Page Banner

United States Department of Agriculture

Agricultural Research Service

Title: Knockdown of Thioredoxin Reductase-1 (Tr) Results in Increased Measures of Cellular Oxidative Damage

Authors
item Hintze, Korry - NDSU, FARGO, ND
item Wald, Karl
item Finley, John

Submitted to: Journal of Federation of American Societies for Experimental Biology
Publication Type: Abstract Only
Publication Acceptance Date: December 1, 2003
Publication Date: March 24, 2004
Citation: Hintze, K.J., Wald, K., Finley, J.W. 2004. Knockdown of thioredoxin reductase-1 (tr) results in increased measures of cellular oxidative damage [abstract]. Federation of American Societies for Experimental Biology Journal. 18:A917.

Technical Abstract: The selenoprotein TR functions as an antioxidant by providing reducing equivalents to thioredoxin and a wide variety of other substrates including dehydroascorbic acid, the ascorbyl free radical, lipoic acid and vitamin K3. To study the antioxidative function of TR, expression in HepG2 cells was knocked down by siRNA technology. Compared to controls (treated with equimolar concentrations of luciferase siRNA), 250 nM TR-specific siRNA reduced TR mRNA expression (measured by RT-PCR) ~83%, antibody-reactive protein ~80% and enzyme activity 82% (4.1 ± 0.3 and 0.75 ± 0.7 mU/prot./min; P < 0.0001, controls and knockdowns, respectively). Knockdown of TR did not affect cell growth (P > 0.05) or cell cycle (measured by flow cytometry) but did increase total glutathione (10.4 ± 1.4 and 7.5 ± 0.2 mM/mg protein; P < 0.05, knockdowns and controls, respectively). Total protein carbonyl formation (measured by western blot) was increased relative to controls in knockdown cells; this occurred in both unstressed (2-fold increase) and cells stressed with 200 uM H2O 2 for ½ h (1.4-fold increase). These data demonstrate that TR knockdown is not detrimental to cell growth or cycle. However, they do suggest that the antioxidant function of TR prevents protein oxidation in HepG2 cells. Also, knockdown cells may have compensated for decreased anti-oxidant status by up-regulating glutathione production.

Last Modified: 11/25/2014
Footer Content Back to Top of Page