Antimigratory Effect of Bac I and II on HT-29 Cells

Cellular migration was measured to assess antimigratory effect of bac I and II either alone or in combination on HT-29 by circular scratch wound migration assay as described previously [13 (link),57 (link)]. Briefly, HT-29 cells were seeded in a complete medium at 1 × 10⁵ cells per well on 96-well flat-bottomed plates and incubated under the standard culture condition until 80% confluent. The medium was replaced with a serum-reduced medium and cells were incubated overnight. A circular wound was made on the cellular monolayer using a 10 µL pipette tip. The serum-reduced medium was changed to a complete medium, supplemented either with vehicle or bac I and/or II, and, to inhibit cell proliferation, 1 μg/mL mitomycin C (Sigma-Aldrich, St. Louis, MO, USA). Images were captured at time 0 and after 48 h on an Eclipse TE2000-U light microscope (Nikon, Tokyo, Japan) at 40× magnification. NIS-Elements BR software (Nikon) was used to measure the area of the wound relative to the area at time 0.