[Kinetic study of cell proliferation in a new wound healing model using tissue cultured corneal endothelial cells]

We studied cell kinetics in the wound healing process using cultured bovine corneal endothelium. We cultured bovine endothelial cells on coverslips coated by hydroxyethyl methacrylate (HEMA) which had on 8 mm HEMA-free zone in diameter, and produced wounds at the center of the monolayer cell sheets using a rotating silicone tip. At various time periods after injury, we added bromodeoxyuridine (BrdU) to the medium and incubated for 12 hours. Then, the specimens with fixed 10% phosphate-buffered formalin and were incubated with a monoclonal antibody against BrdU. The cells incorporating BrdU into DNA were stained by the Avidin Biotin Peroxidase Complex (ABC) method. At 12 hours, no labeled cells were observed. At 24 hours, 14.6 +/- 4.0 cells were stained. The maximum labeling occurred during 48-60 hours after the wound. Seventy two hours after the wound, labeled cells decreased rapidly. Labeled cells were localized within 0.6mm from the wound edge throughout the wound healing process. It can be thought that the wound healing process comprised four phases, i.e., latent, migration, migration plus mitosis, and contact inhibition phases. The first is the latent phase observed during the first six hours after wound infliction. The cells respond to external expansion. The second phase in the migration phase which is last until 24 hours after wound infliction, and occurs mainly through cellular migration, while proliferation has only a minor contribution in this phase. The third is the migration plus mitosis phase. The cell proliferation shows a rapid increase after 72 hours after wound infliction.(ABSTRACT TRUNCATED AT 250 WORDS)