Unfractionated Heparin Ameliorated Pulmonary Glycocalyx Shedding-a Study On The Involvement Of Angpt-2/HPA/MMP-9 Pathway

Objective:Histones,as important mediator in sepsis,can cause the shedding of glycocalyx and dysfunction of the vascular endothelium.Unfractionated heparin(UFH)is known to protect the glycocalyx and thereby inhibit coagulation activation.The exact mechanism is still unknown.Therefore,the aim of this study is to investigate the involvement of angiopoietin(Angpt)-2/heparinase(HPA)/matrix metalloproteinase(MMP)-9 pathway in shedding of glycocalyx.Methods:Mice were divided into three groups: control group,histone group,and histone+UFH group.Histone was injected through the tail vein at a dose of 50 mg/kg.UFH at a dose of 400 U/kg was injected through the tail vein 1h after histone.The control group was given the same volume of saline.Mice were anesthetized and the lung was collected for detecting wet/dry ratio(W/D),lung water content(%),and hematoxylin and eosin(H&E)staining after 4 hours.Angpt-1,Angpt-2,HPA,MMP-9,syndecan-1(SDC-1),tissue factor(TF)and fibrinogen(FIB)m RNA were detected by quantitative real-time polymerase chain reaction(qRT-PCR).Angpt-2,HPA,MMP-9,SDC-1,TF and FIB proteins in lung tissue were detected by western blot.Results:1.H&E staining showed that it is obviously that in histone group(P<0.001 vs control group)the disrupted alveolar architecture,edema,infiltration of inflammatory cells,hemorrhage and thrombosis were observed,and it is alleviated in histone+UFH group(P<0.001).2.W/D ratio and lung water content(%)indicated that histone group showed pulmonary edema compared with the control group.W/D ratio(P<0.001)and the lung water content(%)of the histone+UFH group(P<0.001)were significantly lower than the histone group.3.The results of qRT-PCR showed that histone decreased the expression of SDC-1(P<0.001)and Angpt-1m RNA(P<0.01).Angpt-2(P<0.01),Angpt-2/Angpt-1(P<0.01),HPA(P<0.05)MMP-9(P<0.01),TF(P<0.001)and FIB(P<0.001)m RNA expression were significantly increased in the histone group.The results reversed after UFH treatment.4.Western blot showed that histone reduced the SDC-1 relative protein expression.Angpt-2(P<0.001),HPA(P<0.001),MMP-9(P<0.001),TF(P<0.001)and FIB(P < 0.001)relative protein expression were increased.There was a significant improvement after treatment with UFH as expected.Conclusion:We concluded that UFH can attenuate histone-induced disruption of pulmonary glycocalyx and activation of coagulation in mice possibly by Angpt-2/HPA/MMP-9 pathway.