The Expression Of Wnt5a And Syndecan-1 In Sepsis-induced Acute Lung Injury Of Mice

Objective: Sepsis is a critical illness in ICU,when patients missed the best timing of treatment,it can lead to anti-inflammatory and proinflammatory reaction out of control,human body may produce a large number of inflammatory factors,namely "inflammatory cascade",which is a serious threat to the safety of patients,In China,the mortality in severe sepsis is as high as 48.7%.Acute lung injury is a common complication of sepsis.Because there are many signal pathways involved in lung injury and there is a complex intersection between the various pathways,the current treatment of acute lung injury is not satisfactory,the prognosis is poor.It is very important to make early diagnosis of acute lung injury in clinic.The injury of vascular endothelial cells plays an important role in the pathogenesis of acute lung injury.The surface of vascular is covered with a layer of vascular endothelial cell glycocalyx,its core component is Syndecan-1,the component is vulnerable in serious infections(such as sepsis)and is easily to fall off,resulting in high permeability of lung vascular,more easily result in positive fluid balance,even lead to pulmonary edema and acute respiratory distress syndrome(Acute respiratory distress syndrome,ARDS).Wnt5a(wingless-related integration site-5a)represents one of the nonclassic protein in wnt signaling pathway,as Macrophage effector by combining different Frizzled5 receptor triggers different pathways,for instance,through activate GSK3? to regulate NF-?B which can induced the production of inflammatory factors.This paper intends to establish a mouse model of sepsis induced acute lung injury,to observe lung pathological changes and pulmonary vascular permeability changes,pathological changes of pulmonary vascular endothelial cells,observe serum markers of Wnt5 a and Syndecan-1 at the different time point,To study the changing trend of Wnt5 a and Syndecan-1 in sepsis induced acute lung injury(ALI)in the mouse model.Methods: The whole experiment process was divided into two parts,the first part:The model of sepsis-induced ALI was established by intraperitoneal injection LPS(lipopolysaccharide,serotype O55: B5).120 mice were randomly divided into injured group and control group,The mice in the injured group were injected with 10 mg / kg LPS,And the control group received the same volume of saline.A total of 120 mice were divided into 6 time points: 1h,3h,5h,8h,12 h,24h,and so on.10 rats in each group,The mice were sacrificed by anesthesiaat at every time point.Which used to detect the Pathological changes of lung tissue and the expression of Wnt5 a and Syndecan-1.We kept the blood supernatant after taking the whole blood.the left lung tissues were taken for hematoxylin-eosin staining(HE).The pathological changes of lung tissues were observed under light microscope.We took middle lobe of right lung to observe the structure of pulmonary vascular endothelial cells under electron microscope.Extracted protein of superior lobe of right lung.The expression of Wnt5 a protein was detected by Western blotting.we extracted m RNA of Inferior lobe of right lung,then the expression of Syndecan-1 gene was detected by Real-time PCR.the trend of Wnt5 a and Syndecan-1 in serum were observed by ELISA.The second part:we used the same method to prepare sepsis model.100 mice were randomly divided into injured group and control group.which used to study the change of pulmonary vascular permeability,they were divided into 5 time points: 1h,3h,5h,8h,12 h,10mice in each group.The changes of pulmonary vascular permeability were observed by tail intravenous injection of Evans blue dye.Results: The normal group of pulmonary alveolar wall is complete,the alveolar wall is thin,there were no red blood cells and inflammatory cells exudation in alveolar,without alveolar interstitial veins stasis;after giving lipopolysaccharide(LPS),the degree of lung injury aggravated gradually as time goes by.In the injured group,the alveolar wall of the lung tissue was significantly thickened,a large number of red blood cells and inflammatory cells exudation in the alveolar cavity,some of the alveolar cavities formed hyaline membrane.We observed vascular endothelial cells spacing broadening under TEM.Also we found mitochondria appered different degrees of edema,vascular basement membrane and endothelial cells showed different degrees of separation,there were more lipid vacuoles in matrix.Compared with the control group at the same time,the serum concentration of Wnt5 a in the injury group was gradually increased in 3h,it reached the peak at 5h-8h,then decreased gradually after the injury of about 12 h.The injury groups at 24 h were not obvious compared with control group.The serum concentration of Syndecan-1 maintain a high concentration in 3h after giving lipopolysaccharide(LPS),then gradually decreased.after 8h increased slightly,compared with the control group,in24 h it maintained a high concentration.The level of Wnt5 a protein in serum was detected by Western blotting,the expression of Wnt5 a was increased early in5h-8h.The highest expression level of Syndecan-1 was at 3h.The concentration of Evans blue dye in formamide were increased with the severity of lung injury,lung injury degree reached a peak between 5h-8h.Conclusion: we first discussed the expression and trend of Wnt5 a in acute lung injury mice model.we detected the concentration of Syndecan-1 and Evans blue dye in formamide to evaluate evaluating the injury of pulmonary vascular endothelium,we detected the concentration of these two markers to appraise the severity of pathological changes of lung tissue.In this study,we found that the injury of endothelial cells and the shedding of syndecan-1 apperas in the early stage of acute lung injury.Both of them may be used as early biological markers of sepsis-induced acute lung injury.