IPSC derived cardiac fibroblasts of DMD patients show compromised actin microfilaments, metabolic shift and pro-fibrotic phenotype

Abstract

Duchenne muscular dystrophy (DMD) is a severe form of muscular dystrophy caused by mutations in the dystrophin gene. We characterized which isoforms of dystrophin were expressed by human induced pluripotent stem cell (hiPSC)-derived cardiac fibroblasts obtained from control and DMD patients. Distinct dystrophin isoforms were observed; however, highest molecular weight isoform was absent in DMD patients carrying exon deletions or mutations in the dystrophin gene. The loss of the full-length dystrophin isoform in hiPSC-derived cardiac fibroblasts from DMD patients resulted in deficient formation of actin microfilaments and a metabolic switch from mitochondrial oxidation to glycolysis. The DMD hiPSC-derived cardiac fibroblasts exhibited a dysregulated mitochondria network and reduced mitochondrial respiration, with enhanced compensatory glycolysis to sustain cellular ATP production. This metabolic remodeling was associated with an exacerbated myofibroblast phenotype and increased fibroblast activation in response to pro fibrotic challenges. As cardiac fibrosis is a critical pathological feature of the DMD heart, the myofibroblast phenotype induced by the absence of dystrophin may contribute to deterioration in cardiac function. Our study highlights the relationship between cytoskeletal dynamics, metabolism of the cell and myofibroblast differentiation and provides a new mechanism by which inactivation of dystrophin in non-cardiomyocyte cells may increase the severity of cardiopathy.

Heart-on-a-chip systems: disease modeling and drug screening applications

“…To create a more holistic in vitro model with multiple cell types, methods for differentiating hiPSCs into other cell types, such as endothelial cells (EC) and cardiac fibroblasts (CF), which are responsible for producing the extracellular matrix (ECM) in vivo , are being developed. 117–119 Strategically incorporating different cell types to more accurately represent the heart in vivo could enable more comprehensive studies of cell–cell communication within the heart. However, the controllable placement and selective differentiation of multiple cell types for in vitro modeling has proven to be challenging.…”

Differentiation of Pluripotent Stem Cells for Disease Modeling: Learning from Heart Development

“…The loss of full-length dystrophin, an actin cytoskeleton component, in DMD iPSC-CFs induced metabolic remodeling, causing increased fibroblast activation after pro-fibrotic stimulation. This study highlights the relationship between cytoskeletal dynamics, metabolism, and myofibroblast differentiation while providing a new mechanism by which the loss of dystrophin in CFs may increase the severity of cardiac fibrosis [ 162 ].…”

Molecular and metabolomic characterization of hiPSC-derived cardiac fibroblasts transitioning to myofibroblasts

“…There are multiple protocols published so far to differentiate cardiac fibroblasts from iPSCs; however, most of them share a similar path with various small molecules to initiate the cardiac mesoderm formation followed by epicardial cell conversion to cardiac fibroblasts. To generate the cardiac mesoderm and cardiac progenitors from iPSCs, most protocols used a GSK3 inhibitor (CHIR99021) and a WNT signaling inhibitor (IWR1 or XAV939); after cardiac mesoderm formation, a TGFβ-inhibitor (SB431542) and retinoic acid (RA) promoted conversion to epicardial lineage cells, followed by FGF-2 treatment to generate hiPSC-CFs (Beauchamp et al, 2020; Campostrini et al, 2021; Cumberland et al, 2023; Giacomelli et al, 2020; Hall et al, 2023; Soussi et al, 2023; Whitehead et al, 2022; Yu et al, 2022; Zhang et al, 2022; Zhang et al, 2019a). The differentiation protocols for iPSC-derived lung and dermal fibroblasts differ in the growth factors and chemical signals used on specific lineages to become tissue-specific fibroblasts (Alvarez-Palomo et al, 2020; Itoh et al, 2013; Kim et al, 2018; Mitchell et al, 2023; Tamai et al, 2022; Wong et al, 2015).…”