Cell Migration Assay with MSCs

The procedure of the cell migration assay was conducted according to a procedure reported in previous research [31 (link)]. The Oris seeding stoppers were cultured with 1 × 10⁴ MSCs per well and incubated for 24 and 48 h to reach confluency. Afterwards, the stoppers were removed, with the exception of one which was used in reference for pre-migration. The seeded plates were incubated at 37 °C for further investigation of pre-migration (t = 0 h) and post-migration (24 to 48 h). Afterwards, 200 μL of Calcein AM (2 μM, Sigma-Aldrich, Burlington, MA, USA) was added to each well and stained for 30 min at each time point, with the migration ability observed using a Zeiss Axio Imager A1 fluorescence microscope (White Plains, NY, USA). The semi-quantification of cell migration distance was acquired through Image J 5.0 software (Becton Dickinson, Canton, MA, USA).