Ischemia / Hypoxia Induced Activation of Transcription Factor NF-KB in RAT Cardiac Myocyte
Document Type
Article
Publication Date
12-1-1996
Description
Cytokine genes expression may play an important role in ischemia/reperfusion injury associated with acute myocardial infarction. We have recently observed that occlusion of left anterior descending coronary artery followed by reperfusion increased cardiac TNF-a mRNA expression in rats. The transcription factor NF-kB is a pleiotropic regulator of many genes involved in immuno and inflammatory responses. To elucidate the mechanisms of TNF-a mRNA expression induced by ischemia/reperfusion, we have investigated the activation of NF-kB in the ischemia/hypoxia cardiac myocytes in rats. Cardiac myocytes from adult rat ventricles were isolated by perfusion and incubation with buffer containing collagenase and hyaluronidase (Kao, 1980). The isolated cardiac myocytes were harvested and washed with modified Medium 199 containing 10% calf serum. The cells (2 x 107) were placed in microcentrifuge tubes, covered with mineral oil, and incubated at 37° C for 5 to 120 min. to establish the kinetics of ischemia/hypoxiainduced activation of NF-kB in cardiac myocytes. After incubation, nuclear proteins were extracted from the cardiac myocytes and NF-kB activation was determined by gel electrophoretic mobility shift assay. Ischemia/hypoxia resulted in the stimulation of DNA binding activities of NF-kB in the rat cardiac myocytes at 5 minutes, which peaked within 15 to 30 min. The activation of NF-kB binding to DNA in ischemia/hypoxia cardiac myocytes maintained up to two hours. These results are the first demonstration of ischemia/hypoxia stimulation of transcription factor NF-kB binding activity in the rat cardiac myocyte.
Citation Information
Li, C.; Ha, T.; Browder, W.; and Kao, R. L.. 1996. Ischemia / Hypoxia Induced Activation of Transcription Factor NF-KB in RAT Cardiac Myocyte. FASEB Journal. Vol.10(3). ISSN: 0892-6638