This project focuses on the biochemical pathways involved in signal transduction in the monocyte. Recent studies have examined the role of guanine nucleotide binding proteins (G proteins) in the prostaglandin-cyclic AMP dependent pathway of collagenase production by monocytes. Western blot analysis of the G proteins revealed that the same G proteins were present in the membrane of control and stimulated monocytes. However, ADP-ribosylation assays revealed that cholera toxin (CT) ADP-ribosylated a 46-kDa Gs(alpha) protein only in the membranes of monocytes that had been stimulated with agents such as Con A or LPS. This correlated with the ability of CT to enhance eicosanoid synthesis of activated but not control monocytes, whereas CT elevated cAMP levels in both control and stimulated monocytes. These results suggest that the 46-kDa Gs(alpha) protein may be involved in the regulation of phospholipase activity. Thus, G proteins regulate both phospholipase and adenylyl cyclase in monocytes, events that are an integral part of the activation sequence leading to collagenase production by these cells. In additional studies, since IL-4 has been shown to inhibit PGE2 synthesis by monocytes, this cytokine was examined for its effect on the biochemical events leading to collagenase production. Nanogram amounts of IL4 were shown to not only inhibit PGE2, but to also block the release of arachidonic acid and the subsequent synthesis of metabolites from both the cyclooxygenase and lipoxygenase pathway as well as collagenase. These findings indicated that IL-4 is a potent suppressor of phospholipase activity of the monocyte. Evidence for the suppression of phospholipase by IL-4, and therefore the subsequent PGE2 production, as the mechanism for inhibition of collagenase was demonstrated by the ability of exogenous PGE2 to restore collagenase production in IL-4 treated monocyte cultures.
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