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C.K. Bahler, M.P. Fautsch, T.S. Acott, D.H. Johnson; Factors Influencing Outflow Facility in Cultured Human Anterior Segments . Invest. Ophthalmol. Vis. Sci. 2003;44(13):1156.
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Purpose: To determine why variations in outflow facility occur in cultured human anterior segments. Methods: Cell fragments from monolayer cultured 293 cells were prepared by freeze/thaw (5 minutes on dry ice followed by 5 minutes at 37oC repeated four times) or sonication. Cell fragments from 30,000 (n=4) or 60,000 (n=2) cells were infused into cultured human anterior segments. Human genomic DNA was infused into cultured human anterior segments at concentrations equivalent to 30,000 (n=4), 150,000 (n=4) or 300,000 (n=3) cells. Gelatinase A, B, and stromelysin levels from effluents were measured by zymography and correlated with outflow facility (n=3). Results: Cell fragments decreased outflow facility in a dose-dependent manner: fragments from 30,000 cells (corresponding to a 10% loss of trabecular cells) caused a 27% ± 0.07% (mean ± SEM; p=0.09) decrease in facility; fragments from 60,000 cells decreased facility 36% ± 0.03%. Human genomic DNA also decreased outflow facility in a dose-dependent manner: 30,000 cells: 15% ± 0.05% (p=0.11); 150,000 cells: 39% ± 0.05% (p=0.01); 300,000 cells: 26% ± 0.09%. Most pressures remained elevated for a minimum of 24 hours. Gelatinase A, B, and stromelysin levels did not correlate with facility changes during the culture adaptation period. Conclusions: Cell fragments can cause changes in outflow facility during the culture period. These fragments could come from trabecular cells or from ciliary body cells. Genomic DNA also can decrease outflow facility during culture. An ongoing loss of cells from the trabecular meshwork and ciliary body during culture may shower the meshwork and cause variations in outflow facility.
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