Cell Migration and Invasion Assays

Cell migration was assessed by measuring the movement of cells into a scraped, acellular area that was prepared as described previously.^{48 (link)} The spread extent of wound closure was observed after 0 and 48 h, respectively. Migration was quantified by counting the total number of cells that migrated toward the original wound field. For the invasion assay, Matrigel-coated chambers (BD Biosciences, San José, CA, USA) containing 8-μm pores were used. A total of 2 × 10⁵ concentration cells were seeded into the upper chambers (coated in Matrigel) in serum-free medium. The lower chamber of the Transwell was filled with culture media containing 10% FBS as a chemo-attractant. After the chambers were incubated at 37 °C for 48 h, non-invaded cells on the top of the Transwell were scraped off with a cotton swab. Successfully translocated cells were fixed with 10% formalin. Then, they were stained with 0.1% crystal violet for 30 min and counted under a light microscope. All experiments were performed in triplicate.