Safety Of Trypan Blue In Combination With Light On Arpe–19 And R28 Cells In Vitro

Abstract

Abstract: : Purpose:To evaluate the safety of trypan blue, in combination with light, on retinal pigment epithelial and neurosensory retinal cells. Methods:Human retinal pigment epithelial cells (ARPE–19) and rat neurosensory retinal cells (R28) were cultured and treated with 0.1%, 0.05%, 0.025% and 0.0125% concentration of trypan blue for 2 minutes and then exposed to a Grieshaber surgical halogen light source for 0, 5 or 10 minutes. Toxicity was measured by dye exclusion assay, mitochondrial dehydrogenase assay and tritiated [H³] thymidine incorporation assay. Results:ARPE–19 cells did not show any evidence of toxicity to trypan blue and/or light as measured by the three different assays. R28 cells exposed to 10 minutes of light at any of the concentrations of trypan blue showed a significant reduction in the activity of mitochondrial dehydrogenase (p<0.001). Cells exposed to the higher concentration of trypan blue (0.1%), even without light, showed a significant reduction of mitochondrial dehydrogenase activity (p<0.001). However, 5 minute light exposure with the lower concentrations of trypan blue (0.05%, 0.025% and 0.0125%) did not show any significant reduction in the mitochondrial dehydrogenase activity. R28 cells did not show any significant decrease in cell viability by the dye exclusion assay and [H³] thymidine incorporation assay. Conclusions:Trypan blue is an alternative to indocyanine green (ICG) for membrane peeling for macular hole surgery and proliferative vitreoretinopathy. In this study, trypan blue was non–toxic to the human retinal pigment epithelial cells (ARPE–19) but its effect on mitochondrial function in neurosensory retinal (R28) cells may imply some risk for toxicity.

Keywords: retinal culture • macular holes • vitreoretinal surgery