Modelling ischemia-reperfusion injury (IRI) in vitro using metabolically matured induced pluripotent stem cell-derived cardiomyocytes

Abstract

Coronary intervention following ST-segment elevation myocardial infarction (STEMI) is the treatment of choice for reducing cardiomyocyte death but paradoxically leads to reperfusion injury. Pharmacological post-conditioning is an attractive approach to minimize Ischemia-Reperfusion Injury (IRI), but candidate drugs identified in IRI animal models have performed poorly in human clinical trials, highlighting the need for a human cell-based model of IRI. In this work, we show that when we imposed sequential hypoxia and reoxygenation episodes [mimicking the ischemia (I) and reperfusion (R) events] to immature human pluripotent stem cell-derived cardiomyocytes (hPSC-CMs), they display significant hypoxia resistance and minimal cell death (∼5%). Metabolic maturation of hPSC-CMs for 8 days substantially increased their sensitivity to changes in oxygen concentration and led to up to ∼30% cell death post-hypoxia and reoxygenation. To mimic the known transient changes in the interstitial tissue microenvironment during an IRI event in vivo , we tested a new in vitro IRI model protocol that required glucose availability and lowering of media pH during the ischemic episode, resulting in a significant increase in cell death in vitro (∼60%). Finally, we confirm that in this new physiologically matched IRI in vitro model, pharmacological post-conditioning reduces reperfusion-induced hPSC-CM cell death by 50%. Our results indicate that in recapitulating key aspects of an in vivo IRI event, our in vitro model can serve as a useful method for the study of IRI and the validation and screening of human specific pharmacological post-conditioning drug candidates.

Human pluripotent stem cell-derived cardiomyocytes as a target platform for paracrine protection by cardiac mesenchymal stromal cells

“…Currently improvement in maturation and specificity is achieved by the use of 3D human engineered heart tissue, mechanical or electrical conditioning, and heart-on-chip technologies although throughput and/or accessibility of these methods remain limiting factors [74][75][76][77][78] . Interestingly, low sensitivity to hypoxia/reoxygenation was previously reported for hPSC-CMs 23,79 , and in our experimental setting, although differences were observed between the lines studied in their sensitivity to menadione, the paracrine protective effect was consistent and comparable. Interestingly, higher susceptibility of hPSC-CM to ischemic damage was linked to a polymorphism of ALDH2 and, unsurprisingly, screening a single commercial line failed to capture the known cardiotoxicity of rosiglitazone 80,81 .…”

Human cardiac progenitor cell activation and regeneration mechanisms: exploring a novel myocardial ischemia/reperfusion in vitro model

“…Such results are in accordance with the findings that point to a relevant role of CM maturation stage on cell survival upon I/R injury. In fact, primary embryonic human CMs are resistant to hypoxia, while primary adult human CMs are highly dependent on an adequate oxygen supply, which might be related with the different metabolic phenotypes between the different developmental stages [32, 33]. hiPSC-CMs with an extra maturation step were therefore used in the following studies, since they better reflected the typical loss of cell viability during AMI.…”

Exercise and Cardioprotection: A Natural Defense Against Lethal Myocardial Ischemia–Reperfusion Injury and Potential Guide to Cardiovascular Prophylaxis

“…In recent years, human induced pluripotent stem cells-derived cardiomyocytes have been successfully cultured and provide a human cardiomyocytes model in which to research exercise effects. [76][77][78] However, in relation to the ischemia and reperfusion process that occurs during acute MI, the cultured cardiomyocyte model provides a reductionist approach for examining the cellular mechanisms underlying cardioprotection. Exercise preconditioning through hypoxia reoxygenation is one of the mechanisms of exercise-induced cardioprotection.…”