Stem cell culture was achieved in line with the ISSCR guidelines (https://www.isscr.org/standards-document ). HiPSC lines used in this study were maintained on ESC-qualified Matrigel (Corning), coated plates with mTeSR1 media (Stem cell technology). All cell lines were cultured at 37°C, under normoxic conditions (20% O2, 5% CO2) and culture media was daily replaced. HiPSC were passaged every 4 days at a 1:6 split ratio using passaging solution XF (Miltenyi) for 2 min, at room temperature for dissociation into cell clusters. Cells were grown for 10 passages maximum. A new vial of the cell bank was used when cell reach P30. The MycoAlert kit (LONZA) was employed to confirm the absence of mycoplasma in all cell lines.
The H1-ESC line was developed in the laboratory of Andras Nagy, of which the cells originated from the H1 embryonic stem cell line HA01 (Wi Cell, USA). The CFS were cultured in 6-well culture plates coated with Geltrex™ LDEV-Free, hESC-Qualified, Reduced Growth Factor Basement Membrane Matrix (Thermofisher Scientific). Cells were maintained in mTeSR™ Plus (Stemcell Technologies) that was refreshed daily (1.5mL) and passaged every 4-5 days using ReLeSr (Stemcell Technologies) as per manufacturers’ instructions.
The H1-ESC line was developed in the laboratory of Andras Nagy, of which the cells originated from the H1 embryonic stem cell line HA01 (Wi Cell, USA). The CFS were cultured in 6-well culture plates coated with Geltrex™ LDEV-Free, hESC-Qualified, Reduced Growth Factor Basement Membrane Matrix (Thermofisher Scientific). Cells were maintained in mTeSR™ Plus (Stemcell Technologies) that was refreshed daily (1.5mL) and passaged every 4-5 days using ReLeSr (Stemcell Technologies) as per manufacturers’ instructions.