Figure 5

Ghrelin inhibits inflammatory reactions by combining with GHSR1a and antagonizing TNF-α signaling. (A–C) Ghrelin greatly inhibited the secretion of inflammatory cytokines, such as IL-1β, IL-6, and iNOS, in vitro in a dose-dependent manner, as shown by real-time PCR. (D) Reduced nitrite levels were promoted by the application of ghrelin in a dose-dependent manner, as shown by the Griess experiment. (E,F) iNOS expression is downregulated in response to ghrelin application in a dose-dependent manner, as shown by western blotting. (G,H) IL-1β and IL-6 secretion were changed by ghrelin pretreatment and appeared to be heightened in proportion to elevations in ghrelin concentration, as shown by ELISA. (I–K) The mRNA expression of IL-1β, IL-6 and iNOS was enhanced by application of DLys, implying that DLys antagonizes the effect of ghrelin, as shown by real-time PCR. (L) Addition of DLys attenuates ghrelin’s effect of decreasing nitrite production, as detected by the Griess test. (M,N) Higher iNOS protein levels occurred with DLys application, as shown by western blotting. (O,P) Application of DLys inhibited ghrelin’s antagonizing effect on the secretion of inflammatory cytokines IL-1β and IL-6 induced by stimulation with TNF-α, as shown by ELISA. (*p < 0.005, **p < 0.05, ***p < 0.005, ^###p < 0.01 vs the control group). Scale bar: 100 μm.