Bradykinin and thrombin synergistically potentiate interleukin-1 and tumour necrosis factor induced prostanoid biosynthesis in human dental pulp fibroblasts

The effects of and interactions between the six inflammatory mediators interleukin 1 (IL-1), tumour necrosis factor (TNF), gamma-interferon (IFN-gamma), transforming growth factor-beta (TGF-beta), bradykinin (BK) and thrombin on prostanoid biosynthesis in primary cultures of human, dental, pulp fibroblasts were examined. IL-1 alpha, IL-1 beta, TNF-alpha and TNF-beta caused a time- and concentration-dependent enhancement of prostaglandin E(2) (PGE(2)) formation in the fibroblasts. The onset of action was delayed 1-2 h and maximal response was seen after 24 h. In contrast, BK and thrombin caused a burst of PGE(2) formation with maximal response after 10 min. BK (1 mu M) and thrombin (3 U/ml) synergistically potentiated IL-1 alpha and IL-1 beta stimulated PGE(2) formation in 24 h cultures. The effect of BK and thrombin on IL-1 beta enhanced PGE(2) formation was seen both at suboptimal and optimal concentrations of IL-1 beta without affecting the sensitivity to IL-1 beta. BK and thrombin also synergistically potentiated the stimulatory effect of TNF-alpha and TNF-beta on PGE(2) formation. The synergistic interactions between BK and IL-1 alpha, IL-1 beta and TNF-alpha were seen after 24 h of treatment. BK analogues with affinity to BK B2-receptors, but not to BK B1-receptors, were able to synergistically potentiate IL-1 beta and TNF-alpha-enhanced PGE(2) production. The synergistic stimulation of PGE(2) formation by IL-1, TNF and BK was abolished by indomethacin and flurbiprofen. Preincubation with IL-1 beta and TNF-alpha for 24 h resulted in a substantial amplification of the PGE(2) response to a subsequent 24 h challenge with BK in the absence of cytokine. Similarly, when the pulp fibroblasts were preincubated with or without IL-1 beta or TNF-alpha for 24 h and then challenged with exogenous arachidonic acid for 60 min, PGE(2) formation was significantly enhanced in cytokine pretreated cells. BK potentiated cytokine induced amplification of the PGE(2) response to arachidonic acid. gamma-IFN and TGF-beta did not enhance PGE(2) formation, nor did these cytokines potentiate IL-beta or TNF-alpha-induced PGE(2) production. These data show that proinflammatory mediators such as BK and thrombin act in concert with IL-1 and TNF in stimulating prostanoid formation in human pulpal fibroblasts and that the action of BK is linked to BK B2-receptors. The results are compatible with the view that enhanced metabolism of arachidonic acid, probably due to increased activation or de novo synthesis of cyclooxy-genase(s), is involved in the mechanism by which IL-1, TNF, BK and thrombin interact. (C) 1996 Academic Press Limited