Meeting Abstract
P1.69 Saturday, Jan. 4 15:30 Cholecystokinin inhibits inducible nitric oxide synthase expression in lipopolysaccharide-stimulated macrophages CAPELLARI CARNIO, E*; BRANCO, LGS; SAIA, RS; University of São Paulo; University of São Paulo; University of São Paulo carnioec@eerp.usp.br
Cholecystokinin (CCK) receptors are expressed in macrophages and are up-regulated by inflammatory stimulus. In vitro and in vivo studies have demonstrated the ability of CCK to decrease the production of various pro-inflammatory cytokines. This study investigates the role of CCK on iNOS expression in lipopolysaccharide (LPS)-activated peritoneal macrophages, as well as the intracellular signaling pathways involved in affecting iNOS synthesis. Thioglicollate-elicited macrophages were obtained by peritoneal lavage and cultured in RPMI 1640 medium, 10% fetal bovine serum and antibiotics. Nuclear p65, cAMP and iNOS levels were determined using ELISA kits, CCK receptors and IκBα expression by Western blot and nitrite by Griess method. Data was compared by one-way ANOVA and significant differences obtained using Tukey multiple variances post hoc test. CCK reduced NO production attenuating iNOS mRNA expression (15.49±10.80 vs. 113.16±0.23 AU; p<0.05) and protein formation. Furthermore, CCK inhibited the nuclear factor (NF)-κB pathway reducing IκBα degradation and minor p65-dependent translocation to the nucleus (543.78 ± 84.57% vs. 90.42 ± 9.13%, p<0.05). Moreover, CCK restored the intracellular cAMP content activating the cAMP-protein kinase A (PKA) pathway, which resulted in a negative modulatory role on iNOS expression and nitrite production. In peritoneal macrophages, the CCK-1R expression was predominant and up-regulated by LPS (0.61±0.08 vs. 0.30±0.09 AU; p<0.05). The pharmacological studies confirmed that CCK-1R subtype is the major receptor responsible in the biological effects of CCK. These data suggest an anti-inflammatory role for the peptide CCK in modulating iNOS-derived NO synthesis, possibly controlling the macrophage hyper-activation through NF-κB, cAMP-PKA and CCK-1R pathways.
