The authors declare that there is no conflict of interest that could be perceived as prejudicing the impartiality of the research reported.
Article first published online: 18 FEB 2013
Copyright © 2012 Wiley Periodicals, Inc.
Journal of Cellular Biochemistry
Volume 114, Issue 4, pages 844–853, April 2013
How to Cite
Chang, S.-Y., Kim, D.-B., Ryu, G. R., Ko, S.-H., Jeong, I.-K., Ahn, Y.-B., Jo, Y.-H. and Kim, M.-J. (2013), Exendin-4 inhibits iNOS expression at the protein level in LPS-stimulated Raw264.7 macrophage by the activation of cAMP/PKA pathway. J. Cell. Biochem., 114: 844–853. doi: 10.1002/jcb.24425
Seo-Yoon Chang and Dong-Bin Kim contributed equally to this work.
- Issue published online: 18 FEB 2013
- Article first published online: 18 FEB 2013
- Accepted manuscript online: 23 OCT 2012 11:57AM EST
- Manuscript Accepted: 8 OCT 2012
- Manuscript Received: 29 AUG 2012
- National Research Foundation of Korea. Grant Number: 5-2010-A0154-00139
- EXENDIN-4 (EX-4);
- Raw264.7 MACROPHAGE
Glucagon-like peptide-1 (GLP-1) and its potent agonists have been widely studied in pancreatic islet β-cells. However, GLP-1 receptors are present in many extrapancreatic tissues including macrophages, and thus GLP-1 may have diverse actions in these tissues and cells. Therefore, we examined the mechanism by which exendin-4 (EX-4), a potent GLP-1 receptor agonist, inhibits lipopolysaccharide (LPS)-induced iNOS expression in Raw264.7 macrophage cells. EX-4 significantly inhibited LPS-induced iNOS protein expression and nitrite production. However, Northern blot and promoter analyses demonstrated that EX-4 did not inhibit LPS-induced iNOS mRNA expression and iNOS promoter activity. Electrophoretic mobility shift assay (EMSA) showed that EX-4 did not alter the binding activity of NF-κB to the iNOS promoter. Consistent with the result of EMSA, LPS-induced IκBα phosphorylation and nuclear translocation of p65 were not inhibited by EX-4. Also, actinomycin D chase study and the promoter assay using the construct containing 3′-untranslated region of iNOS showed that EX-4 did not affect iNOS mRNA stability. Meanwhile, cycloheximide chase study demonstrated that EX-4 significantly accelerated iNOS protein degradation. The EX-4 inhibition of LPS-induced iNOS protein was significantly reversed by adenylate cyclase inhibitors (MDL-12330A and SQ 22536), a PKA inhibitor (H-89) and PKAα gene silencing. These findings suggest that EX-4 inhibited LPS-induced iNOS expression at protein level, but not at transcriptional mechanism of iNOS gene and this inhibitory effect of EX-4 was mainly dependent on cAMP/PKA system. J. Cell. Biochem. 114: 844–853, 2013. © 2012 Wiley Periodicals, Inc.