Proteinase activated receptor-2 (PAR-2) is a novel G-protein coupled receptor, that is activated by means of proteolytic cleavage (Macfarlane et al, 2001) and has both pro and anti-inflammatory actions depending upon the system examined. PAR-2 has been linked to the stress-activated protein kinases, JNK and p38 MAP kinase and also NFκB (Kanke et al, 2001; Sabri et al, 2001), pathways known to be involved in pro-inflammatory responses in several cell types. The cytokine TNFα is also an important mediator of pro-inflammatory responses, and indeed signals through both the SAPK and NFκB pathways. Since both TNFα and PAR-2 activators are released during inflammation, we examined the possibility of crosstalk/synergy between PAR-2 and TNFα at the level of the SAP kinases and NFκB signalling pathways. The skin epithelial cell line NCTC2544 stably expressing PAR-2(Clone G), was generated and incubated with trypsin or PAR-2 activating peptide (AP) and TNFα. The SAPK and NFκB activation was examined in whole cell lysates, which had been resolved by Western blotting, using phospho-specific antibodies. In Clone G cells trypsin (50nM), SLIGKV-OH (200μM) and TNFα (20ng/ml) caused a significant increase in phospho-p38 MAP kinase and NFκB levels. Preliminary results failed to show evidence of synergy as prolonged TNFα pre-treatment (1-24hrs) failed to modify PAR-2 mediated signalling, whilst PAR-2 activation failed to potentiate SAP kinase and NFκB activation mediated by low concentration of TNFα (1ng/ml). However, surprisingly, activation of PAR-2 using low concentrations of trypsin (1-20nM) or AP (1-30µm) resulted in a reduced/inhibitory effect on the ability of TNFα to signal through p38 and NFκB. The same effect was observed in the EAHy926 cell line (endothelial cells), which endogenously express PAR-2. The phenomenon was not observed for PAR-4 activation. Studies investigating the signalling pathways involved in this response will be presented. These results indicate a potential mechanistic explanation for both the anti and pro-inflammatory actions of PAR-2, dependant upon the occupancy of the receptor.