Purpose: This study used high-resolution research confocal laser scanning microscopy to study the epithelium of fresh bovine corneas treated with two contact lens solutions.
Methods: Fresh bovine corneal buttons were stained with a combination of the cell-permeable mitochondrial- and nuclear-specific dyes rhodamine 123 and Hoechst 33342 or acridine orange and imaged with a confocal laser scanning microscope. High-resolution X,Y images of the corneal epithelium were taken with a Zeiss 510 confocal laser scanning microscope at a 0.44-062 -microm Z step, and three-dimensional images of the entire thickness of the corneal epithelium were reconstructed. Semiautomated quantitative MatLab software analysis was applied to count the mitochondria in each optical slice. The effects of treatment for 30 minutes with OPTI-FREE Express and ReNu MultiPlus No Rub were investigated and compared to Hanks' balanced salt solution controls.
Results: This study showed the feasibility of using the confocal laser scanning microscope with fluorescent dyes to study organelles throughout the epithelial layers of fresh unfixed bovine corneas. The mitochondria in the intermediate and basal epithelium are abundant and range in length from less than 1 microm to threads greater than 15 microm, with a tendency to accumulate around the nuclei. Image analysis indicated significantly fewer mitochondria only in the superficial epithelium after treatment with ReNu MultiPlus No Rub treatment. OPTI-FREE Express treatment produced significantly fewer mitochondria not only in the superficial but also in the intermediate epithelial layers.
Conclusions: OPTI-FREE Express has the potential for deeper mitochondrial effects compared to ReNu MultiPlus No Rub and Hanks' balanced salt solution controls.
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