Thrombin and PAR-1 activating peptide increase iNOS expression in cytokine-stimulated C6 glioma cells.

Thrombin (THR) plays a key role in the brain under physiol. and pathol. conditions. Several of the biol. activities of thrombin have been shown to be mainly driven through activation of protease-activated receptor-1 (PAR-1)-type thrombin receptor. Here we have studied the effect of THR and PAR-1-activating peptide (PAR1-AP), SFLLRN, on cytokine-induced expression of inducible nitric oxide (iNOS), a prominent marker of astroglial activation using the rat C6 glioma cells. In this cell line, THR (1-10 U/mL) and PAR1-AP (1-100 mM) induced a significant concn.-dependent increase both of IFN-g- (250 U/mL) or TNF-a- (500 U/mL) induced NO release. The obsd. increase of NO prodn. was related to an enhancement of iNOS expression as measured in cell lysates prepd. from different treatments by using SDS-PAGE followed by western blot anal. The effect of THR, but not that of PAR1-AP, was significantly inhibited by hirulog (60 mg/mL), a specific and stoichiometric THR inhibitor or by cathepsin-G (40 mU/mL), an inhibitor of PAR-1. In conclusion our data suggest a role for THR through activation of PAR-1 in the induction of astroglial iNOS, and further support the hypothesis that THR may function as an important pathophysiol. modulator of the inflammatory response.