BACTERIAL LIPOPOLYSACCHARIDE INDUCES EARLY AND LATE ACTIVATION OF PROTEIN-KINASE-C IN INFLAMMATORY MACROPHAGES BY SELECTIVE ACTIVATION OF PKC-EPSILON

Experiments from our and other laboratories have shown that specific i nhibitors of protein kinase C (PKC) inhibited the secretion of nitric oxide, TNF alpha, and IL-1 beta from lipopolysaccharide (LPS)-stimulat ed macrophages, suggesting an important role for PKC in the inflammato ry response. The present study was designed to investigate the mechani sm whereby LPS stimulates PKC activity in inflammatory macrophages. Mo use macrophages were stimulated with 0-1 mu g/ml LPS for 0-18 hours, a nd PKC activity was detected in cell lysates. PKC isoform specificity was determined by blocking PRC activity with isoform-specific antibodi es. Treatment of macrophages with 1 mu g/ ml LPS induced a two-fold in crease in PKC activity within 15 minutes and an additional more signif icant peak of PKC activity appeared 3 hours post-LPS stimulation. A lo wer dose of LPS (10 ng/ml) induced the later peak only. The enhancemen t in PKC activity induced by LPS occurred in both the cytosol and memb rane fractions, but the enhancement in the membrane fraction was signi ficantly greater than in the cytosol. The increase in PKC activity in both peaks was abolished only by the addition of anti-PKC-epsilon anti body. The present experiments suggest that PKC activation is an import ant pathway in the LPS-induced secretory response of macrophages and t hat PEC-epsilon is the major isoform involved. (C) 1997 Academic Press .