| dc.description.abstract | Interferon-β (IFN-β) is a crucial component of innate immune system which enhances IFN-stimulated genes (ISGs) expression to promote the host’s immunity. Numerous data indicated that various pathogenic structures, such as lipopolysaccharide (LPS), can up-regulate IFN-β production by activating toll like receptor (TLR) signal pathways. Many of these responses can be modulated by cAMP signaling. Type 4 cAMP-specific phosphodiesterases (PDE4s) are the predominant cAMP-hydrolyzing enzymes in most immune cells and, thereby important in modulating cAMP concentration in these cells. In this study, we aimed to determine whether PDE4 is involved in regulation of IFN-β production in mouse macrophages, and its underlying mechanisms. By stimulation of Raw264.7 macrophages with LPS, we found a dose-dependent increase in IFN-β mRNA expression and the IFN-β mRNA level reached maximum at 3 h after LPS treatment. The PDE4 inhibitor rolipram effectively suppressed the IFN-β expression with the IC50 of approximately 0.115 µM. Such inhibiting effect of rolipram was mediated by activating cAMP signaling because the cAMP analog dibutyryl-cAMP produced the same inhibitory effect in Raw264.7 cells. In addition, the Epac inhibitor ESI-09 did not reverse the inhibitory effect of rolipram, while the PKA inhibitor Rp-8-CPT-cAMP reversed the inhibition by approximately 57 %, suggesting that the inhibitory effect of rolipram on the IFN-β expression was mediated by activation of the cAMP/PKA, but not cAMP/Epac signal pathway. Moreover, the PKA activator 6-Bnz-cAMP significantly decreased the LPS-induced IFN-β expression at the level similar to that of rolipram. Using the peritoneal macrophages isolated from PDE4 null mice and the corresponding wild-type mice, we further found that the LPS-induced IFN-β expression in PDE4B-/- macrophages, but not PDE4A-/- or PDE4D-/- macrophages, was significantly decreased, and the level of expression was similar to that of PDE4B+/+ macrophages treated with rolipram. Additionally, rolipram did not further decrease the IFN- expression in PDE4B-/- macrophages. Contrarily, PDE4A-/- and PDE4D-/- macrophages, like their wild-type macrophages, exhibited significant reduction in IFN-β expression in the presence of rolipram. These results demonstrated that the rolipram effect on the IFN-β expression was mediated by inhibition of PDE4B. To explore whether inhibition of IFN-β expression by rolipram acts through regulation of TRIF-dependent signal pathway, the activation of the transcription factor IRF3 and its upstream factor TBK1 was monitored by western blotting. The results showed that LPS-induced IRF3 phosphorylation in Raw264.7 cells was elevated with time until 3 h, but was not altered by rolipram until 5 h after LPS stimulation. LPS-induced TBK1 phosphorylation was unchanged by rolipram up to 5 h of LPS stimulateion. Collectively, our data demonstrated that inhibition of PDE4B is sufficient to block LPS-induced IFN-β expression via activation of cAMP/PKA signal pathway in mouse macrophages but not by regulation of TLR4/TRIF/TBK1/IRF3 signal pathway. | en_US |