Abstract
Purpose: :
Boric acid is associated with cell proliferation, regulation of glutathione and possibly reactive oxygen species (ROS) levels. SLC4A11 is mutated in patients with Fuchs corneal dystrophy and a recent publication has indicated that this protein is a sodium borate transporter. This suggests that regulation of boric acid concentration in corneal endothelial cells may influence proliferation and ROS production. Therefore, we tested the effect of boric acid depletion and supplementation on proliferation and ROS production of bovine corneal endothelial cells (BCEC) and also HEK293 cells.
Methods: :
BCEC and HEK293 cells at 30-50% of confluence were serum-deprived for 16 hrs. Then cells were incubated with either boric acid depleted medium alone or supplemented with different concentrations of boric acid for 24-48 hrs. MTT assay was carried out to test cell growth and viability. For ROS assays, confluent BCEC and HEK293 were incubated with normal medium, boric acid depleted medium or depleted medium supplemented with different concentrations of boric acid for different time points between 1 to 12 hrs, then loaded with the ROS sensitive dye, 5-(and 6)-carboxy-2’, 7’ dichlorodihydrofluorescein diacetate (carboxy-H2DCFDA) and fluorescence was measured by flow cytometry.
Results: :
MTT assay showed no significant effect of boric acid depletion and of supplemented boric acid (30-300 µM) on either cell type. Significant inhibition of proliferation was observed at 3000 µM (25 and 11% inhibition in BCEC and HEK293 respectively, p<0.05). Incubation of BCEC and HEK293 cells with boric acid depleted as well as supplemented medium (30-300 µM) for 1 to 12 hrs did not have any effect on ROS production.
Conclusions: :
At physiological concentrations (below 100 µM), boric acid has no effect on cell proliferation. The inhibitory effect of boric acid is observed at high concentrations. Boric acid depletion or supplementation has no effect on cell ROS at the time points studied. These results indicate that boric acid is not an important regulator of proliferation and ROS levels in BCEC and HEK293 cells.
Keywords: cornea: endothelium • proliferation • oxidation/oxidative or free radical damage