Abstract
Despite advancements in imaging and interventional technologies, radiocontrast media continue to pose a risk of contrast-induced acute kidney injury (CI-AKI) in a subset of patients prone for this complication. Iohexol, which is a non-ionic, monomeric, iodinated low osmolar radiocontrast agent induces direct renal cytotoxicity and renal ischemia resulting in reactive oxygen species (ROS) formation. Besides, 1,25-dihydroxyvitamin D3, the active form of Vitamin D is reported to have an antioxidative role. In this study, we intended to demonstrate the significance of antioxidant effect of 1,25-dihydroxyvitamin D3 on Vero cells exposed to Iohexol in vitro. The cytotoxicity induced by Iohexol was tested in Vero cell line (ATCC® CLL-81™) by viability assay (MTT assay). Varying doses of Iohexol containing increasing concentrations of iodine (17.5 to 300 mg/ml) were used and corresponding morphological changes of cell viability were studied in relation to a control group. Total ROS is detected using PathScan® Total ROS1 Sandwich ELISA Kit (CST, MA, USA). Vero cells treated with or without Iohexol in the presence or absence of 1,25-dihydroxyvitamin D3 were analyzed after staining the cells with DAPI (4’6-diamidino-2-phenylindole). Lipid peroxidation was determined by TBARS (Thiobarbituric acid reactive species) assay. Antioxidants including total thiol content were assessed by Elmman’s method, Catalase by Colorimetric method and Superoxide dismutase (SOD) by Nitroblue tetrazolium assay. In our preliminary experiments, it was found that in comparison with the control cells, the cell viability dramatically decreased in Iohexol-treated cells in a time- and dose-dependent manner (Figures 1,2). ROS buildup and corresponding increase in lipid peroxidation were seen in Iohexol-treated cells (Figures 3,4,5). These effects were significantly reduced in cells exposed to 1,25-dihydroxyvitamin D3. Vero cells displayed decreased viability in the presence of Iohexol in a dose- and time-dependent manner. Increase in lipid peroxidation occurred along with a concomitant reduction in antioxidant levels indicating generation of ROS. Furthermore, 1,25-diihydroxyvitamin D3 reduced the production of ROS and thereby increased the cell viability.
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