Urate oxidase knockdown decreases oxidative stress in a murine hepatic cell line.
Cleveland-BM; Leonard-SS; Klandorf-H; Blemings-KP
Oxidat Med Cell Longev 2009 Apr; 2(2):93-98
Humans, birds, and some primates do not express the uric acid degrading enzyme urate oxidase (UOX) and, as a result, have plasma uric acid concentrations higher than UOX expressing animals. Although high uric acid concentrations are suggested to increase the antioxidant defense system and provide a health advantage to animals without UOX, knockout mice lacking UOX develop pathological complications including gout and kidney failure. As an alternative to the knockout model, RNA interference was used to decrease UOX expression using stable transfection in a mouse hepatic cell line (ATCC, FL83B). Urate oxidase mRNA was reduced 66% (p < 0.05) compared to wild type, as measured by real time RT-PCR. To determine if UOX knockdown resulted in enhanced protection against oxidative stress, cells were challenged with hexavalent chromium (Cr(VI)) or 3-morpholinosydnonimine hydrochloride (SIN-1). Compared to wild type, cells with UOX knockdown exhibited a 37.2 ± 3.5% reduction (p < 0.05) in the electron spin resonance (ESR) signal after being exposed to Cr(VI) and displayed less DNA fragmentation (p < 0.05) following SIN-1 treatment. Cell viability decreased in wild type cells (p < 0.05), but not cells with UOX knockdown, after treatment with SIN-1. These results are consistent with an increased intracellular uric acid concentration and an increased defense against oxidative stress.
Animal-studies; Antioxidants; Antioxidation; Biochemical-analysis; Biological-effects; Cell-biology; Cell-function; Cell-metabolism; Cell-morphology; Cellular-reactions; Hepatic-microsomal-enzymes; Hepatocytes; Laboratory-animals; Liver-cells; Liver-function; Liver-microsomal-metabolism; Metabolism; Oxidation-reduction-reactions; Oxidative-metabolism; Oxidative-processes;
Author Keywords: urate oxidase; RNA interference; oxidative stress; uric acid; SIN-1
Kenneth P. Blemings, West Virginia University, PO Box 6108, Morgantown, West Virginia 26506
Oxidative Medicine and Cellular Longevity