Abstract
Purpose: :
The crystalline lens is a unique cellular organ that performs metabolic processes while maintaining optical functionality. The mitochondria play a vital role in providing the cell with the energy necessary for these metabolic processes. Menadione has long been known as a cytotoxin that specifically targets mitochondria. Upon entering the mitochondria, reactive oxygen species are formed inducing the formation of the permeability transition pore which renders the mitochondria inactive. The purpose of this study was to investigate the effectiveness of menadione sodium bisulfite as a mitochondrial uncoupler in the lens and to determine the relationship between lens metabolism and optical function.
Methods: :
Bovine lenses were extracted from whole bovine eyes and treated for 30 minutes at room temperature with 50 µM (n=6), 200 µM (n=4), 600 µM (n=5) and 1000 µM (n=6) menadione (w/v) in medium. A control (n=5) was treated with medium only. Lenses were rinsed 4 times then placed in fresh culture medium. Focal lengths of laser beams passing through the lenses at varying eccentricities were measured at 4, 24, 48, 144, and 216 hours post-treatment. Mitochondrial activity and dynamics were analysed using the Rhodamine 123, a mitochondria-specific marker for intact and functioning mitochondria, and were visualised using confocal microscopy. The images captured are analyzed using Zeiss and MATLAB software.
Results: :
The results show a concentration and time-dependent decrease in optical function that became apparent as early as 24 hours after treatment with 1000 µM and 600 µM menadione treatment groups after which damage was sustained throughout the experiment. Lenses exhibited optical degradation for 200 µM treatment group 48 hours after treatment while no change in optical function was observed for 50 µM treatment group. Confocal microscopy images show that changes in mitochondrial activity and dynamics precede optical changes.
Conclusions: :
These results show menadione to be an effective mitochondrial uncoupler and that mitochondrial function is directly related to optical function.
Keywords: mitochondria • metabolism • oxidation/oxidative or free radical damage