Production and characterization of the advanced oxidation protein products (AOPP) in bovine plasma proteins exposed to either hypochlorous acid or cumene hydroperoxyde.

Advanced oxidation protein products (AOPP) are novel markers of protein oxidation described in plasma of uremic patients and related to neutrophil activation. The observation that AOPP were higher in plasma of cows affected by late embryonic loss raised our interest in their biological role in cattle. Chlorinating agents, such as HOCl, induce protein modifications, which could be used as biomarkers of pathological events. In this study, we aimed to characterise bovine serum albumin (BSA; concentration) chlorination by bovine neutrophils, and to measure the AOPP production. BSA was incubated for 1, 2 and 3 hours with neutrophils isolated from healthy heifers (N=7), either activated or not activated with PMA. HOCl production was significantly higher in PMA-activated (400.5 ± 36.3 moles*hour) than in not-activated cells (126.8 ± 19.2 moles*hour; P<0.05). In parallel experiments (N=5), BSA was exposed to the same HOCl concentration observed in activated neutrophils over the same time-intervals. Levels of chloramines, dityrosines and carbonyls were significantly higher (P<0.05) when BSA was incubated with PMA-activated neutrophils or HOCl. Not-activated neutrophils produced a tangible amount of chloramines and dityrosines, likely attributable to basal HOCl release. PMA-activated cells generated significantly less (P<0.05) di-tyrosines (16.9 ± 1.1 nmol/mg*hour) and carbonyls (852,845 ± 299,271 OD unit*hour) than HOCl alone (di-tyrosines: 30.2 ± 1.5 nmol/mg*hour; carbonyls: 2,020,282 ± 726,031 OD unit*hour). In conclusion, bovine neutrophils produce enough HOCl to generate AOPP “in vitro”. However, the use of HOCl alone cannot fully mimic the complex events leading to BSA chlorination and AOPP production by activated neutrophils.