| BackgroundAcute lung injury(ALI) is one of the refractory complications which is associated with several clinical disorders,including severe sepsis,all kinds of shock,trauma,surgery and so on.Acute lung injury and its more severe form,the acute respiratory distress syndrome(ARDS),are generally refractory to clinical control and have a continuing high morbidity in critical ill patients.Pathogenic factors of ALI is many,but the common pathophysiology disorders are the diffuse injury of the alveolar epithelial cells,endothelial cells and pulmonary interstitial substance.The pathogenesis of ALI is complicated,activated neutrophils accumulated, inflammatory factor and media released,oxidative stress and apoptosis play a key role in lung tissue injury.SIRS is the root cause.So far,the therapeutic methods for ALI have not been improved. Respiratory support is still one of major methods for treating ALI which based on the treatment of primary disease.Pharmacological treatment especially inhaled anesthetic become a hot research topic at present. Sevoflurane is an inhaled anesthetic agent.Many researches have indicated that sevoflurane has a benefical protective function on lung tissue suffered from ALI.However,the mechanisms of action of sevoflurane on ALI have been not yet clarified.Heme oxygenase 1(HO-1) represents the inducible isoform of the microsomal HO family.HO-1 gene expression is up-regulated in response to a variety of stimuli and catalyzes the oxidation of heme to biliverdin-IXa,iron,and carbon monoxide.The HO system exerts major antioxidative,antiinflammatory,antiapoptotic,and vasodilatory characteristics.Many efforts have been made to modulate or preinduce HO-1 and to optimize its protective potency against subsequent noxious stimuli.However,most inducers of HO-1(e.g.,oxidative stress,hypoxia, hemin,cobalt chloride,radiation) have numerous side effects and therefore cannot be used in patients.Therefore,it is essential to identify substances or conditions that induce HO-1 without endangering the patient.We recently want to know that the volatile anesthetic sevoflurane is able to specifically up-regulate HO-1 gene expression or not.In the current study,we investigated the actions and mechanisms of sevoflurane post-treatment in ALI induced by LPS.ObjectiveThe present study was undertaken to assess the protective effects of sevoflurane on LPS-induced acute lung injury(ALI),investigate the influence of sevoflurane with different concentrations on tectology, oxygenation,inflammatory media and oxidative stress of the lung,then determine the conformable concentration.Determine whether the protective effects of sevoflurane on ALI is associated with HO-1.Methods1.A rat model of ALI was established according to document, healthy male SD rats anesthetized by intraperitoneal injection of 20%urethane 1g/kg which were tracheotomized and mechanically ventilated,then femoral vein injection of LPS at a dose of 5mg/kg.All groups were sacrificed respectively after 6 hours,lung biopsies were obtained to determine if the model of ALI successed.2.Forty-eight male SD rats anesthetized by intraperitoneal injection of 20%urethane 1g/kg which were tracheotomized and mechanically ventilated for 30 min were randomly divided into six groups,eight of each,group A(control group) were vena femoralis injected with 1ml/kg saline and continuous ventilated for 6 hours,group B(1.5MACSevo group) were vena femoralis injected 1ml/kg saline,2 hours later inhalated 1.5 MAC and continuous ventilated for 4 hours,group C (LPS+1.5MACSevo group) were vena femoralis injected LPS 5mg/kg,2 hours later inhalated 1.5 MAC and continuous ventilated for 4 hours,group D(LPS+1.0MACSevo group) were vena femoralis injected LPS 5mg/kg,2 hours later inhalated 1.0 MAC and continuous ventilated for 4 hours,group E(LPS+0.5MACSevo group) were vena femoralis injected LPS 5mg/kg,2 hours later inhalated 0.5 MAC and continuous ventilated for 4 hours,group F(LPS group) were vena femoralis injected with LPS 5mg/kg and continuous ventilated for 6 hours.Blood pressure was measured directly from artery during above processes mentioned, mean arterial pressure and arterial blood gas were both measured and recorded at three time points T0(LPS or saline injection rightly before), T1(2 hours after injection),T2(6hours after injection).All groups were sacrificed respectively by losing blood at T2 time point.The dry/wet (D/W) ratio,the activity of superoxide dismutase(SOD) myeloperoxidase (MPO),and malondialdehyde MDA contents in lung tissues were determined and lung biopsies were also obtained.The expression of ICAM-1,CINC-1 mRNA and protein were detected in lung tissue.3.Forty-eight male SD rats anesthetized by intraperitoneal injection of 20%urethane 1g/kg which were tracheotomized and mechanically ventilated for 20 min were randomly divided into six groups,eight of each,group A(control group),group B(ZnPP group),group C(1.0MACSevo group),group D(LPS+1.0MACSevo group),group E(ZnPP+LPS+1.0MACSevo group),group F(LPS group).rats in group B,E were vena femoralis injected 10mg/kg ZnPP,while 1mg/kg saline in other groups.Ten minutes later,rats in group D,E,F were vena femoralis injected LPS 5mg/kg,while 1mg/kg saline in other groups.Two hours later,1.0 MAC sevoflurane was inhalated for 4 hours continuously in group C,D,E.All groups were sacrificed respectively by losing blood. The dry/wet(D/W) ratio,the activity of superoxide dismutase(SOD) myeloperoxidase(MPO),HO-1 and the contents of malondialdehyde MDA in lung tissues were determined and lung biopsies were also obtained.The expression of ICAM-1,CINC-1,and HO-1 protein was detected by western-blotting while the expression of ICAM-1,CINC-1, and HO-1 mRNA was detected by RT-PCR in lung tissues.4.Forty-eight male SD rats anesthetized by intraperitoneal injection of 20%urethane 1g/kg which were tracheotomized and mechanically ventilated for 20 min were randomly divided into six groups,eight of each,group A(control group),group B(1.0MACSevo group),group C(LY294002 group),group D(LPS+1.0MACSevo group),group E(LY294002+LPS+1.0MACSevo group),group F(LPS group).Rats in group C,E were vena femoralis injected 10mg/kg LY294002,while 1mg/kg saline in other groups.Ten minutes later,rats in group D,E,F were vena femoralis injected LPS 5mg/kg,while 1mg/kg saline in other groups.Two hours later,1.0 MAC sevoflurane was inhalated for 4 hours continuously in group B,D,E,but not in group A,C,F.All groups were sacrificed respectively by losing blood.The dry/wet(D/W) ratio,the activity of superoxide dismutase(SOD) myeloperoxidase(MPO),HO-1 and the contents of malondialdehyde MDA in lung tissues were determined and lung biopsies were also obtained.The expression of ICAM-1,CINC-1,HO-1,PI3K,piPA3K,Akt,piAkt protein was detected by western-blotting while the expression of ICAM-1,CINC-1,and HO-1 mRNA was detected by RT-PCR in lung tissues.Results1.Compared with control group,PaO2,MAP and oxygenation index decreased in group LPS despite at 2 or 6 h,the lung biopsies at 6h prove the model of ALI is successed.2.Compare to group A(control group),SOD activity was obvious decreased,while MPO activity and MDA content were increased in lung homogenates of group C(LPS+1.5MACSevo group),D(LPS+1.0MACSevo group),E(LPS+0.5MACSevo group),F(LPS group)(p＜0.05),moreover the expression of ICAM-1,CINC-1 mRNA and protein were increased in lung tissues of group C,D,E,F too(p＜0.05).Compared to group F,SOD activity was increased while MPO activity,MDA content and the expression of ICAM-1,CINC-1 mRNA and protein were decreased in group C,D,E(p＜0.05),especially in group C and D,the difference between group C and group D was not significant except MAP in T2.Group B(1.5MACSevo group) were not obvious different from group A except MAP.3.The activity of HO-1,expression of HO-1 mRNA and protein in group F(LPS group) were higher than group A(control group)(p＜0.05), while these in group D(LPS+1.0MACSevo group) were higher than group F(p＜0.05),the activity of HO-1,expression of HO-1 mRNA and protein in group E(ZnPP +LPS+1.0MACSevo group) were lower than group D(p＜0.05).HO-1 mRNA and protein in group B(ZnPP group),and group C(1.0MACSevo group) were not obvious different from group A (p＞0.05).4.The activity of HO-1,expression of HO-1 mRNA and protein, piPA3K,piAkt protein in group F(LPS group) were higher than group A(control group)(p＜0.05),while these in group D(LPS+1.0MACSevo group) were higher than group F(p＜0.05),the activity of HO-1, expression of HO-1 mRNA and protein piPA3K,piAkt protein in group E(LY294002+LPS+1.0MACSevo group) were lower than group D (p＜0.05),but the differences of pathologic change,SOD,MDA,MPO,the expression of ICAM-1 and CINC-1 were not statically significant(p＞0.05).Conclusion 1.Vena femoralis injected LPS 5mg/kg could induce ALI in rats. Oxidative stress and inflammatory reaction involved in pathogenesis of ALI.2.Sevoflurane post-treatment with different concentrations, especially 1.0MAC sevoflurane post-treatment could decrease oxidative stress and inflammatory reaction in ALI induced by LPS.3.The protective effect of sevoflurane post-treatment on LPS-induced ALI were produced by uptaking of HO-1 activity,HO-1 mRNA and protein expression.4.The phosphorylation of PI3K/AKT signal transduction pathway, the increase of HO-1 activity,expression of HO-1 mRNA and protein involved in acute lung injury induced by endotoxin.5.Effect of sevoflurane on protection of HO-1 on LPS-induced ALI was partly mediated by PI3K/AKT signal transduction pathway.
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