Lens Optical Function and Loss of Mitochondria in Bovine Lenses Cultured With Elevated Glucose in vitro

Abstract

Abstract: : Purpose:This study investigates whether lens optical function and the mitochondrial integrity of epithelial and superficial cortical lens fiber cells of the bovine lens are sensitive indicators of early damage associated with high glucose in vitro. Methods:Bovine lenses were cultured in glucose-supplemented culture medium (65-195 mM glucose, with or without serum) for 7 days at 37₀C and 4-5% CO₂. Lens optical function was analysed daily using the Scantox^TM In Vitro Lens Assay System (Harvard Apparatus, Holliston, MA). On the last day the mitochondria were stained using 20 µM rhodamine 123 fluorescence and viewed by confocal microscopy. Results:No significant differences in optical or mitochondrial properties were seen between serum and serum-free groups. Concentration-dependent loss of sharp focus was measured by the variability of back vertex distance. This was evident for all glucose concentrations. Significant change (p<0.05) was seen at day one for the 195 mM glucose treatment group only. At that point, loss of sharp focus increased from 0.32±0.02 mm (SEM) initially, to 0.90±0.05 mm (SEM) (serum group, n=13, and serum-free, n=7). However, sharpness of focus returned to the level of controls by day 3. Sharpness of focus in the 65 mM glucose group (n=7 for serum-free and n=15 for serum groups) or the 130 mM glucose group (n=7 for serum-free and n=10 for serum) did not change significantly from controls. After 7 days a concentration-dependent decrease in number and length of mitochondria of epithelial and superficial cortical fibre cells was seen. The depth below which the mitochondria of the superficial cortical fibre cells disappear changed significantly (p<0.05) from controls (178.60±13.70 mM SEM) in all treatment groups, decreasing to 79.80±6.58 mM (SEM), 114.00±13.16 mM (SEM) and 102.60±14.72 mM (SEM) in the 195, 130 and 65 mM glucose group lenses, respectively. Conclusion:These results indicate that serum has no effect on lens damage. The results also show that significant loss of sharp focus at higher glucose concentrations occurs within one day. All concentrations of glucose cause a decline in mitochondrial integrity. Moreover, the significant concentration-dependent loss of mitochondria shows that the mitochondria of superficial cortical fibre cells are most sensitive to glucose damage.

Keywords: 387 diabetes • 500 optical properties • 475 mitochondria