| Methanol has neurotoxic actions on the human retina due to its metabolite formic acid, which is a mitochondrial toxin. While optic nerve damage has been documented in both human methanol poisonings and in non-human primate studies, recent investigations using a methanol intoxicated rat model showed an apparent direct retinotoxic action of formate. Electroretinogram (ERG) studies in these animals strongly implicate photoreceptor damage, and morphologic changes were seen both in retinal photoreceptors and in cells of the underlying retinal pigment epithelium (RPE). Here we provide similar evidence for direct retinal effects in humans as shown in an acute, fatal case of methanol poisoning. Further, we developed a cell culture model to analyze the formate sensitivity of retinal cells incorporating the metabolic acidosis that accompanies methanol poisoning. We found greater ATP depletion and cytotoxicity upon formate exposure in the photoreceptor (661W) as compared to the RPE (ARPE-19) cell line. Differences in cytotoxicity were not due to differences in formate accumulation, which was similar in both cell types. Cell death from formate exposure is believed to result from formate inhibition of the electron transport chain leading to a reduction in ATP too low to perform essential cell functions, or to an increase in reactive oxygen species (ROS) secondary to electron transport blockade. Therefore, we also examined whether the cell types differ in the antioxidants necessary to detoxify ROS. Compared to ARPE-19, 661W cells showed lower amounts of glutathione, glutathione peroxidase, catalase protein, and catalase activity. Formic acid produced decreases in glutathione and glutathione peroxidase in both cells types and glutathione inhibition with buthionine L-sulfoximine (BSO) produced greater ATP depletion and cytotoxicity in both cell types. In contrast, formate exposure produced decreases in catalase protein and activity in 661W cells, but increases in ARPE-19 cells. Treatment with the catalase activity inhibitor 3-amino-1,2,4-triazole (AT) increased the formate sensitivity of only ARPE-19 cells. ARPE-19 cells therefore may be less susceptible to formate toxicity due to higher levels of antioxidants, especially catalase, which increases on formate treatment and which has a significant cytoprotective effect for the RPE cell line. We further conclude that 661W photoreceptor and ARPE-19 cell lines retain cell type-specific differences in glutathione/glutathione peroxidase and catalase, with the ARPE-19 cultures showing high antioxidant levels as expected for cells derived from the RPE. Cell lines provide an essential model for investigating cell type differences in the effects of formate and for probing why some cells are more sensitive to mitochondrial toxins than others. |
