Ibuprofen Protects Rat Livers From Oxygen-Derived Free Radical-Mediated Injury After Tourniquet Shock

Abstract

Rats subjected to tourniquet shock suffer a severe form of circulatory shock, tissue and organ oxidative stress, and final multiple system organ failure (MSOF) and death of the animals within 24 h of tourniquet release. The oxidative damage observed in hind-limb muscle tissue after reperfusion does not by itself account for the final systemic and lethal MSOF. We have postulated that organ failure has its genesis in a primary perfusion abnormality, e.g. the hind limbs, which is followed by secondary hypoperfusion of other organs, such as the liver, as has been shown to be the case in several septic shock models. It has also been shown that injured or necrotic tissue can activate neutrophils, Küpffer cells, platelets, and both the complement and coagulation cascades. In turn, complement activation also leads to neutrophil and Küpffer cell activation as assessed by their capacity to generate oxyradicals. Herein we have evaluated the potential protective effect of ibuprofen on hepatic oxygen-derived free radical production, as well as its effects on both polymorphonuclear leucocyte (PMN) activation and liver infiltration. The protective effect of ibuprofen on hepatic oxidative injury was assessed by determining total thiol groups (SH), thiobarbituric acid-reactive substances (TBARS), and by the release of aspartic acid (AsT) and alanine (AIT) aminotransferases in control animals, in animals subjected to 5 h of tourniquets, and in animals after 2 h of hind-limb reperfusion. Liver infiltration by PMNs was determined by histology after staining with cosin-hematoxylin, and PMN activation by their capacity to reduce nitro blue tetrazolium (NBT). Our results show that total hepatic thiol content decreased significantly, over and above the the normal circadian decrease in liver glutathione, after the 5 h tourniquet period (from 6.16 ± 0.97 to 4.07 ± 0.21 μmoles/g w.w). The decrease in liver thiols in animals pretreated with ibuprofen was not significantly different from that in control animals (from 5.76 ± 0.21 to 4.69 ± 0.19, μmoles/g w.w), and could be accounted for by the circadian effect. A further significant decrease was observed in the control (3.01 ± 0.12μmoles/g w.w), but not in the ibuprofen pretreated rats (4.65 ± 0.16 μmoles/g w.w), after the 2 h reperfusion period. TBARS production remained essentially unchanged during the tourniquet period in both the control and ibuprofen pretreated animals (average 260 nmoles/g w.w), but increased significantly after hind-limb reperfusion in the control animals (386.9 ± 18.5 nmoles/g w.w), but not in the ibuprofen treated rats (267.2 ± 7.4 nmoles/g we.w). The protective effect of ibuprofen was also evident in plasma aminotransferase levels (AsT and AIT) which increased 14 and 6-fold, respectively, during the experimental period in the untreated rats, and only 6 and 3-fold in the animals pre-treated with the drug. No significant differences were observed in PMN liver infiltration in any of the animals, nor at any of the different time periods under study. Nevertheless, our results indicate that there is a 3-fold increase (over control values) in the number of circulating activated PMNs after hind-limb reperfusion in the non-protected control rats, and only a 2-fold increase in those protected by ibuprofen. It is concluded that ibuprofen: a) protects rat livers from the oxidative stress which results after 2 h of reperfusion of rat hind limbs subjected to 5 h of ischemia by means of tourniquets, and b) significantly decreases the number of NBT-positive PMNs in the systemic circulation after hind-limb reperfusion.

Key Words:

• Oxidative stress
• ibuprofen
• oxyradicals
• liver
• tourniquet shock
• ischemia
• reperfusion
• free radicals