Study On Melatonin’s Protective Effect With Mechanism On The Inflammation Model Of New-born Mice Brain Slices Induced By Lipopolysaccharide

ObjectiveAn inflammation model of mouse brain slice is established to study the melatonin’s protective effect and mechanism through melatonin receptors on the central nervous system inflammation induced by Lipopolysaccharide(LPS).The study provides a theoretical basis for further exploring melatonin’s effecting mechanism on central nervous system inflammation and related diseases.MethodsBrain slices were prepared from 10-day-old mice and divided into four experiment groups: CTRL group,LPS group(24h LPS treatment),LPS+MEL group(6h MEL pretreatment,24 h LPS treatment),and MEL group(6h MEL treatment).Enzyme-linked immunosorbent method was used to detect the content of inflammatory factor IL-6 in brain slices,and protein imprinting method was used to detect the expression levels of microglial-specific proteins CD11 b,melatonin receptor MT1,and MT2 in brain slices.In order to further study and verify this experiment,2-phenyl-N-acetyltryptamine(Luzindole,LUZ),an antagonist of melatonin receptor MT1 and MT2,was added to the brain slice medium co-treated by LPS and MEL.The experiment was further divided into:CTRL group,LPS group(24h LPS treatment),LPS+MEL group(6h MELpretreatment and 24 h LPS treatment),MEL group(6h MEL treatment),LPS+MEL+LUZ group(6h MEL & LUZ co-pretreatment and 24 h LPS treatment).Enzyme-linked immunosorbent method was used to detect the contents of inflammatory factors IL-1β and IL-10 in brain slices,and protein imprinting method was used to detect the expression levels of MT1 and MT2 in brain slices.Results1.Using LPS to induce in vitro cultured brain slices and create a mouse brain inflammation model,the releasing of inflammatory factors and expression levels of CD11 b protein increased after 24 hours.After melatonin pretreatment,the releasing of IL-6 and expression level of CD11 b protein both decreased,meanwhile LPS also reduced the protein expression levels of melatonin receptors MT1 and MT2.2.After adding LUZ,the antagonist of melatonin receptor MT1 and MT2,the protein expression level of MT1 reduced significantly,and the levels of inflammatory factors IL-1β and IL-10 increased when compared to the CTRL group and the LPS+MEL group,which further proved that melatonin reduces the neuro-inflammatory response caused by LPS through MT1.Conclusion1.Mouse organ-type brain inflammation was successfully caused by inducing in vitro cultured brain slices with LPS.2.Melatonin can reduce LPS-induced neuro-inflammatory response.3.Melatonin may protect LPS-mediated neuro-inflammatory response through the melatonin receptor MT1.