Bioreactor-based 3D human myocardial ischemia/reperfusion in vitro model: a novel tool to unveil key paracrine factors upon acute myocardial infarction

During acute myocardial infarction (AMI), Ischemia/Reperfusion (I/R) injury causes cardiomyocyte (CM) death and loss of tissue function, making AMI one of the major causes of death worldwide. Cell-based in vitro models of I/R injury have been increasingly used as a complementary approach to preclinical research. However, most approaches use murine cells in 2D culture setups, which are not able to recapitulate human cellular physiology, as well as nutrient and gas gradients occurring in the myocardium.In this work we established a novel human in vitro model of myocardial I/R injury using CMs derived from human induced pluripotent stem cells (hiPSC-CMs), which were cultured as 3D aggregates in stirred tank bioreactors. We were able to recapitulate important hallmarks of AMI, including loss of CM viability with disruption of cellular ultrastructure, increased angiogenic potential, and secretion of key proangiogenic and proinflammatory cytokines. Conditioned medium was further used to probe human cardiac progenitor cells (hCPCs) response to paracrine cues from injured hiPSC-CMs through quantitative whole proteome analysis (SWATH-MS). I/R injury hiPSC-CM conditioned media incubation caused upregulation of hCPC proteins associated with migration, proliferation, paracrine signaling, and stress response-related pathways, when compared to the control media incubation.Our results indicate that the model developed herein can serve as a novel tool to interrogate mechanisms of action of human cardiac populations upon AMI. During acute myocardial infarction (AMI), Ischemia/Reperfusion (I/R) injury causes cardiomyocyte (CM) death and loss of tissue function, making AMI one of the major causes of death worldwide. Cell-based in vitro models of I/R injury have been increasingly used as a complementary approach to preclinical research. However, most approaches use murine cells in 2D culture setups, which are not able to recapitulate human cellular physiology, as well as nutrient and gas gradients occurring in the myocardium. In this work we established a novel human in vitro model of myocardial I/R injury using CMs derived from human induced pluripotent stem cells (hiPSC-CMs), which were cultured as 3D aggregates in stirred tank bioreactors. We were able to recapitulate important hallmarks of AMI, including loss of CM viability with disruption of cellular ultrastructure, increased angiogenic potential, and secretion of key proangiogenic and proinflammatory cytokines. Conditioned medium was further used to probe human cardiac progenitor cells (hCPCs) response to paracrine cues from injured hiPSC-CMs through quantitative whole proteome analysis (SWATH-MS). I/R injury hiPSC-CM conditioned media incubation caused upregulation of hCPC proteins associated with migration, proliferation, paracrine signaling, and stress response-related pathways, when compared to the control media incubation. Our results indicate that the model developed herein can serve as a novel tool to interrogate mechanisms of action of human cardiac populations upon AMI. Brief Commentary Maria J. Sebastião et al.BackgroundAcute Myocardial infarction (AMI) and inherent Ischemia/ Reperfusion (I/R) injury remains a leading cause of death worldwide. I/R research has relied on animal models and murine cell cultures that despite central to cardiovascular research, do not fully recapitulate human cellular physiology.Translational SignificanceIn this work we developed and characterized a 3D human cellular in vitro I/R model. This system represents a step closer to the human cardiomyocyte milieu and a novel tool to interrogate human cardiac cell populations upon AMI. In order to demonstrate the relevance of the 3D bioreactor I/R model developed to interrogate AMI molecular mechanisms of action, we analyzed human cardiac progenitor cells’ response to paracrine cues from injured human cardiomyocytes. Acute Myocardial infarction (AMI) and inherent Ischemia/ Reperfusion (I/R) injury remains a leading cause of death worldwide. I/R research has relied on animal models and murine cell cultures that despite central to cardiovascular research, do not fully recapitulate human cellular physiology. In this work we developed and characterized a 3D human cellular in vitro I/R model. This system represents a step closer to the human cardiomyocyte milieu and a novel tool to interrogate human cardiac cell populations upon AMI. In order to demonstrate the relevance of the 3D bioreactor I/R model developed to interrogate AMI molecular mechanisms of action, we analyzed human cardiac progenitor cells’ response to paracrine cues from injured human cardiomyocytes.