| Objective: Acute lung injury(ALI)is a life-threathening critical illness,in which acute respiratory distress syndrome(ARDS)is its severe form.The pathogenesis of ALI is complicated,and the mortality rate is high.The uncontrolled inflammatory reaction is the key mechanism underlying the development of ALI.AMP-activated protein kinase(AMPK)is an important metabolic regulator in the cell,but recent studies also found that AMPK has profound anti-inflammatory effects in vitro and vivo models.Metformin(MET),which is commonly used in clinic,exerts hypoglycemic and anti-inflammatory effects by activating AMPK.This study investigated the potential pharmacological effects of MET in ALI and the underlying mechanismscMethod: Male BALB/c mice were injected intraperitoneally with lipopolysaccharide(LPS)to induce ALI associated withsystemic infection.MET(400 mg/kg)was injected intraperitoneally 30 min before LPS injection.In order to investigate whether the pharmacological effect of MET is related to AMPK,AMPK inhibitor Compound C(15 mg/kg)was injected intraperitoneally 30 min before MET injection or AMPK activator A-769662(30 mg/kg)was used to replace MET and injected intraperitoneally 30 min before LPS injection.In order to explore whether AMPK play roles via activation of sirtuin 1(SIRT1)or inhibition of mammalian target of rapamycin complex 1(mTORC1),SIRT1 inhibitor EX527(10 mg/kg)or mTORC1 activator 3-BDO(100 mg/kg)was injected intraperitoneally 30 min before A-769662 injection.The mice were all decapitated 18 h after LPS injection,plasma and tissue were collected.The proinflammatory cytokines tumor necrosis factor-α(TNF-α)and Interleukin-6(IL-6)in tissue and plasma were detected by ELISA.HE staining was used to observe the morphological changes of lung tissue.The total protein and phosphorylation level of AMPK and mTORC1 downstream target protein P70S6K1 and 4EBP1 in lung tissues were detected by Western blot.Results:(1)MET treatment inhibited the expression of proinflammatory cytokines TNF-α and IL-6 in the plasma and lung tissues of mice induced by LPS,along with alleviating lung histological abnormalities.(2)MET treatment also increased the phosphorylation level of AMPK.The anti-inflammatory and protective effects of MET were blocked by the AMPK inhibitor Compound C.(3)Consistently,the AMPK activator A-769662 mimiced the anti-inflammatory and protective effect of MET in ALI,as evidenced by the reduced plasma level of IL-6 and the alleviated lung histological abnormalities.(4)The inhibitory effects of the AMPK activator A-769662 on the production of IL-6 could not be blocked by the SIRT1 inhibitor EX527,but the mTORC1 activator 3-BDO significantly blocked the inhibitory effect of A-769662 on IL-6 and lung tissue injury.(5)LPS exposure increased the phosphorylation of mTORC1 downstream target protein P70S6K1 and 4EBP1 in lung tissue,but MET inhibited P70S6K1 and 4EBP1 phosphorylation.The inhibitory effects of MET on P70S6K1 and 4EBP1 were blocked by the AMPK inhibitor Compound C.Conlusion: The experimental data of this study suggest that MET might have crucial anti-inflammatory and protective benefits in LPS-induced ALI model,and the underlying mechanisms involve the down-regulation of mTORC1 activity via activation of AMPK by MET.This study reveals a new mechanism of MET play an anti-inflammatory effect via AMPK,suggesting that MET may have potential application value in ALI. |
