Dopamine Via Dopamine D2-like Receptor Inhibits Inflammatory Response Induced By LPS In Mice Peritoneal Macrophages

Objective To investigate the regulatory effect and mechanism of dopamine and dopamine receptor on the inflammatory response of mice peritoneal macrophages induced by lipopolysaccharide（LPS）.Methods C57 mice were selected to collect mice peritoneal macrophages by thioglycolate broth peritoneal cavity lavage method.1 Mice peritoneal macrophages were pretreated with dopamine at concentrations(10^-6,10^-5,10^-4mol/L) and time（0.5,1,2 h）, then stimulated with LPS（1μg/ml） for 6 h. The tumor necrosis factor-α（TNF-α） in cell culture supernatant was detected by enzyme-linked immuno sorbent assay（ ELISA） and the expression of Toll-like receptor 4（TLR4） and pro-interleukin-1β（pro-IL-1β） were detected by Western-blot. The cell viability was detected via cell counting kit-8（CCK8）.2 Mice peritoneal macrophages were treated as following groups: control（ normal medium）、 LPS stimulation group（mice peritoneal macrophages were stimulated by 1μg/ml LPS for 6 h） 、 dopamine pretreatment group（mice peritoneal macrophages were pretreated with dopamine, then stimulated by 1μg/ml LPS for 6 h）、dopamine receptor antagonist group（mice peritoneal macrophages were pretreated with dopamine D1-like or D2-like receptor antagonist, then stimulated with dopamine and LPS）、dopamine receptor agonist group（mice peritoneal macrophages were pretreated with dopamine D1-like or D2-like receptor agonist, then stimulated by LPS）. The TNF-α in cell culture supernatant was detected by ELISA and the expression of TLR4 and pro-IL-1β were detected by Western-blot.3 TLR4^+/+ and TLR4^-/- mice peritoneal macrophages were treated according to the experiment design respectively : control（normal medium）、LPS stimulation group（mice peritoneal macrophages were stimulated by 1μg/ml LPS for 6 h）、dopamine pretreatment group（mice peritoneal macrophages were pretreated with dopamine, then stimulated by 1μg/ml LPS for 6 h）. The TNF-α in cell culture supernatant was detected by ELISA and the expression of TLR4 and pro-IL-1β were detected by Western-blot.Results1 ELISA and Western blot results showed, compared with LPS stimulation group, dopamine pretreatment（10-4 mol/L, 2 h）could markedly inhibit the expression of TNF-α, TLR4 and pro-IL-1β of mice peritoneal macrophages induced by LPS（P<0.05）. CCK8 results showed there’s no significance of cell viability between each group（P>0.05）.2 Compared with dopamine pretreatment group, D2-like receptor antagonist could significantly reduce the inhibitory effect of dopamine on expression of TNF-α, TLR4 and pro-IL-1β, while dopamine D1-like antagonist had no obvious inhibition（P>0.05）;conversely, compared with LPS stimulation group, dopamine D2-like receptor agonist could inhibit TNF-α expression（P<0.05）, and dopamine D1-like agonist showed no effect on TNF-α expression（P> 0.05）.3 Compared with TLR4^+/+ MPM LPS stimulation group, expression of TNF-α and pro-IL-1β of TLR4^+/+ dopamine pretreatment group decreased. While compared with TLR4^-/- MPM LPS stimulation group, TLR4^-/-dopamine pretreatment group showed no inhibition on expression of TNF-α and pro-IL-1β（P> 0.05）.Conclusion1 Dopamine inhibits mice peritoneal macrophages TNF-α secretion and pro-IL-1β expression induced by LPS,and the effect does not result from lowering cell viability.2 Dopamine via D2-like receptor inhibits the mice peritoneal macrophages inflammatory response induced by LPS.3 The inhibiton of dopamine on mouse peritoneal macrophages inflammatory response induced by LPS depends on TLR4.