Objective: Tumor Necrosis Factor alpha (TNF alpha) induces programmed cell death and contributes to cardiac ischemia/reperfusion injury. Paradoxically, we have recently demonstrated that low doses of TNF alpha can induce cardiac preconditioning (PC). We hypothesized that the production of free radicals participates in this cardioprotective program.
Methods: Control isolated rat hearts underwent 30 min regional ischemia and 120 min of reperfusion. A second group of hearts received a low dose of TNF alpha (0.5 ng/ml) for 7 min followed by 10 min washout prior to I/R. In other groups, the antioxidant N-2-mercaptopropionyl glycine (MPG) (1 mM) was given for 15 min prior to I/R alone or during TNF alpha perfusion. Infarct size was determined at the end of the reperfusion period. Ventricular catalase and superoxide dismutase activities were assessed as an index of oxidative stress and free radical production was directly measured by the oxidation of 1-hydroxy-3-carboxy-pyrrolidine (CP-H) to paramagnetic 3-carboxy-proxyl (CP.) using electron spin resonance spectroscopy.
Results: TNF alpha reduced the infarct/area at risk (I/AAR) ratio (7.2+/-1.7% vs. 36.5+/-1.7% for controls, p<0.05). MPG reduced the cardioprotective effect of TNF alpha (I/AAR ratio: 20.5+/-3.3%, p<0.05). TNF alpha-perfusion increased catalase activity in the ventricles (15.8+/-1.2 I.U./mg for controls vs. 19.9+/-1.1 I.U/mg for TNF alpha, p<0.05). Proof of formation of free radicals was increased CP formation in the coronary effluent during TNF alpha infusion (24.2+/-4.5 for TNF alpha vs. 11.9+/-1.5 arbitrary units for controls, p<0.05), with decreased CP after addition of MPG.
Conclusions: Our data provide firm evidence for a production and role of free radicals in TNF alpha-induced cardioprotection.