimulated with ET-1. Differentiated Isl1+Sca1+cKit+ pCPCs exhibited significantly higher levels of MEF2c, GATA4, miR-29a, and miR-21 as well as Cx43 and BNP with Ang II stimulation. pMx-MGT-transfected Isl1+Sca1+cKit+ pCPCs showed significant elevations in MEF2c, GATA4, and BNP expressions when stimulated with ET-1. Our model demonstrates that in vitro stimulation leads to successful Isl1+Sca1+cKit+ pCPC hypertrophy with upregulation of cardiac remodeling associated genes and profibrotic miRNAs and offers great possibilities for further investigations of disease mechanisms and treatment.

Highlights

Cardiac hypertrophy is characterized by the increase of the heart size, as well as cardiomyocyte size, and mass in response to pathological and physiological mechanisms [1,2,3]
Specialized adult cardiac progenitor cell (CPC) populations can be found in mammals in interstitial places of the heart near cardiomyocytes and are known to express progenitor cell markers like stem cell antigen-1 (Sca-1), insulin gene enhancer protein-1 (Isl-1), or the stem cell growth factor receptor c-kit [7,8]
We evaluated porcine CPCs (pCPCs) hypertrophy by measuring the mean cell area, quantification of cellular proteins with immunofluorescence staining, and mRNA and miRNA expression focusing on GATA4 and myocyte-specific enhancer factor 2C (MEF2c) as well as miR-21 and miR-29a

Summary

Introduction

Cardiac hypertrophy is characterized by the increase of the heart size, as well as cardiomyocyte size, and mass in response to pathological and physiological mechanisms [1,2,3]. Isl-1-positive CPCs originate from the second heart field in cardiac development, and they have been shown to give rise to cardiac cells of different lineages such as cardiomyocytes, smooth muscle cells, and vascular endothelial cells [9,10]. They are part of the outflow tract, right/left atrium, and ventricle as well as vessels from the inflow tract, but they are reported to be very rare in proportion to other cells [10,11,12]

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