Tesr e8 medium

For the generation of iPSC-CFs a protocol developed by Zhang et al. was used^{13 (link)}. Briefly, human iPSCs were dissociated with 1 mL/well 0.5 µM EDTA solution (Invitrogen) at RT for 5 min and seeded on Vitronectin XF (StemCell Technologies) coated 6-well plates at a density of 15.000–30.000 cells/cm² in TeSR-E8 medium (StemCell Technologies) supplemented with 5 μM ROCK inhibitor (Y-27632) (Tocris) for 24 h. Cells were cultured for 6–7 days in TeSR-E8 medium with medium changes every other day until they reached 100% confluency and differentiation started (day 0). At day 0, the medium was changed to 2.5 mL/well RPMI + B27 without insulin (Gibco) and supplemented with 12 µM CHIR99021 (Tocris) for 24 h (day 1). After day 1, the medium was changed to 2.5 mL RPMI + B27 without insulin for 24 h (day 2). Afterwards, the medium was changed to 2.5 mL/well of the CFBM medium (Table S1) supplemented with 75 ng/mL bFGF (StemCell Technologies). Cells were refreshed with 2 mL/well CFBM supplemented with 75 ng/mL bFGF every other day until day 20 when RNA was collected, and cells were dissociated using TrypLE Select (10x) (Thermo Fisher) for 10 min at 37 °C. After dissociation, cells were cultured in DMEM + 10% Fetal bovine serum. For the first two passages, 5 μM ROCK inhibitor was added for 24 h to help cell attachment. Cells between passage 3–6 were used for experiments.

Human H9 (WA09) embryonic stem cells purchased from WiCell Research Institute, Inc hESCs were cultured on Matrigel (Thermo Fisher Scientific)‐coated culture dishes in the TeSR^™‐E8^™ medium (STEMCELL Technologies). Cells were passaged every 5 days by treating with 0.5 mmol/L EDTA (Thermo Fisher Scientific) for 3 minutes at 37°C and transfering at 1:20 ratio onto fresh Matrigel‐coated plates containing TeSR^™‐E8^™ medium with 10 μmol/L Y‐27632 (Tocris). Next day, we changed the medium with fresh TeSR^™‐E8^™ medium without Y‐27632.