Characterization of advanced oxidation protein products (AOPP) production by bovine neutrophils

Advanced oxidation protein products (AOPP) are novel markers of protein oxidation first characterized in uremic patients and related to hypochlorous acid (HOCl) production by activated neutrophils. To establish how oxidants contribute to the pathology of inflammation it is important to identify physiologically relevant targets of oxidation and to develop specific biomarkers to quantify the oxidative damage. Here, we characterized the relative reactivity of HOCl (0-25 mM) and cumene hydroperoxyde (CuOOH; 0-100 mM) with bovine serum albumin (BSA), bovine gamma globulins (bG) and whole bovine plasma. HOCl concentrations higher than 25 mM induced a massive protein denaturation, bG being the most susceptible. AOPP levels significantly increased in a dose-dependent manner in both HOCl-treated BSA and bG (P<0.01), whereas AOPP levels in plasma were significantly higher than the unexposed control only at 25mM HOCl concentration (P<0.05). The AOPP yield was lower when CuOOH was the oxidant agent. The AOPP levels were significantly higher (P<0.01) than the unexposed control when BSA was treated with 10 mM CuOOH, and bG and plasma were treated with 100 mM CuOOH. A significant (P<0.05) dose-dependent dityrosine and carbonyl group formation was observed in both BSA and bG exposed to HOCl. A significant dose-dependent dityrosine production was induced by CuOOH in BSA (P<0.01), bG (P<0.001) and whole plasma (P<0.05). Strong positive correlations were observed between AOPP and dityrosine in HOCl-treated BSA (r=0.977, P<0.001), bG (r=0.962, P<0.001) and plasma (r=0.811, P<0.01). Correlations between AOPP and dityrosine were weaker CuOOH-treated BSA (r=0.572, P<0.05), bG (r=0.812, P<0.01) and plasma (r=0.672, P<0.05).