Measurement of oxidative stress in human liver by EPR spin-probe technique

A method for the measurement of reactive oxygen species (ROS) in human hepa tic tissue has been developed. The method is based on the EPR detection of the nitroxide radical produced by reaction of the hydroxylamine spin-probe bis(1-hydroxy-2,2,6,6-tetramethyl-4-piperidinyl)decandioate with ROS genera ted under pseudo-physiologic conditions in fine needle biopsies of healthy (10 controls) and diseased (22 patients) human liver. Measures of malonalde hyde in 9 liver biopsies (3 controls and 6 patients) have also been obtaine d by high pressure liquid chromatography and values parallel those obtained by the spin-probe technique. The amount of ROS found in healthy human live r (median = 1.8 x 10(-11) mol/mg) was significantly lower than values found in liver affected by hepatitis B (median = 5.8 x 10(-10) mol/mg; p < 0.02) or by hepatitis C (median = 2.7 x 10(-9) mol/mg; p < 0.003) as well as com pared to some other non-viral liver diseases (NVLD): autoimmune hepatitis, primary biliary cirrhosis, primary schlerosing cholangitis (median = 9.8 x 10(-9) mol/mg; p < 0.005). NVLD also showed significantly higher ROS levels compared to hepatitis B (p < 0.04) and hepatitis C (p < 0.04). The mechanism, potentiality and limitations of our method are discussed.