[Anti-inflammatory effect of Syk inhibitor in LPS stimulated macrophages].

OBJECTIVE

To investigate the role of spleen tyrosine kinase (Syk)on the Toll-like receptor 4 (TLR4) signal transduction cascade.

METHODS

RAW264.7 macrophages were treated with different concentrations of Syk inhibitor and then stimulated with 100 ng/mL lipopolysaccharide (LPS). The cell viability was tested by the Alarmarblue assay. The NO production was detected by Griess reagents and cytokines in culture supernatants were quantified by ELISA. The expression of iNOS was detected by Western blotting. Moreover, the activation of the transcription factors NF-κB and activator protein 1 (AP-1) was investigated using the reporter cell line RAW-Blue(TM);.

RESULTS

The Syk inhibitor had no effect on the cell proliferation even at concentrations of 14 μmol/L. However, the NO production (P<0.01) and iNOS expression resulting from LPS stimulation were inhibited, when the inhibitor concentration exceeded 1.5 μmol/L, and the TNF-α and IL-6 production were inhibited even at lower concentrations (P<0.01). Moreover, inhibition of the activation of both the transcription factors NF-κB and AP-1 was observed.

CONCLUSION

The Syk inhibitor shows a significant anti-inflammatory property due to its inhibitory effect on NO and cytokine production by LPS stimulated macrophages. This effect correlated with the reduced activation of both transcription factors NF-κB and AP-1, which places Syk downstream of the TLR4 signal pathway but upstream of the point of divergence of both transcription factors.