Mesenpro growth supplement

Commercially available BM‐MSCs purchased from Lonza (Verviers, Belgium) were cultured in MesenPro RS Basal Medium (Life Technologies, Inc.) supplemented with 2% MesenPro Growth Supplement (Life Technologies, Inc.) and 1% GlutaMax Supplement (Life Technologies, Inc.) according to the manufacturer's instruction 30. iPS‐MSCs were generated and maintained according to our established protocols as described in our previous studies 31, 32, 33. The iPS‐MSC clone, N1‐iPS‐MSC, was used in our study. This clone was generated from human iPS cells reprogrammed from human fibroblast cells (American Type Culture Collection catalog no. CCL‐186) via viral transduction with human cDNAs of Klf4, Sox2, Oct4, and c‐Myc, as described in our previous studies 18, 31, 32. iPS‐MSCs were derived from iPSCs according to our established clinically compliant protocol 33, 34. iPS‐MSCs within the ninth passages were used throughout this study 18.

The HEK293 cell line (CRL-1573, ATCC) was maintained under standard conditions in DMEM (Lonza) supplemented with 10% fetal bovine serum (FBS), 1% GlutaMAX, and 10 mg/mL penicillin-streptomycin (all from Life Technologies). Umbilical cord blood-derived hMSCs (PCS-500-010, ATCC) were cultured in MesenPRO RS medium supplemented with MesenPRO growth supplement (Life Technologies). Cells were cultured at 37°C in a humidified atmosphere with 5% CO₂ and 20% O₂. hiPSCs were maintained under feeder-free conditions on Matrigel (BD Biosciences) in mTeSR1 medium (STEMCELL Technologies). Cultures were dispersed to single-cell suspensions using Accutase (STEMCELL). Cell survival before nucleofection was promoted with the Rho-associated kinase (ROCK) inhibitor Y-27632 (10 μM; Selleckchem). hiPSCs were generated in P.M.'s laboratory from human CD34⁺ cells from peripheral blood mononuclear cells using OKSM polycistronic SeV vectors (Muñoz-López et al., 2016 ). hiPSC research was approved by the ISCIII Steering Committee on pluripotent stem cell research.