DLL4 High Expression Promotes The Capillarization Of Liver Sinusoidal Endothelial Cell And Liver Fibrogenesis

Background and aims: Liver sinusoidal endothelial cells(LSECs)undergo capillarization preceding the onset of fibrosis,and recognized as the gate of liver fibrosis.DLL4,a ligand of the Notch signaling pathway,is predominantly expressed in endothelial cells and maintains liver sinusoidal homeostasis.We aim to explore the role of DLL4 in the capillarization of LSEC and liver fibrogenesis.To further reveal the mechanism of liver fibrosis and provide a new perspective of liver fibrosis treatment.Methods: DLL4 expression and distribution were assessed by immunohistochemistry(IHC)and immunofluorescence(IF)in HBV related liver cirrhotic patients and CCl4-induced murine liver fibrosis model.CD31,v WF,the markers of capillary endothelial cell,were dynamically observed by IHC;while fenestrae and basement membrane formation of LSECs were dynamically examined by scanning electron microscopy and transmission electron microscopy respectively in mice.DLL4 and the related Notch signal genes,basement membrane materials collagen type I,III,IV,laminin and fibronectin were measured by PT-PCR and western blott in LSECs.Mice with CCl4-induced liver fibrosis were treated with modified small interfering RNA targeting DLL4.Results: We found DLL4 was up-regulated in the sinusoids of human fibrotic liver and CCl4-induced murine liver fibrosis model,mainly in LSECs.Consistent with DLL4 high expression in CCl4 murine model,CD31 and v WF were also up regulated,meahwhlie fenestrae decrsaesed and basement membrane formed underlying LSECs.Compared with control group,DLL4,CD31,v WF,the related Notch sigal genes and basement membrane molecules collagen type I,III,IV,laminin and fibronectin were increased in LSECs isolated from 4 and 6-week CCl4-induced mice fibrotic liver.GO and pathway analysis confirmed DLL4 induced the production of basement membrane molecules in LSEC.DLL4 overexpression also increased the tension of sinusoid through endothelin-1 synthesis in LSECs and enhancing the coverage of hepatic stellate cell,in addition to injure the e NOS-s GC signaling.Hypoxia-inducible factor-1α caused the overexpression of DLL4 in LSECs during liver fibrosis.Finally,inhibiting DLL4 expression in vivo alleviated LSEC capillarization and CCl4-induced liver fibrosis.Conclusion: DLL4 mediating Notch signaling accelerated LSEC capillarization.DLL4 played an important role in the pathological sinusoidal remodeling during liver fibrogenesis.