The SIRT1 Inhibitor EX-527 Suppresses MTOR Activation And Alleviates Acute Lung Injury In Mice With Endotoxiemia

Objective: Acute lung injury(ALI)with high morbidity and mortality is a common critical disease in clinic.It has great impact on quality of life affecting the patient's recovery.It is generally regarded that silencing information regulator 1(sirtuin1,SIRT1),a longevity factor in mammals,acts as a negative regulator of inflammation.A series of in vitro and in vivo experiments have shown that SIRT1 can down-regulate the transcription of inflammation-related genes by regulating the level of acetylation of histones,which has important implications for limiting inflammation and reducing inflammatory injury.However,there are also a few experiments found that SIRT1 has a positive effect on the expression of inflammatory genes and inflammatory lesions in tissues,and plays an important role in the development of inflammation.This study investigate the pathophysiological significance and the underlying mechanism of SIRT1 selective inhibitor EX-527 in the development of ALI induced by systemic lipopolysaccharide(LPS)exposure.Method: Experimental animal models were established using 7-8 weeks old specific pathogen free(SPF)grade male Balb/C mice.Intraperitoneal injection of LPS(20 mg/kg,dissolved in 0.9% physiological saline)induces systemic inflammation and ALI.To investigate the potential roles of SIRT1,the animals were received EX-527(10 mg/kg)administration 0.5 h prior to LPS challenge.In order to explore the underlying mechanisms,the mice received the mTOR activator 3BDO 0.5 h prior to EX-527 administration,and LPS was injected at 0.5 h later.Next,the animals were returned to their cages and provided with food and water ad libitum.The animals were anesthetized and sacrificed at 18 h after LPS challenge.The plasma samples were harvested for determining TNF-? and IL-6 by ELISA.The lung sections were stained with hematoxylin and eosin for histopathological evaluation under a light microscope.The myeloperoxidase(MPO)activity in the lung tissue were detected by MPO detection kit.The levels of TF and PAI-1 in lung tissues were detected by Western blot.The total protein level and phosphorylation level of mTOR downstream target protein 4E-BP1 were analyzed by Western blot.Results:(1)The LPS-challenged mice exhibited a significant increase in TNF-? and IL-6,which was significantly decreased after EX-527 administration.(2)Treatment with EX-527 attenuated LPS-induced lung histological abnormalities in lung tissue,which was accompanied with decreased myeloperoxidase level,which is widely regarded as a molecular marker of neutrophil infiltration.(3)The levels of TF and PAI-1 significantly increased in the LPS-treated group,but EX-527 pre-treatment could suppress these alterations.(4)Treatment with the SIRT1 inhibitor EX-527 significantly suppressed LPS-induced elevation of 4E-BP1 phosphorylation.(5)Co-treatment with 3BDO abolished the inhibitory effects of EX-527 on LPS-induced 4E-BP1 phosphorylation.Consistently,the suppressive effects of EX-527 on IL-6 induction and lung injury were also partially reversed by 3BDO.Conlusion: This study suggests that selective inhibition of SIRT1 by EX-527 might alleviate endotoxemia-associated acute lung injury partially via suppression of mTOR,which implies that SIRT1 selective inhibitors might have potential value for the pharmacological intervention of inflammatory lung injury.