Discussion
Previous studies has shown the noticeable anti-inflammatory effects of NR4a1. In this study, we explored the mechanism of NR4a1 through bioinformatics analysis. We employed the GSE68167 dataset and selected 108 DEGs to be analyzed using DAVID and KEGG pathway and GO term analysis. BP analysis mainly shows signal transduction, positive regulation of chemokine production and inflammatory response. CC terms displayed enrichment of DEGs mainly in cytoplasm, membrane, and cytosol; while MF indicated the protein binding, calcium ion binding and callagen binding. KEGG showed the enrichment of DEGs mainly in cytokine-cytokine receptor, JAK-STAT signaling pathway and cAMP signaling pathway. We used STRING and Cytoscape to construct the gene network, where 49 DEGs were mapped and top Hub gene
IFNG, IL10, Fos, IL19, PDE4B, Cnr1, MMP13, NPY, Rtn1, UCHL1 were ranked by CytoHubba. Among these Hub genes, IFNG encodes IFNγ, IL10 and IFNγ are considered the important inflammatory cytokine and NR4a1 defects macrophage showed the decreased IL10 while the increased IFNγ production [10,33,34]. NR4a1 regulates the expression of early gene, including Fos in cancer [35]. Fos together with c-jun forms AP-1 plays a key role in inflammatory type macrophage [36]. UCHL1 has been reported to exert the anti-inflammatory effects via MAPK and NFκB signaling pathway in LPS stimulated macrophage [37]. Phosphodiesterase 4B(PDE4B), a member of the phosphodiesterase (PDEs) family, serving as a selectively hydrolytic enzyme of the second signal messenger cyclic adenosine monophosphate (cAMP) to play a crucial role in cAMP signaling pathway [15,25]. PDE4B, NPY and FOS were found to be enriched in cAMP signaling pathway, where the former is critical in signal transduction of inflammation [15,25,30]. The inhibitor of PDE4B has been shown exert an effective validity in treating various inflammatory diseases [15]. PED4B defected macrophage shows the decreased LPS induced NFκB activation and reactive oxygen species level [30,38].
Our data demonstrated that NR4a1 showed the effects of increasing cAMP and p-PKA levels through down-regulating PED4B expression. We also found that NR4a1 can inhibit LPS induced NF-κB signaling activation and the mRNA level of IL-6 and IL-1β by down-regulating PED4B expression. The anti-inflammatory function of NR4a1 has been proved to be related to restrict NFκB signaling pathway [33,39] Interestingly, cAMP activated PKA could effectively inhibit the NFκB regulated gene transcription [10,27].Thus, we hypothesized that NR4a1 may regulates the NFκB signaling pathway through PDE4B-cAMP-PKA signaling, which is also worthy to further detecting. In addition, bioinformatics analysis showed that the effect of NR4a1 in macrophage may be related to Fos, UCHL1, MMP13. JAK-STAT signaling pathway and cytokine-cytokine interaction may be regulated by NR4a1.