Table 1 Transgenic approaches for human induced pluripotent stem cells skeletal muscle differentiation. The bibliographic research was performed until December 2020 using PubMed and SCOPUS databases. The search queries used for titles and/or abstracts starting from 2006 (year of the first report on iPSCs) were [(induced pluripotent stem cells) AND (skeletal muscle)]; [(skeletal muscle cell) AND (differentiation) AND (induced pluripotent stem cells)]; [(induced pluripotent stem cells) AND (myogenesis) OR (myogenic differentiation)]. Review articles, book chapters, and conference abstracts/papers were not included.
Ref. | iPSC origin and lines | Culture conditions in proliferation | Transgene and overexpression system | Myogenic progenitor derivationa | Terminal differentiationa | Culture type | Myogenic cell selection | Functional tests | In vivo engraftment |
|---|---|---|---|---|---|---|---|---|---|
Darabi et al., 201211 | Fibroblasts (IPRN13.13, IPRN14.57) | Matrigel® coating with mTeSR1 | PAX7 (dox-inducible lentiviral system) | IMDM, 15% FBS, 10% HS, 1% chick embryo extract, 50 µg/mL AsAc, 4.5 mM MTG (11 days); same medium as before with dox 0.75 μg/mL (4 days); sorted for PAX7+ (GFP+); same medium as before with dox 0.75 μg/mL and human FGF2 (5 ng/mL) (2 weeks) Markers GE: T, MYOD1 PE: MRFs, CD29/44, CD56/63/105, CDH15, ITGA7 | logDMEM, 5% HS (7 days) Markers GE: DMD, MYH PE: MYH, MYOG | EBs and then adherent | Progenitor purification (EBs): FACS for iPAX7+ (GFP+) | n/a | Cell type: myogenic progenitors iPAX7+ Endpoint: 2 months Outcome: successful engraftment with fibers human DMD+ and LMNA. Increase in tetanic, absolute, and specific force, but no influence on fatigue tests. No tumor formation after 46 weeks (injected in TA muscles of NSG mice) |
Tedesco et al., 201212 | Fibroblast and myoblasts | iMEFs with KO DMEM; 25% KOSR, 2 mM L-glut, 1 mM Na pyr, 100 IU mL pen, 100 mg/mL strep, 1% NEAA, 0. 2 mM 2-ME, 10 ng/ml human FGF2 | MYOD1 (tamoxifen-inducible lentiviral system) | Generation of mesangioblast-like cells (HIDEMs): Matrigel® coating with α-MEM, pen (100 IU/mL), strep (100 mg/mL), 10% FBS, 1% NEAA, 0.2% 2-ME (14 days); MegaCell DMEM (7 days) Markers GE: n/a PE: CD44, CD13, CD146, PDGFRA, MYOD1 | 4OH-tamoxifen or standard tamoxifen (5 days) Markers GE: MYOD1, MYOG, SGCA PE: MYH, MYOD1 | Adherent | Progenitor purification: FACS for SSEA1- | n/a | Cell type: HIDEMs from LGMD2D iPSCs GFP+ Endpoint: 1 month Outcome: 5–7% cell survival, SGCA+ fibers are present and DPC was reconstituted (fibers SGCB+/SGCG+ ). Increase in tetanic force ex vivo. Repopulation of alkaline phosphatase+ pericyte pool.(injected in TA muscles of Sgcanull/scid/beige mice) |
Tanaka et al., 201313 | Fibroblasts (201B7, 253G1, 253G4) | Collagen I or Matrigel® coating with primate ES medium with 4 ng/mL FGF2 | MYOD1 (dox-inducible PiggyBac transposon-Tet-ON system) | Collagen I or Matrigel® coating; primate ES medium, 10 µM ROCK inhibitor (1 day), added 1 µg/mL dox (1 day); α-MEM, 5% KOSR, 50 mU/L pen/50 mg/L strep, and 100 mM 2-ME (5 days) Markers GE: CKM, MYOD1, MEF2C PE: n/a | DMEM with 5% HS, 50 mU/L pen, 50 mg/L strep, 10 ng/mL IGF-1, 2 mM L-glut and 100 mM 2-ME (7 days) Markers GE: MYOG, DMD, microarray PE: DMD, MYH, ACTA1 | Adherent | FACS for iMYOD1+ (mCherry+) | Contraction upon ESt at day 14 (100 V, 3 ms, 1 Hz) | Cell type: myogenic progenitors (day 6 of differentiation) Endpoint: 28 days Outcome: cells displayed fusion potential and the expression of human DMD and human SYNE1 (injected in TA muscles of NOD/Scid-DMD mice) |
Abujarour et al., 201424 | FTc01-C1 and FTc01-C2 | Matrigel® coating with DMEM/F12, 20% KOSR, 1% NEAA, 2 mM L-glut, 100 mM 2-ME, 10 ng/mL FGF2 | MYOD1 (dox-inducible lentiviral system) | DMEM, 10% FBS, 1 µg/mL dox (4 days) Markers GE: n/a PE: n/a | logDMEM, 5% HS (3 days) Markers GE: n/a PE: MYOD1, MYH, MYOG | Adherent | n/a | n/a | n/a |
Quattrocelli et al., 201514 | Fibroblasts and mesangioblasts | iMEFs with DMEM/F12 with 20% KOSR, 1% pen/ strep, 1% L-glut, 1% NEAA, 0.2% 2-ME, and 5 ng/mL FGF2 | PAX3/7 (transient overexpression by pSPORT6.1 plasmid) | DMEM, 2% HS, 1% ITS, 100 ng/mL noggin, 1 mM TGF-β inhibitor (10 days) Markers GE: T, MEOX1, TBX5, PAX3/7, KDR PE: MYH2 | n/a | Adherent | PDGFRA/B+CD44+ of the EBs before differentiation | n/a | Cell type: iPAX3/7 cells PDGFRA/B+CD44+ Endpoint: 2 months Outcome: muscle fiber regeneration with SGCB expression in Sgcbnull/Rag2null/ γcnull mice and DMD expression in dystrophic mouse |
Shoji et al., 201525 | Fibroblasts | iMEFs with primate ES medium with 4 ng/mL FGF2 | MYOD1 (dox-inducible PiggyBac transposon-Tet-ON system) | Matrigel® or collagen I coating, 20% KOSR replacement media, 100 μ g/mL neomycin sulphate (1 day); 20% KOSR replacement media, 1 μg/mL dox (1 day); 5% KOSR/α-MEM media, dox, 2-ME (5 days) Markers GE: MYOD1, MYOG, DMD, CKM, TPM2 PE: n/a | DMEM, 2% HS (7 days) Markers GE: DMD, TPM2 PE: ACTA1, CKM, MYH, ultrastructure | Adherent | Myogenic progenitors: FACS for iMYOD1+(mCherry+) | Detection of Ca2+ influx upon ESt (12 V, 50 ms, 0.2 Hz) (day 9 of differentiation) | n/a |
Lenzi et al., 201626 | Fibroblasts | hESC-qualified Matrigel® coating with Nutristem-XF | MYOD1 (dox-inducible PiggyBac transposon-Tet-ON system) | DMEM/F12 with GlutaMAXTM, 20% KOSR, 1X NEAA, 100 U/mL pen, 100 μg/mL strep, 0,1 mM 2-ME, (5 days); 200 ng/mL dox (2 days) Markers GE: T PE: n/a | Skeletal Muscle Cell Differentiation Medium (Promocell), 100 U/mL pen, 100 μg/mL strep, 200 ng/mL dox (5 days) Markers GE: DMD, MYOD1, MYOG, MYH, MEF2C PE: MYH2, MYOG | Adherent | n/a | Patch-clump recordings of ACh-evoked currents, and intracellular Ca2+ release with ACh stimulations | n/a |
Uchimura et al., 201751 | 414C2, 409B2 | Matrigel® coating with StemFit AK02N, 0.5% pen/strep | MYOD1 (tetracycline-inducible system) | StemFit AK02N, 0.5% pen/strep (1 day); primate ES Cell media, 0.5% pen/strep (1 day); same medium as before, 1 μg/mL dox (1-2 days); α-MEM, 0.5% pen/strep, 5% KSR, 200 μM 2-ME, 1 μg/mL dox (6–7 days) Markers GE: MYOD1, MYOG, MYH PE: MYH2, MYOG | DMEM, 0.5% pen/strep, 2 mM L-glut, 200 μM 2-ME, 5% HS, 10 ng/mL IGF-1, 5 μM SB431542 (2-3 days); DMEM, 0.5% pen/strep, 2 mM L-glut, 200 μM 2-ME, 2% HS, 10 ng/mL IGF-1, 5 μM SB431542 (7 days) Markers GE: n/a PE: MYH2 | Adherent | n/a | n/a | n/a |
Sato et al., 2016204 | n/d | iMEFs with DMEM/F12, 20% KOSR, 1% GlutaMAXTM, 0.01% 2-ME, NEAA, FGF2 | MYOD1 constitutive expression (lentiviral system with EEF1A1 promoter) | Collagen I coating, α-MEM, 5% KOSR (7–10 days) Markers GE: PAX7/3, MYOD1, PE: n/a | Markers GE: MYH2, Ach receptor PE: MYH1, ultrastructure | Adherent | n/a | n/a | n/a |
Rao et al., 201815 | H9, TRiPS, GM2525646, and Fucci | Matrigel® coating with E8 medium | PAX7 (dox-inducible lentiviral system) | Matrigel® coating, with E6 media, 10 µM CHIR99021 (2 days); E6 media, 1 µg/mL dox (18 days) Markers GE: MYHs PE: MYOD1, PAX3, MYF5, MYOG, sarcomeric α-actinin, PAX7 | logDMEM, 10% FBS, Fetuin (500X), hEGF (1000X), DE (1000X), pen (100 unit/mL), strep (50 µg/mL) until 80% confluence (for 2D culture) or 4 days (for 3D culture); logDMEM, N-2 Supplement (100X), pen G (100 unit/mL) (~ 2 weeks) Markers GE: n/a PE: sarcomeric α-actinin, ACh receptor, MYH, Ca2+ handling genes | Adherent vs 3D | Myogenic progenitors: FACS for iPAX7+ (GFP+) | ESt at 20% stretch (40 V/cm, 10 ms): twitch force per cross-sectional area of 0.8 mN/mm2 Ca2+ transients recording after 1, 2, 4 weeks (in vitro) upon ESt (10 ms pulse, 3 V/mm), and from muscle explants 7-15 days post-implantation | Cell type: progenitors (day 14 of differentiation) Endpoint: 15 days Outcome: the bundles were vascularized and remained functional after explant (injected dorsally or in TA muscle of NSG or nude mice) |
Selvaraj et al., 201946 | Fibroblasts (PLZ, TC-1133, MNP-120, MNP-119) | Matrigel® coating with mTeSR1 | PAX7 (dox-inducible lentiviral system) | IMDM basal medium, 15% FBS, 10% HS, 1% pen/strep, 1% GlutaMAXTM, 1% KOSR, 50 µg/mL AsAc, 4.5 mM MTG (altogether defined “myogenic medium”), 10 µM CHIR99021 (2 days); myogenic medium, 200 nM LDN193189, 10 µM (SB431542 (1 day); myogenic medium, 1 µg/mL dox (3 days); gelatin coating, myogenic medium, 1 µg/mL dox, 5 ng/mL FGF2 (4 days) Markers GE: MEOX1, TCF15, PAX3, FOXC2 PE: n/a | KO DMEM, 20% KOSR, 1% NEAA, 1% GlutaMAXTM, 1% pen/strep, 10 µM SB431542, 10 µM DAPT, 10 µM DE, 10 µM PD0325901, 10 µM Forskolin (5 days) Markers GE: MYOG, MYOD1, MYH2, MYH3, MYH8, MYH7 PE: MYH8, TTN, DES | EBs then adherent | Myogenic progenitors: FACS for iPAX7+ (GFP+) | 3D construct with bovine fibrinogen, thrombin, and growth factor reduced Matrigel® 3D. ESt at 20% stretch (10 ms, 0.5 Hz, or 20 Hz): twitch force of 0.4 mN. | n/a |
