Cardiomyocyte Differentiation from hiPSCs

hiPSCs were seeded into 10-cm tissue culture plates and then allowed to grow for ~4 days until the cells became ~70% confluent. CMs were differentiated from iPSCs as previously described (79 (link)) [protocol originally adapted from Lian et al. (35 (link))]. Media were changed to RPMI 1640 + B27 + ascorbic acid (RPMI/B27/AA) containing the GSK3 inhibitor CHIR99021 (Selleck Chemicals) (6 μM) on day 0 of CM differentiation. On day 2 of differentiation, media were changed to RPMI/B27/AA containing the Wnt antagonist IWP-4 (Reprocell) (5 μM). On day 4 of differentiation, media were changed to fresh RPMI/B27/AA, and on day 7 of differentiation, basal media were changed to RPMI/B27 + insulin (10 μg/ml), with media subsequently changed every 2 to 3 days with RPMI/B27/insulin. Beating of CMs was usually observed between days 7 and 9 of differentiation. CMs were dissociated using the STEMdiff Cardiomyocyte Dissociation Kit (StemCell Technologies) and replated onto Geltrex-coated plates for further assay.

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Lin Z., Garbern J.C., Liu R., Li Q., Mancheño Juncosa E., Elwell H.L., Sokol M., Aoyama J., Deumer U.S., Hsiao E., Sheng H., Lee R.T, & Liu J. (2023). Tissue-embedded stretchable nanoelectronics reveal endothelial cell–mediated electrical maturation of human 3D cardiac microtissues. Science Advances, 9(10), eade8513.

Publication 2023

CHIR99021 IWP-4 STEMdiff Cardiomyocyte Dissociation Kit

Ascorbic acid Assay Cardiomyocyte Cells Chir99021 Gsk3 Hipscs Insulin Ipscs Lian Stemcell Tissue

Corresponding Organization : Brigham and Women's Hospital

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Variable analysis

independent variables

Culture duration (4 days)
CHIR99021 concentration (6 μM)
IWP-4 concentration (5 μM)
Insulin concentration (10 μg/ml)

dependent variables

Cardiomyocyte (CM) differentiation efficiency
Beating of CMs (observed between days 7 and 9 of differentiation)

control variables

Cell type (hiPSCs)
Culture vessel (10-cm tissue culture plates)
Culture medium (RPMI 1640 + B27 + ascorbic acid)
Dissociation and replating method (STEMdiff Cardiomyocyte Dissociation Kit and Geltrex-coated plates)

controls

Positive control: Protocol for CM differentiation from iPSCs previously described in the referenced publication (link), which was originally adapted from the protocol by Lian et al. (link)

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