Mechanism Of Estrogen Regulating PMNs Function Through AMPK/mTOR Signaling Pathway

Neutrophils(polymorphonuclear neutrophils,PMNs)are an important part of the innate immune system.Increased susceptibility to inflammatory diseases in the mammary gland and uterus during perinatal period is closely related to decreased function of PMNs.Peripheral estrogen(estrogen,E)of bovine plays an important role in the regulation of PMNs function.As an important liposoluble steroid hormone,the cellular regulatory effect of E is mainly achieved through slow genomic pathway and rapid non-genomic pathway mediated by estrogen receptor(estrogen receptors,ERs).Changed in the immune PMNs function is always accompanied by cell energy metabolism,and AMPK/mTOR is the key signaling pathway in the regulation network of intracellular energy metabolism.The purpose of this study was to explain the signaling mechanism between the state of intracellular energy metabolism and the level of E in the process of regulating the immune function of PMNs,and to provide theoretical basis for evaluating the direct correlation between peripheral E and immune function of PMNs in perinatal bovine.The venous blood of bovine was collected to separate PMNs,and induced by LPS,E and ICI in vitro for 0h,1h,2h,4h and 8h.The expression of ER?,ER? and GPR30,the key signals of immunoactive signal pathway TLR2,TLR4,MyD88,NF-?B,the protein expression and phosphorylation of AMPK/mTOR,the key signals of cell energy metabolism,were detected by qPCR and Western Blot.The results showed that low concentration of E significantly inhibited the expression of TLR2,TLR4,MyD88 and NF-?B(p<0.05),and significantly down-regulated the expression of mTOR and phosphorylation(p<0.01).No effect on AMPK expression.High concentration of E significantly promoted the expression of TLR2,TLR4,MyD88 and NF-?B(p<0.05),activated AMPK/mTOR signaling pathway,promoted energy metabolism and enhanced cell viability.The results showed that PMNs of bovine expressed ER?,ER? and GPR30,in a concentration-dependent and time-dependent manner in the regulation of PMNs immune function mediated by E.At the same time,low concentration E decreased PMNs activity by inhibiting the activation of AMPK/mTOR pathway,and high concentration E enhanced PMNs activity by activating AMPK/mTOR pathway.