The Study Of Vascular Endothelial Cells Activation In Antiphospholipid Syndrome

Antiphospholipid syndrome(APS) is a non-organ specific autoimmune disease manifested by recurrent fetal loss,arterial occusions,repeated venous thrombosis and thrombocytopenia.Obstetrical features presently include recurrent recurent spontaneous abortion,preeclampsia,intrauterine growth restriction,placental disfuction, premature labour,and much attention was payed to APS in obstetric field.Severe pregnancy complications may be attribute to thrombosis. Endothelial cell activation is the principal reason of thrombosis,and can be measured by expression of endothelial cell adhesion molecules. Several overseas studies have demonstrated that aPL antibodies activate endothelial cells(ECs) in vitro,as demonstrated by enhanced expression of adhesion molecules,but domestic studies is fewer.Binding of APA to these cells has been assumed to result from its interaction with membrane phospholipids.β₂-GP1has been found to be an important autoantigen in the antiphospholipid antibody syndrome.It is now generally accepted that most antiphospholipid antibodies recognizeβ2GP I bound to the cardiolipin-coated microplates used in clinical"anticardiolipin"assays。there is said to be a receptor ofβ₂-GP1 on endothelial cells surface.Recent overseas studies indicated e that annexin A2 mediates the binding ofβ₂-GP1 to endothelial cells,annexin A2 is the receptor ofβ₂-GP1on endothelial cells surface for Endothelial cell activation.There are three part in the paper.The first part was about investigating the effects of umbilical vein endothelial cells activation by the serum of patients with antiphospholipid syndrome.HUVECs were cultured in vitro,and treated with APS serum,and adhesion molecule ICAM-1å’ŒE-selectin mRNA was measured by real-time fluorescent quantitative-polymerase chain reaction and its antigen by flow cytometry.the second part was about investigating the effects of Annexin A2 mediates endothelial cell activation in antiphospholipid syndrome. We found anti-annexin A2 antibodies cause endothelial cell Activation viia non-Fc receptor pathway.These observations suggest a novel pathway for endothelial activation in APS that is initiated by crosslinking or clustering of annexin A2 on the endothelial surface. anti-annexin A2 monomeric Fab can block endothelial activation, treat by annexin A2 RNA interference also can block endothelial activation.the third part was about investigating whether fluvastatin interfering with ICAM-1 expression of HUVECs treated by APS serum,so that blocked endothelial cell activation in APS.we may provide a rationale for using statins as a therapeutic tool in treatment of APS. Objective:To investigate the effects on expression of ICAM-1 and E-selectin in vascular endothelial cells induced by by the serum of patients with antiphospholipid syndromeMethods:The serum was collected from 10 patients with APS and 10 normal controls,Human umbilical vein endothelial cells(HUVEC) were treated with the two kind of serum respectively.Adhesion molecule ICAM-1å’ŒE-selectin mRNA was measured by real-time fluorescent quantitative-polymerase chain reaction and its antigen by flow cytometry.Results:APS serum significantly increased the expressions of ICAM-land E-selectin on HUVEC-12 were from mRNA levels and protein level,compared with control serum(P＜0.05).Conclusious:umbilical vein endothelial cells can be activated by APS serum,and increased expression of adhesion molecules. Chapter 2Annexin A2 mediates endothelial cell activation in antiphospholipid syndromeObjective:To investigate the effects of Annexin A2 mediates endothelial cell activation in antiphospholipid syndrome.Methods:Human umbilical vein endothelial cell(HUVEC) were treated with anti-annexin A2 antibodies and anti-annexin A2 fragments F(ab')2,F(ab),and adhesion molecule ICAM-1 mRNA of HUVEC was measured by real-time fluorescent quantitative-polymerase chain reaction and its protein by flow cytometry.Then HUVEC in APS serum group was pretreated with anti-annexin A2 F(ab) or Annexin A2 siRNA,and adhesion molecule ICAM-1mRNA of HUVEC was measured by real-time fluorescent quantitative-polymerase chain reaction and its protein by flow cytometry.Results:Incubation of HUVEC with anti-annexin A2 antibodies and bivalent anti-annexin A2 F(ab₎2 fragments both led to a higher ICAM-1 expression in mRNA and protein,but incubation of HUVEC with monomeric Fab couldn't do it. HUVEC p retreated with anti-annexin A2 Fab significantly reduced the ICAM-1 response to anti-annexin A2 antibodies or APS serum.HUVEC pretreated with Annexin A2 RNA interference also significantly reduced the ICAM-1 response to APS serum.Conclusious:Anti-annexin A2 antibodies cause endothelial cell activation via non-Fc receptor pathway.These observations suggest a novel pathway for endothelial activation in APS that is initiated by crosslinking or clustering of annexin A2 on the endothelial surface.Anti-annexin A2 monomeric Fab can block endothelial activation in APS,and annexin A2 RNA interference can also do it. Objective:The present study was undertaken to investigate whether fluvastatin interferes with endothelial cell activation induced by the serum of patients with antiphospholipid syndrome.Methods:human umbilical vein endothelial cells were treated with serum from APS patients or normal controls or with medium alone,cells were pretreated with fluvastatin(0,2.5,5 and 10μmol/L)before treated with APS serum.Adhesion molecule ICAM-1mRNA was measured by real-time fluorescent quantitative -polymerase chain reaction and its antigen by flow cytometry.Results:Incubation of EC with APS serum led to a higher ICAM-1 expression compared with incubation with control serum,fluvastatin treatment of the cells significantly reduced the ICAM-1 response to APS serum,and this effect was dose dependent(P＜0.05).Conclusions:fluvastatin exerts a protective effect towards APS serum at the EC level by decreasing leukocyte adhesion molecule expression on endothelial cells.our data provide a rationale for using statins as a therapeutic tool in treatment of APS.