The transcription factor nuclear factor B (NF-B) regulates multiple immediate-early gene expressions involved in immune and inflammatory responses and cellular defenses. Ischemia-reperfusion induces many immediate-early gene expressions, but little is known about the NF-B activation in myocardium during ischemia and reperfusion. This study demonstrated that ischemia alone rapidly induced NF-B activation in the myocardium of isolated working rat hearts. Electrophoretic mobility shift assay showed that NF-B binding activity significantly increased in the nucleus after 5 min of ischemia and remained elevated for up to 30 min. Western blot analysis suggested that the levels of inhibitory IB protein in the cytoplasm became markedly decreased at 4, 5, 7.5, and 10 min of ischemia but were gradually restored following 10 min of ischemia. Reduction of IB protein in the cytoplasm by ischemia resulted in NF-B translocation to the nucleus. Northern blot hybridization showed that IB mRNA levels were not significantly elevated during myocardial ischemia. Pyrrolidine dithiocarbamate, an antioxidant, significantly inhibited the loss of IB protein from the cytoplasm and prevented NF-B binding activity in the nucleus. Reperfusion following short periods of ischemia augmented NF-B binding activity in the nucleus induced by ischemia. The results suggest that early activation of NF-B induced by ischemia in the myocardium could be a signal mechanism for controlling and regulating immediate-early gene expression during ischemia-reperfusion.

inhibitory protein IB; nuclear factor B; reperfusion; antioxidant; reactive oxygen species

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