In vitro study for staining and toxicity of rose bengal on cultured bovine corneal endothelial cells

It has recently been reported that rose bengal (RB) is actually not a vital dye and that it has an intrinsic toxic effect. Therefore, we conducted a study to determine its staining capability and the presence of intrinsic toxicity. We also examined its effects on the proliferation of cultured bovine corneal endothelial cells. The cells stained despite short exposure (1 s) to rose bengal under a fluorescein photomicroscope. The cell nucleus stained first, followed by the cytoplasm; the intensity was directly dependent on the exposure time. Although the entire cell layer did not stain at an RB concentration of 0.001%, the cell layer stained diffusely above a concentration of 0.01%. At a 5% concentration, areas of cellular detachment and lysis were noted. The degree of cellular change was directly proportional to the concentration of RB and the exposure time. Cellular damage included cell swelling, intracytoplasmic vacuole formation, cell detachment, and lysis. The presence of light augmented the toxic effect of RB on cultured endothelial cells and was statistically more significant than RB's intrinsic toxic effect. Exposure to RB concentrations > 0.01% induced statistically significant concentration-dependent inhibition of cell proliferation. These results suggest that rose bengal may not actually be a vital dye on cultured bovine corneal endothelial cells and that it has intrinsic inhibitory and toxic effects with additional phototoxic effects on corneal endothelial cells. Furthermore, cell proliferation was markedly inhibited after exposure to RB.