Facsuite software version 10

According to the MSC criteria of the International Society for Cell Therapy (ISCT),⁸⁵ immunophenotypic analysis of WJ‐MSCs was performed by flow cytometry to determine the expression of the following markers: CD44, CD73, CD90, CD105, CD14, CD11b, HLA‐DR (MHC‐II), CD34, CD45, and CD19 (BD Biosciences). At least 10 000 events were acquired on a BD FACSVerse (BD Biosciences, New Jersey), and the results were analyzed with BD FACSuite software version 10 (BD Biosciences). The differentiation of WJ‐MSCs was tested according to the procedure outlined in a previous report.⁸⁶

Lentiviral vectors were produced by the transient transfection of 293T cells, as described previously (16 (link)). The 293FT cells (~5×10⁶ cells) in logarithmic growth phase were inoculated into 10 cm culture dishes and cultured for 24 h in a humidified 5% CO₂ atmosphere at 37°C. The vectors were subsequently transfected into the 293FT cells using Lipofectamine^® 2000 and incubated overnight under the same conditions. The following day, DMEM containing 10% FBS was changed and the viral supernatants were collected following 48 h under the same conditions, filtered using 0.45 µm pore size filters and stored at −80°C. For the determination of infectious titers, 293FT cells were infected with lentivirus (CyPA shRNA and Control shRNA) (dilution, 1:10) and incubated overnight at 37°C with 5% CO₂. The cells were subsequently washed in PBS and cultured for an additional 48 h under the same conditions. GFP-positive cells were counted using a BD FACSVerse™ flow cytometer and BD FACSuite software (version 1.0) (both BD Biosciences, Franklin Lakes, NJ, USA).