Methods to produce induced pluripotent stem cell-derived mesenchymal stem cells: Mesenchymal stem cells from induced pluripotent stem cells

Table 2 Protocols to produce induced pluripotent stem cell-derived mesenchymal stem cells by embryoid bodies approaches

Ref.	iPSC origin	iPSC to iMSC protocol	Time	Application	Citations
Ahfeldt et al[56], 2012	Foreskin fibroblast	(1) iPSC cultured into low-adhesion dishes for EB formation with DMEM + 15% FBS + L-Glutamine; (2) EB plated into gelatin-coated dishes with DMEM + 15% FBS + L-Glutamine; and (3) Cells plated with Mensenchymal Progenitor Cell (MCP) medium: DMEM + 15% FBS + L-Glutamine + bFGF	12	Producing white and brown adipocytes from hPSCs	194
Chen et al[57], 20121	Lung fibroblast	SB431542 inhibitor differentiation method (feeder free); iPSC cultured in inhibitor differentiation medium: KOSR medium + SB431542 (TGFβ inhibitor)Without bFGF to enhance differentiation. Embryoid body differentiation method: (1) EB formation in KOSR medium; and (2) EB cultured with MSC medium: DMEM + 10% FCS + L-Glutamine + gentamicin + p/s	17	Generation of iMSC with TGF-beta inhibitor	136
Villa-Diaz et al[58], 2012	Dermal fibroblasts	(1) EB formation in suspension cultured into ultra-low-attachment plates; and (2) EB plated on gelatin-coated dishes with MSC medium: α-MEM + 10% FBS + L-Glutamine + NEAA + FGF2	21	iMSC from iPSC cultured on synthetic substrate (PMEDSAH)	262
Wei et al[59], 20121	Dermal fibroblasts	(1) EB formation through cardiac differentiation protocol involving cardiomyogenic medium CARM: High-Glucose DMEM + L-Glutamine + NEAA + Selenium Transferrin + β-ME + SB 203580 (p38-MAPK inhibitor); and (2) EB plating on gelatin-coated plates with DMEM + 2% FBS	21	Generation of iMSC	64
Shao et al[60], 2013	MSC	(1) iPSC cultivated in suspension in the differentiation medium: KO DMEM + 20% FBS+ 1% NEAA + β-ME+ L-Glutamine for EB formation; and (2) Embryoid bodies plated on gelatin-coated dishes	19	iMSC DNA methylation profiles	48
Jeong et al[46], 20141	NA	(1) iPSCs cultured in iMSC-inducing medium: DMEM/F12 + 20% KOSR + SB431542 (TGFβ inhibitor); (2) EB grown on matrigel + DMEM/F12 + 0.5% BSA + 10% ITS + SB431542 (TGFβ inhibitor); and (3) Outgrowth grown with DMEM/F12 + FBS (10%) + p/s	17	Duchene muscular dystrophy	17
Miao et al[61], 2014	Dermal fibroblasts	EB cultured with DMEM + 10% FBS	NA	Myocardial infarctus	38
Tang et al[21], 2014	bone marrow	(1) iPSC cultured in ultra-low attachment plate to form EB with differentiation medium: DMEM/F12 + 20% KSR + MEF medium + NEAA + L-Glutamine + β-ME; and (2) EB plated into gelatin-coated plates + MSC growth medium: DMEM + 10% FBS + L-Glutamine + p/s	20	iMSC and calcium phosphate scaffold for bone regeneration	88
Sheyn et al[20], 2016	Dermal fibroblasts	(1) iPSC plated into PCR plates to form EB with IMDM medium: MDM media + KOSR + NEAA + β-ME + PSA antifungal-antibacterial solution; (2) EB transferred to poly-HEMA-coated flasks; (3) Attached EB (aiMSCs) and Transferred EB (tiMSCs) cultured into gelatin-coated flask with medium + TGF-β1; and (4) Medium switched for DMEM + 10% FBS + L-Glutamine + p/s	10	Generation of iMSC and repair bone defect	60
Eto et al[55], 20181	Skin fibroblast	(1) iPSCs treated with CTK (collagenase type IV + 0.25% trypsin + KSR) and transferred to petri dishes to form EB with: DMEM/F12 + 20% KOSR + glutamine + NEAA + BMP4 + p/s; (2) Specific Differentiation or Mesodermal Differentiation: EB cultured on collagen-coated plates + αMEM + 10% FBS + bFGF + BMP4 + Activin A + LiCl + p/s; or Neuroepithelial differentiation: αMEM + 10% FBS + β-ME + RA; and (3) FACS: PDGFR-α+ and VEGFR2+ cells resuspended on collagen-coated plates with αMEM + 10% FBS + 20% KOSR	10	iMSC from mesoderm or neuroepithelium differentiation	7
Nachlas et al[51], 20181	NA	(1) iPSC cultured in suspension (to promote cell aggregate) with differentiation media: KO-DMEM + β-ME + L-Glutamine + 20% FBS + NEAA + p/s, then cells were cultured on gelatin coated plates; and (2) Cells cultured with iMSC media: KO-DMEM + L-Glutamine + 10% FBS + NEAA + p/s	12	Generation of valve interstitial-like cells from iMSC	14
Karam et al[62], 2020	PBMC	(1) EB cultured into ultra-low attachment plates with differentiation medium: Low-Glucose DMEM + 15% FBS + p/s; (2) Later, RA is added to enhance EB formation; (3) EB plating into gelatin/matrigel coated plates + differentiation medium; and (4) Later bFGF is added	14	Generation of iMSC and adipocytes	NA
Huang et al[63], 2020	PBMC	(1) iPSC cultured in suspension to form EB; and (2) Cells plated into gelatin-coated plates with α-MEM + FGF2	NA	Repair of acute kidney injury	NA

iPSC origin refers to the cell type used for reprograming; Time is indicated as the minimum number of days required to obtain iMSCs; Citations show numbers on March 2020.

¹In references indicate the study includes methods in more than one protocol category. bFGF: Basic fibroblast growth factor; β-ME: β-Mercaptoethanol; BMP4: Bone morphogenetic protein 4; BSA: Bovine serum albumin; CDM: Chemical defined medium; DMEM/F12: Dulbecco's modified Eagle’s medium F12; EB: Embryoid body; EGF: Epidermal growth factor; FBS: Fetal bovine serum; FCS: Fetal calf serum; FGF2: Fibroblast growth factor 2; HEPES: 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid; hiFBS: Heat inactivated fetal bovine serum; hPL: human platelet lysate; IMDM: Iscove's modified Dulbecco's media; iMSCs: Induced pluripotent stem cell-derived mesenchymal stem cells; iPSC: Induced pluripotent stem cell; ITS: Insulin-transferrin-selenium; KO-DMEM: Knockout Dulbecco's modified Eagle’s medium; KOSR: Knock-out serum replacement; KSR: Knockout serum replacement; α-MEM: Minimum essential medium Eagle; NA: Not available; PBMC: Peripheral blood mononuclear cell; p/s: Penicillin-streptomycin; SRM: Serum replacement medium; TGF: Transforming growth factor; VEGFR: Vascular endothelial growth factor.