New recellularizable cardio-matrix scaffold as a base for alternative in vitro models for the cardiotoxicity risk prediction
Authors: Bernava G , Buaron M , Iop L 
Published: May 2024

Abstract

Funding Acknowledgements
Type of funding sources: Public Institution(s). Main funding source(s): SID 227944/22
DOR 2331157/23

Background
Cardiotoxicity is the occurrence of cardiac dysfunction induced by pathophysiological stimuli as a result of toxic effects. A considerable number of drugs, including chemotherapics, antibiotics antidepressants but also antiarrhythmics, could cause an alteration of cardiac physiology, and then cardiac damage. These include disturbances in ventricular repolarization and QT interval, arrhythmias, bradycardia, tachycardia, decreases in left ventricular ejection fraction, and congestive heart failure.
Precinical evaluation of drug cardiotoxicity has used animal models, which tend to be expensive, have lo throughput, and have limitations as not ever faithfully reflect the human pathophysiology, while current two-dimensional cellular models revealed to be not sufficient and specific.

Purpose
The aim of this study is to develop a myocardial tissue-like scaffold, in order to realize an efficient and easy-replicable in vitro model for drug cardiotoxicity prediction. This must be able to mimic the morphological, cellular, and electrophysiological complexity of the intact adult human myocardium.

Material and Methods
Starting from fresh porcine hearts, cardiac specimens were isolated from left ventricles using a scalpel, sectioned with a vibratome and punched with a biopsy puncher obtaining myocardial patches. Thus, samples were decellularized through a novel serial decellularization treatment based on osmotic shock and detergents. The decellularization procedure exploited a new automated, dynamic perfusion device for a standardized and optimized removal of cardiac cells and non-ECM proteins. These samples were analysed for decellularization effectiveness by DNA quantification, histology and immunofluorescence staining to evaluate cell removal and extracellular matrix integrity. Moreover, they were tested for cytocompatibility using human mesenchymal stem cells and cardiac progenitors, also derived from induced pluripotent stem cells. Cellular adhesion, behaviour, vitality, proliferation, and possible apoptotic effects were tested, too.

Results
In order to advance a robust bioelectronics model, an automated platform was developed to generate a three-dimensional, bioengineered replica of human myocardial tissue. With respect to the common decellularization protocol characterized by mild agitation, results based on the automated, dynamic treatments have shown a superior ability to obtain acellular, biocompatible scaffolds in terms of cell elements’ removal and extracellular matrix preservation, as well as time- and cost-effectiveness. Indeed, these decellularized scaffolds allow cell penetration, adhesion and survival.

Conclusions
Further experiments will be focused on evaluating functional bioengineered myocardial tissues with electrophysiological and molecular analysis to evaluate this new model's throughput capacity. Finally, the validation of these bioelectronic platforms will be performed with drugs with known cardiotoxic effects.