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I. Jalbert, R. Augusteyn, F. Stapleton; The Effect of Atmospheric Composition and Hydrostatic Pressure on Keratocyte Viability: Preliminary Experiments . Invest. Ophthalmol. Vis. Sci. 2005;46(13):2190.
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Purpose: The aim of this study was to investigate the short–term effects of anoxia and increased pressure on keratocyte viability in tissue culture. Methods: Primary cultures of bovine and porcine keratocytes were exposed to anoxia using a humidified chamber filled with 100% nitrogen. Hydrostatic heads of tissue culture medium equivalent to pressures of 400 and 1,000 Pa were also established. Negative controls were exposed to air and positive controls to D–sorbitol (Sigma–Aldrich, Sydney, Australia). Test conditions were applied for six hours and keratocyte viability assessed immediately afterwards using vital staining (Hoeschst 33258, annexin V–EGFP, propidium iodide). Results: No viable keratocytes could be detected following six hours of anoxia. The percentage of viable cells reduced from 95% during normal atmospheric conditions to 58% and 41% after 6 hours exposure to 400 and 1,000 Pa of hydrostatic pressure, respectively. Raised hydrostatic pressures induced a combination of necrosis and apoptosis in keratocyte cultures. Conclusions: Oxygen deprivation and physiologically relevant levels of pressures are able to directly impact keratocyte survival in primary culture. These findings may underpin some of the recent clinical observations described in the stroma of soft contact lens wearers.
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