Shedding Of Syndecan-4 From Vascular Endothelial Cells Impairs Endothelial Migration And Angiogenesis In DM

Background:Syndecan-4(synd4),a ubiquitous cell surface heparin sulfate proteoglycan,acted as an independent receptor for numerous cellular processes.In the study,we investigated whether synd4 shedded from vascular endothelial cells in diabetes mellitus(DM)and the role of synd4 in the pathogenesis of impaired angiogenesis.Methods:First,we detected the concentration of soluble synd4 in the plasma of fifty type 2 diabetic patients by enzyme-linked immunosorbent assay(ELISA).Then DM rats were induced by a high dosage of streptozotocin(STZ).We analyzed the change of soluble synd4 in the plasma once a week by ELISA and synd4 staining in endothelial cells of aorta.Human umbilical vein endothelial cells(HUVECs)were used to assess the mechanism of synd4 shedding.Advanced glycation end products(AGEs)were added to stimulate synd4 shedding.Anti-RAGE and NAC were used to inhibit the shedding of synde induced by AGEs.We use aortic ring assays and matrigel plug to evaluate the angiogenesis of db/db,synd4-/-,and wild type mice in vitro and vivo.Aortic rings were embedded in fibrin.After 6 days,immunofluorescence staining was performed using BS1 lectin-FITC.Matrigel plug premixed with bFGF was injected subcutaneously in db/db,synd4-/-,and wild type mice.After one week of the procedure,the level of angiogeneses was viewed by embedding in paraffin and staining using HE stain.Then we transfected the HUVECs with Lenti-synd4,and observed the migration of AGEs-elicited and synd4 knockdown HUVECs using a 2D-chemotaxis slide in a live cell system.Finally,we transfected the aortic rings of db/db mice and premixed the magrigel plug with Ad-synd4,trying to rescue the impaired angiogenesis in aortic ring and matrigel plug assays respectively.Results:Plasma concentration of synd4 of the diabetic patients were extremely low compared with the health.Soluble synd4 concentration of the DM rats was higher than that of the normal ones at the first week after onset of DM and declined to lower as times goes on.We also confirmed that synd4 on surface of endothelial cells of the aorta in rats was diminished 13 weeks after diabetes established.We found that synd4 shedded from HUVECs in the presence of 200?g/ml AGEs.When pretreatment with anti-RAGE antibody and NAC(a ROS inhibitor),the shedding was limited.Once synd4 was exhausted from endothelial cells,the angiogenesis was impaired.In both the matrigel plug assay and aortic ring assay,the neovasculerization was impaired in db/db and synd4-/-mice.The migration of AGEs-elicited and synd4 knockdown HUVECs was damaged in the live cell system.Finally,re-expressed synd4 could partly rescue the angiogenesis dysfunction in db/db mice.Conclusion:We have demonstrated that the shedding of synd4 from vascular endothelial cells played a key role in the diabetes-related impairment of angiogenesis and identified synd-4 as a potential therapeutic target for the vascular complications of diabetes mellitus.