Neutrophil chemoattractants generated in two phases during reperfusion of ischemic myocardium in the rabbit. Evidence for a role for C5a and interleukin-8

Abstract

The neutrophil chemoattractants generated in a model of myocardial infarction in the anesthetized rabbit were investigated. Coronary artery occlusion was followed by reperfusion for periods from 5 min to 4.5 h. Extracts of myocardial tissue in normal and post-ischemic zones were tested for C5a and interleukin-8 (IL-8) using specific radioimmunoassays. In the post-ischemic zone, immunoreactive C5a was detected within 5 min and rose progressively to reach a plateau at 3-4.5 h. In contrast, immunoreactive IL-8 concentrations rose after a delay and were highest at the last time point tested, 4.5 h. Myeloperoxidase activity levels, an index of neutrophil accumulation, rose progressively as the concentrations of chemoattractants increased. Using cation exchange and reversed phase HPLC, immunoreactive C5a and IL-8 co-eluted with authentic standards. Fractions taken at the C5a and IL-8 peaks from reversed phase HPLC exhibited neutrophil aggregating activity which was neutralized by the respective antibody used in the radioimmunoassays. Depletion of circulating neutrophils virtually abolished immunoreactive IL-8 in the post-ischemic myocardial tissue. These observations suggest a sequential release of chemoattractants: the first, C5a is generated in interstitial fluid, followed by IL-8 generated by infiltrating neutrophils. Thus, over the time period studied, IL-8 generation would be expected to be indirectly dependent on C5a production.