Directed Myogenic Differentiation of Human Induced Pluripotent Stem Cells

Abstract

Patient-derived induced pluripotent stem cells (iPSCs) have opened the door to recreating pathological conditions in vitro using differentiation into diseased cells corresponding to each target tissue. Yet for muscular diseases, a method for reproducible and efficient myogenic differentiation from human iPSCs is required for in vitro modeling. Here, we introduce a myogenic differentiation protocol mediated by inducible transcription factor expression that reproducibly and efficiently drives human iPSCs into myocytes. Delivering a tetracycline-inducible, myogenic differentiation 1 (MYOD1) piggyBac (PB) vector to human iPSCs enables the derivation of iPSCs that undergo uniform myogenic differentiation in a short period of time. This differentiation protocol yields a homogenous skeletal muscle cell population, reproducibly reaching efficiencies as high as 70-90 %. MYOD1-induced myocytes demonstrate characteristics of mature myocytes such as cell fusion and cell twitching in response to electric stimulation within 14 days of differentiation. This differentiation protocol can be applied widely in various types of patient-derived human iPSCs and has great prospects in disease modeling particularly with inherited diseases that require studies of early pathogenesis and drug screening.

Key transcription factors in the differentiation of mesenchymal stem cells

“…MyoD overexpression was found to inhibit Twist-1 through miR-206 induction, resulting in an increase in muscle cell differentiation (Koutalianos et al, 2015). Overexpression of MyoD1, using a vector to human-induced pluripotent stem cells, promotes these cells to undergo myogenic differentiation (Shoji et al, 2015). The overexpression of TAZ, which is known as a modulator of osteogenic and adipogenic differentiation of MSCs, was found to increase MyoD-mediated myogenic differentiation.…”

Differentiation of control and ALS mutant human iPSCs into functional skeletal muscle cells, a tool for the study of neuromuscolar diseases

“…Transduced iPSCs represent a stable and expandable population, in which doxycycline treatment and medium switch are sufficient to induce myogenic differentiation in few days. Even if we do not provide comparative analysis, a survey in the literature suggests similar timing and efficiency of differentiation with our system compared to analogous methods (Tanaka et al, 2013, Shoji et al, 2016). …”

“…It has been previously shown that inducible expression of a MyoD transgene carried by a piggyBac vector could induce iPSC muscle differentiation (Tanaka et al, 2013, Shoji et al, 2016). The enhanced version of piggyBac (epB) carries an engineered terminal repeat, which increases transposition efficiency in human pluripotent stem cells (Lacoste et al, 2009, Rosa and Brivanlou, 2011, Rosa et al, 2014).…”

Augmentation of Musculoskeletal Regeneration: Role for Pluripotent Stem Cells

“…Myogenic induction of ESCs and iPSCs has also been shown by inducing expression of Met Activating Genetically Improved Chimeric Factor-1 [112]. The myogenic regulatory factor Myo-D has also proved to be useful for the induction of myogenesis in iPSCs [113]. Here, transplantation of the differentiated cells ameliorated the dystrophic phenotype, restored depleted progenitors and improved functional capacity of the muscle.…”