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C. Martinez-Garcia, J. Blanco-Mezquita, R. Torres, J. Merayo-LLoves; Keratocites Apoptosis after Laser in Situ Keratomileusis . Invest. Ophthalmol. Vis. Sci. 2003;44(13):893.
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Purpose: Apototosis is a controlled death of cells that occurs in development, homeostasis and wound-healing. Keratocytes apoptosis has been studied as an initiating event in the wound-healing response following refractive surgery, scrape-photorefractive keratectomy (PRK), transepithelial PRK and laser assisted keratomileusis (Lasik). Most of these studies have been carried out in animals without Bowman’s layer and in short periods of time after surgery. This study was undertaken to demonstrate the timing of this controlled death during wound-healing after LASIK surgery. Methods: Laser assisted in situ keratomilesius was performed in adults hens with 48 µ m laser ablation after a 80µ m thick hinged corneal flap had been made. Hens were killed 4, 5, 8, 10, 12, 24 and 48 hours, 3, 4 and 7 days and 1 and 2 months after surgery and globes were fixed suitably. Central corneal was observed by fluorescent microscopy using the cell death marker, tdt-mediated dUTP nick-end labelling TUNEL assay. Cellular morphologic changes were evaluated by an electron microscope examination. Results: Soon after surgery (4 and 5 hours), we observed through fluorescent microscopy, the first positive-TUNEL cells in the area of lamellar cut. In the following hours (6 and 8) the number of apoptotic cells increased, in the same area and we could find some of them in the deep stroma. From ten to twelve hours apoptosis was massive and we could find clusters of death cells between flap and the stromal bed. The number of positive TUNEL cells in healing line after 24, 48 and 72 hours decreased little by little and some of them were situated in deep stroma. These positive cells could be found drawing the line where the flap and stroma were healing after one or two months. Observed by electron microscopy these cells have spindle shape, cell shrinkage, chromatin condensation, membranes integrity and are free, without junctions between them and with extracellular matrix. Apoptotic keratocites, macrophages and apoptotic bodies make up aggregates of cells localised between the flap and stromal bed, but we couldn’t see here inflammatory cells. Conclusions: There is a first wave of apotosis due to laser damage and possibly a late, continuous and slow death necessary in wound-healing remodelling.
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