Electrophysiological abnormalities in induced pluripotent stem cell‐derived cardiomyocytes generated from Duchenne muscular dystrophy patients

Abstract

Duchenne muscular dystrophy (DMD) is an X‐linked progressive muscle degenerative disease, caused by mutations in the dystrophin gene and resulting in death because of respiratory or cardiac failure. To investigate the cardiac cellular manifestation of DMD, we generated induced pluripotent stem cells (iPSCs) and iPSC‐derived cardiomyocytes (iPSC‐CMs) from two DMD patients: a male and female manifesting heterozygous carrier. Dystrophin mRNA and protein expression were analysed by qRT‐PCR, RNAseq, Western blot and immunofluorescence staining. For comprehensive electrophysiological analysis, current and voltage clamp were used to record transmembrane action potentials and ion currents, respectively. Microelectrode array was used to record extracellular electrograms. X‐inactive specific transcript (XIST) and dystrophin expression analyses revealed that female iPSCs underwent X chromosome reactivation (XCR) or erosion of X chromosome inactivation, which was maintained in female iPSC‐CMs displaying mixed X chromosome expression of wild type (WT) and mutated alleles. Both DMD female and male iPSC‐CMs presented low spontaneous firing rate, arrhythmias and prolonged action potential duration. DMD female iPSC‐CMs displayed increased beat rate variability (BRV). DMD male iPSC‐CMs manifested decreased I f density, and DMD female and male iPSC‐CMs showed increased I Ca,L density. Our findings demonstrate cellular mechanisms underlying electrophysiological abnormalities and cardiac arrhythmias in DMD.

DMD Pluripotent Stem Cell Derived Cardiac Cells Recapitulate in vitro Human Cardiac Pathophysiology

“…Such observations remain to be validated in human DMD. Furthermore, to this day, there are only limited studies of human DMD cardiac cell (CC) models, describing some of the discrepancies eventually leading to DMD-CM damage (Eisen et al, 2019;Pioner et al, 2020). In the present study, we thus focused, for the first time, on evaluating the impact of dystrophindeficiency on some molecular properties of the excitationcontraction coupling (ECC) and flight-or-fight response in some patient-specific DMD human pluripotent stem cell-derived cardiac cells (DMD-hPSC-CCs) as well as in CRISPR/Cas9 engineered hPSC-CCs (summarized in Graphical abstract).…”

Drug Development and the Use of Induced Pluripotent Stem Cell-Derived Cardiomyocytes for Disease Modeling and Drug Toxicity Screening

“…These prominent scientists discovered that by inserting four transcription factors (Oct3/4, Sox2, Klf4 and c-Myc) into fully-differentiated somatic cells, they are transformed/reprogrammed into iPSCs, which feature the two key properties of stem cells: they can divide indefinitely and are pluripotent. Once iPSCs are generated, they can give rise to practically any cell type, such as cardiomyocytes [ 87 , 88 , 89 , 90 , 91 ], neurons [ 92 , 93 , 94 ], pancreatic cells [ 95 , 96 ] and skeletal muscle cells [ 97 , 98 ]. The clinical implications of iPSCs are beyond imagination, since iPSCs can differentiate into a broad range of cell types which can be used for cell therapy, and tissue replacement and regeneration.…”

“…Further, the membrane sealant Poloxamer 188 attenuated the increase in [Ca 2+ ] i and suppressed apoptosis in the mutated cells by repressing caspase-3 activation [ 236 ]. In a recent study from Binah’s lab [ 91 ], we investigated iPSC-CMs from one male and one female DMD patients. Although DMD is an X-linked disorder, in some cases female patients exhibit clinical abnormalities [ 237 ].…”

“…iPSC-CMs from the male patient exhibited decreased amplitude of the key pacemaker current I f , while iPSC-CMs from the female DMD patient presented with increased beat rate variability. In addition, both female- and male-derived iPSC-CMs showed increased I Ca,L density [ 91 ].…”

“Betwixt Mine Eye and Heart a League Is Took”: The Progress of Induced Pluripotent Stem-Cell-Based Models of Dystrophin-Associated Cardiomyopathy

“…In the last few years, iPSC-CMs have been widely used to model cardiac diseases including DAC, and more than 20 original articles reported the use of iPSC-derived cells from MD patients. Starting from the first iPSC line derived from skin fibroblasts of a DMD patient in 2008 [ 341 ], numerous other iPSC cell lines derived from different somatic-cell sources obtained from DMD and BMD patients were generated and published [ 133 , 253 , 302 , 342 , 343 , 344 , 345 , 346 , 347 , 348 , 349 , 350 , 351 , 352 , 353 , 354 , 355 , 356 , 357 , 358 , 359 , 360 , 361 , 362 , 363 , 364 , 365 , 366 , 367 , 368 ]. These works demonstrated that it is possible to use iPSC-derived cells carrying specific DMD mutations to reproduce MD pathogenesis and to verify new therapeutic approaches.…”