Cardiac spheroids as promising in vitro models to study the human heart microenvironment

Liudmila Polonchuk,Carmine Gentile,Mamta Chabria,Gemma Figtree,Laura Badi,Michael J Davies,Jean-Christophe Hoflack

doi:10.1038/s41598-017-06385-8

Abstract

Three-dimensional in vitro cell systems are a promising alternative to animals to study cardiac biology and disease. We have generated three-dimensional in vitro models of the human heart (“cardiac spheroids”, CSs) by co-culturing human primary or iPSC-derived cardiomyocytes, endothelial cells and fibroblasts at ratios approximating those present in vivo. The cellular organisation, extracellular matrix and microvascular network mimic human heart tissue. These spheroids have been employed to investigate the dose-limiting cardiotoxicity of the common anti-cancer drug doxorubicin. Viability/cytotoxicity assays indicate dose-dependent cytotoxic effects, which are inhibited by the nitric oxide synthase (NOS) inhibitor L-NIO, and genetic inhibition of endothelial NOS, implicating peroxynitrous acid as a key damaging agent. These data indicate that CSs mimic important features of human heart morphology, biochemistry and pharmacology in vitro, offering a promising alternative to animals and standard cell cultures with regard to mechanistic insights and prediction of toxic effects in human heart tissue.

Highlights

Engineering of three-dimensional myocardial tissues presents several advantages for cultures of primary cardiomyocytes (CMs) to study human heart biology, physiology and pharmacology[1,2,3]
To investigate the roles played by the microenvironment in the human heart we have developed a 3D in vitro “cardiac spheroid” model of the human heart
Hanging drop cultures of cells normally found within the human heart were prepared using human coronary artery endothelial cells (ECs), cardiac fibroblasts derived from iPSCs and either human primary adult cardiomyocytes or iPSC-derived cardiomyocytes. hCMs were isolated from frozen heart sections, retaining the biochemical, morphological and physiological features of fresh human heart tissue (Fig. S1a–c)

Summary

Introduction

Engineering of three-dimensional myocardial tissues presents several advantages for cultures of primary cardiomyocytes (CMs) to study human heart biology, physiology and pharmacology[1,2,3]. Fernandes et al.[13] have shown that stem cell-derived CMs and cardiac progenitor cells of different sources have similar regenerative properties in an in vivo myocardial infarction model in mice, a deeper understanding of the requirements for optimal engineering of a human heart model in vitro is important[14]. Recent in vitro and in vivo data suggest an alternative mechanism to target DOX/ROS-mediated cardiotoxic effects, which may involve NO synthases (NOS)[36,37,38,39,40,41] This led us to investigate if, and how, NO mediates DOX-induced cardiotoxic effects in human heart cells, and the dual cardioprotective and cardiotoxic role of this species, which highlights the importance of cellular crosstalk

Methods

Results

Conclusion