title: Hemorrhagic activity and mechanism of FIIa, a fibrinolytic enzyme from Agkistrodon acutus venom.
abstract: OBJECTIVE
  To study the local hemorrhagic activity of a fibrinolytic enzyme (FII(a)) from Agkistrodon acutus venom and its mechanism.
  
  
  METHODS
  The local hemorrhagic activity was determined by subcutaneous injection on the back of mouse. The effects of FIIa on factor X, prothrombin, gelatin, and collagen were visualized by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Platelet aggregation assays were performed in rat platelet-rich plasma (PRP). Human umbilical vein endothelial cells (HUVEC) were cultured and passaged in complete M199 medium. Cell viability and nuclear morphology change were determined by fluorescein diacetate (FDA) staining and Hoechst 33258 staining, respectively.
  
  
  RESULTS
  The minimum hemorrhagic dose (MHD) of FIIa was 89 microg. In vitro, FII(a) (0.25 g/L) degraded factor X, prothrombin, collagen, and gelatin, and dose-dependently (0.25, 0.50, 0.75, and 1.00 g/L) inhibited the platelet aggregation induced by ADP in rat PRP. When HUVEC in culture treated with FII(a), HUVEC showed detachment in a dose-dependent manner, but no apoptosis sign was observed.
  
  
  CONCLUSIONS
  FIIa had local hemorrhagic activity, and the mechanism was related to the degradation of factor X, prothrombin, gelatin, and collagen, the inhibition of ADP-induced platelet aggregation, and inducement of HUVEC detachment.
authors.affiliation: Department of Pharmacology, Zhongshan Medical College, Sun Yat-Sen University, Guangzhou 510080, China.
authors.forename: Qing-qing
authors.initials: QQ
authors.lastname: Wang
authors.forename: Jia-shu
authors.initials: JS
authors.lastname: Chen
authors.forename: Xiu-xia
authors.initials: XX
authors.lastname: Liang
authors.forename: Peng-xin
authors.initials: PX
authors.lastname: Qiu
authors.forename: Yi-wen
authors.initials: YW
authors.lastname: Wang
authors.forename: Guang-mei
authors.initials: GM
authors.lastname: Yan
mesh_terms: D017836:Agkistrodon; D000818:Animals; D002478:Cells, Cultured; D003094:Collagen / Q000378:metabolism; D003435:Crotalid Venoms / Q000737:chemistry* / Q000302:isolation & purification / Q000494:pharmacology*; D042783:Endothelial Cells / Q000166:cytology / Q000187:drug effects*; D005170:Factor X / Q000378:metabolism; D005260:Female; D005343:Fibrinolytic Agents / Q000494:pharmacology*; D006801:Humans; D008297:Male; D008666:Metalloendopeptidases; D051379:Mice; D010974:Platelet Aggregation / Q000187:drug effects; D011516:Prothrombin / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D014471:Umbilical Veins / Q000166:cytology
chemical_list: D003435:Crotalid Venoms; D005343:Fibrinolytic Agents; D011516:Prothrombin; D005170:Factor X; D003094:Collagen; C478420:FIIa; D008666:Metalloendopeptidases