Development of high content imaging methods for cell death detection in human pluripotent stem cell-derived cardiomyocytes.
Human pluripotent stem cell-derived cardiomyocytes (hPSC-CM) are being investigated as a new source of cardiac cells for drug safety assessment. We developed a novel scalable high content microscopy-based method for the detection of cell death in hPSC-CM that can serve for future predictive in vitro cardio-toxicological screens. Using rat neonatal ventricular cardiomyocytes (RVNC) or hPSC-CM, assays for nuclear remodelling, mitochondrial status, apoptosis and necrosis were designed using a combination of fluorescent dyes and antibodies on an automated microscopy platform. This allowed the observation of a chelerythrine-induced concentration-dependent apoptosis to necrosis switch and time-dependent progression of early apoptotic cells towards a necrotic-like phenotype. Susceptibility of hPSC-CM to chelerythrine-stimulated apoptosis varied with time after differentiation, but at most time points, hPSC-CM were more resistant than RVNC. This simple and scalable humanized high-content assay generates accurate cardiotoxicity profiles that can serve as a base for further assessment of cardioprotective strategies and drug safety.
National Heart and Lung Institute, Imperial College, ICTEM, Hammersmith Campus, Du Cane Rd., London, UK.
Maxime
M
Mioulane
Gabor
G
Foldes
Nadire N
NN
Ali
Michael D
MD
Schneider
Sian E
SE
Harding
D000818:Animals; D000831:Animals, Newborn; D017209:Apoptosis / Q000187:drug effects*; D057205:Automation, Laboratory; D053119:Benzophenanthridines / Q000633:toxicity*; D053470:Caspases, Effector / Q000378:metabolism; D002460:Cell Line; D002463:Cell Membrane Permeability / Q000187:drug effects; D053144:Cell Nucleus Shape / Q000187:drug effects; D058948:Cell Tracking / Q000379:methods*; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation; D057166:High-Throughput Screening Assays / Q000379:methods*; D006801:Humans; D007091:Image Processing, Computer-Assisted; D057026:Induced Pluripotent Stem Cells / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D053078:Membrane Potential, Mitochondrial / Q000187:drug effects; D008856:Microscopy, Fluorescence; D008929:Mitochondria, Heart / Q000187:drug effects / Q000473:pathology; D032383:Myocytes, Cardiac / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D009336:Necrosis; D010641:Phenotype; D051381:Rats; D013997:Time Factors
D053119:Benzophenanthridines; C016299:chelerythrine; D053470:Caspases, Effector