Effects Of Simvastatin On The Autophagy Of LPS-induced Human Neutrophils And Release Of Inflammatory Mediators

Part one: Effects of simvastatin on the autophagy of human neutrophils induced by LPSObjective To investigate the effects of simvastatin on autophagy of neutrophils activated by LPSMethods Peripheral blood neutrophils(n = 10)from healthy adults were divided into five groups:Group?(Control group),group ?(LPS group),group ?(LPS + simvastatin group),group ?(LPS + simvastatin + 3-MA group)and group?(LPS + 3-MA group).group?was blank control group,group?,?,?,?were added lipopolysaccharide(final concentration of 1ug / ml),cultured 12 h,thencentrifugedandstorage each cell culture supernatant in a refrigerator at-80 ?,Group II was given equal volume of PBS,group?was added simvastatin(final concentration of6uM),group? wasaddedsimvastatin(Final concentration of 6uM)and 3-MA(final concentration of 5m M),group?wasadded3-MA(final concentration of 5uM)only,Cultured for 24 hours,then centrifugedandstorage each cell culture supernatant in a refrigerator at-80??the expression of beclin-1 mRNA was detected by RT-PCR,the expression of Beclin 1 and LC3-II protein was detected by Western Blot.Results Compared with group ?:the expression of Beclin 1 mRNA and protein?LC3-II protein expression in group ?????has no obvious difference(P > 0.05),and the expression of Beclin 1 mRNA and protein?LC3-II protein were increasedingroup?(P<0.05);Compared with group ?:the expression of Beclin 1 mRNA and protein?LC3-II protein expression in group?increased(P> 0.05),group?and?haveno significant differenceswithgroup?in the expression ofBeclin 1 mRNA and protein?LC3-II protein(P> 0.05).Compared with group ?,the expression of Beclin 1 mRNA and protein?LC3-II protein in group??weredecreased(P <0.05).Conclusion Simvastatin can enhance the autophagy activity of neutrophils induced by LPS.LPS does not significantly promote autophagy for neutrophils,and 3-MA can inhibit the autophagy effect of simvastatin.Part two: Effects of simvastatin on the inflammatory mediators released by LPS-induced human neutrophilsObjective To investigate the effects of simvastatin on inflammatory factors produced by LPS-activated neutrophils and to explore whether the anti-inflammatory effect of simvastatin is related to autophagy.Methods Take out the frozensupernatantfrom-80 ? refrigerator,room temperature melting.The concentrations of TNF-? and IL-6 in the supernatant were measured by ELISA.Results Compared with group ?,the levels of TNF-? and IL-6 in group ?,? and? were increased after treatment with LPS for 12 h(P <0.05),whilethe levels of TNF-?and IL-6 in group? hadno significantly difference(P> 0.05);Compared with group ?,the levels of TNF-? and IL-6 in group ?weredecreased(P <0.05),the levels of TNF-? and IL-6 in group ? and ? were not significantly different(P> 0.05);Compared with group?,the levels of TNF-? and IL-6 in group?????wereincreased(P <0.05).Conclusion LPS can promote neutrophils release of inflammatory mediators,simvastatin makes LPS activated neutrophils,release IL-6 and TNF-?.When Autophagy inhibitor 3-MA inhibitestheautophagy of neutrophils,the level of of IL-6andTNF?ingroup? hadno differencewithgroup ?,and use 3-MA processing LPS activation of neutrophils,also can be observed in the IL-6,TNF alpha level no difference with the group?,suggest autophagy can affect the release of inflammatory mediators and theanti-inflammatory effect of simvastatin,may be related to its increase of autophagy.