Clinically relevant metabolic studies of the cornea using redox fluorometry are proposed. Corneal redox fluorometry, a method originally developed in this laboratory, measures the biochemical state of the cornea in a non- invasive and non-destructive manner that has the capability of clinical application. The method applied clinically, would be invaluable both as a diagnostic tool and as a method for evaluation of treatment. No other currently available method provides the same essential information for the clinician. Corneal redox fluorometry measures the fluorescence of reduced pyridine nucleotides and oxidized flavoproteins and this provides an index of cellular oxidation-reduction (redox) state. Measurements of this fluorescence and the amount of light scattering in the tissue as a function of corneal depth will be made using confocal microscopy to enable separation of the endothelial and epithelial signals from the stromal signal. In addition, the redox state of single cells and the differences in metabolic state between individual cells within the corneal endothelium and within the corneal epithelium will be measured. The experiments proposed are clinically relevant and designed to provide knowledge that will be directly applicable to providing better health care. The measurements will be made in perfused corneas in vitro and in animal in vivo. The proposed experiments will focus on investigating the corneal metabolic changes that occur during decreased oxygen availability, wound healing, corneal storage, and corneal disease. The non-invasive measurements will be correlated with histological and electron microscopic studies made on the same tissue. The possibility of applying the methods developed to clinical diagnosis and treatment will also be investigated and animal measurements necessary to obtain approval for human measurements will be made.
