INVOLVEMENT OF PROTEIN-KINASE-C AND PROTEIN-TYROSINE KINASE IN LIPOPOLYSACCHARIDE-INDUCED TNF-ALPHA AND IL-1-BETA PRODUCTION BY HUMAN MONOCYTES

Bacterial LPS stimulates human monocytes to secrete inflammatory cytok ines, which are involved in several disease processes. However, the me chanism of LPS activation of cytokine expression and secretion is not completely understood. In this study, we investigated the signal trans duction pathways involved in LPS-stimulated TNF-alpha and IL-1 beta se cretion. TNF-alpha and IL-1 beta secretion were completely blocked by protein kinase C (PKC) and cyclic nucleotide-dependent protein kinase inhibitor, H-7, but were not affected by H-89, a specific cyclic nucle otide-dependent protein kinase inhibitor. In addition, LPS was found t o induce activation of PKC, reaching maximal activity at 30 min and re turning to unstimulated levels after 60 min. LPS stimulation only slig htly increased intracellular levels of diacylglycerol, the natural act ivator of PKC, and pretreatment of monocytes with the diacylglycerol-k inase inhibitor, R59022, did not affect LPS-stimulated TNF-alpha secre tion. LPS-induced PKC activation was found not to be affected by block ing of the LPS receptor, CD14, with mAb or by inhibition of protein ty rosine kinase with herbimycin A. However, these agents suppressed LPS- induced TNF-alpha secretion and TNF-alpha mRNA accumulation. The resul ts suggest that TNF-alpha and IL-1 beta secretion after LPS stimulatio n of human monocytes requires the activation of protein tyrosine kinas e and PKC, upstream to the activation of gene transcription. The activ ation of PKC by LPS is probably mediated by a diacylglycerol-independe nt pathway.