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A self-renewing biomimetic skeletal muscle construct engineered using induced myogenic progenitor cells

ABSTRACT: A self-renewing biomimetic skeletal muscle construct engineered using induced myogenic progenitor cells

PROVIDER: PRJNA918395 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR22972108.fastq.gz	Fastqsanger.gz
	SRR22972109.fastq.gz	Fastqsanger.gz
	SRR22972110.fastq.gz	Fastqsanger.gz
	SRR22972111.fastq.gz	Fastqsanger.gz

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A self-renewing biomimetic skeletal muscle construct engineered using induced myogenic progenitor cells

Project description:Skeletal muscle is a highly organized and regenerative tissue that maintains its homeostasis primarily by activation and differentiation of muscle stem cells. Mimicking an in vitro skeletal muscle differentiation program that contains self-renewing adult muscle stem cells and aligned myotubes has been challenging. Here, we set out to engineer a biomimetic skeletal muscle construct that can self-regenerate and produce aligned myotubes using induced myogenic progenitor cells (iMPCs), a heterogeneous culture consisting of skeletal muscle stem, progenitor and differentiated cells. Utilizing electrospinning, we fabricated polycaprolactone (PCL) substrates that enabled iMPC-differentiation into aligned myotubes by controlling PCL fiber orientation. Newly-conceived constructs contained highly organized multinucleated myotubes in conjunction with self-renewing muscle stem cells, whose differentiation capacity was augmented by Matrigel supplementation. Additionally, we demonstrate using single cell RNA sequencing (scRNA-seq) that iMPC-derived constructs faithfully recapitulate a step-wise myogenic differentiation program. Notably, when the constructs were subjected to a damaging myonecrotic agent, self-renewing muscle stem cells rapidly differentiated into aligned myotubes, akin to skeletal muscle repair in vivo. Taken together, we report on a novel in vitro system that mirrors myogenic regeneration and muscle fiber alignment, and can serve as a platform to study myogenesis, model muscular dystrophies or perform drug screens.

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Project description:Fate decisions in the embryo are controlled by a plethora of microenvironmental interactions in a three-dimensional niche. To investigate whether aspects of this microenvironmental complexity can be engineered to direct myogenic human induced pluripotent stem cell (hiPSC) differentiation, we screened cell types present in the developmental or adult stem cell niche in heterotypic suspension embryoids. We identified embryonic endothelial cells and fibroblasts as highly permissive for myogenic specification of hiPSCs. After two weeks of sequential Wnt and FGF pathway induction, these three-component embryoids (iTCEs) are enriched in Pax7 positive embryonic-like myogenic progenitors (eMPs) that can be isolated by flow cytometry. Myogenic differentiation of hiPSCs in iTCEs relies on a specialized structural microenvironment and depends on MAPK, PI3K/AKT, and Notch signaling. After transplantation in a mouse model of Duchenne muscular dystrophy, eMPs repopulate the stem cell niche, reactivate after repeated injury and, compared to adult human myoblasts, display enhanced fusion and lead to stronger muscles. Altogether, we provide a two-week protocol for efficient and scalable suspension-based 3D derivation of Pax7 positive myogenic progenitors from hiPSCs.

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