title: On the molecular mechanisms for the highly procoagulant pattern of C6 glioma cells.
abstract: BACKGROUND
  That there is a correlation between cancer and procoagulant states is well-known. C6 glioma cell line was originally induced in random-bred Wistar-Furth rats and is morphologically similar to glioblastoma multiforme, the most common aggressive glioma resistant to therapeutic interventions.
  
  
  OBJECTIVE
  In this study we analyzed the molecular mechanisms responsible for the highly procoagulant properties of C6 glioma cells.
  
  
  METHODS
  The presence of tissue factor (TF) and phosphatidylserine (PS) in C6 cells was investigated by flow-cytometric and functional analyses. The assembly of extrinsic tenase, intrinsic tenase and prothrombinase complexes on these cells was studied using enzymatic assays employing plasma or purified proteins.
  
  
  RESULTS
  TF was identified by flow-cytometric and functional [factor (F) Xa formation in the presence of cells and FVIIa] assays. Alternatively, conversion of FX into FXa was also observed in the presence of C6 cells, FIXa and FVIIIa. This effect was both cell- and FVIIIa-dependent, being consistent with formation of the intrinsic tenase complex. C6 cells were also able to activate prothrombin in the presence of FXa and FVa, thus supporting formation of the prothrombinase complex. This ability was similar to positive controls performed with PS-containing vesicles. Accordingly, exposure of PS on C6 cells was demonstrated by flow cytometry employing specific anti-PS antibodies. In addition, annexin V, which blocks PS binding sites, inhibited FX and prothrombin conversion by their respective C6-assembled activating complexes.
  
  
  CONCLUSIONS
  C6 glioma cells support all procoagulant reactions leading to robust thrombin formation. This ability results from concomitant TF exposure and from the presence of the anionic lipid PS at the outer leaflet of cell membrane. Therefore, this animal cell line may be used to explore new aspects concerning the role of blood coagulation proteins in tumor biology, especially those affecting the central nervous system.
authors.affiliation: Instituto de Bioquímica Médica, Centro de Ciências da Saúde, Universidade Federal do Rio de Janeiro, Avenida Bauhínia 400, Cidade Universitária, Ilha do Fundão, Rio de Janeiro 21941-590, Brazil.
authors.forename: R S
authors.initials: RS
authors.lastname: Fernandes
authors.forename: C
authors.initials: C
authors.lastname: Kirszberg
authors.forename: V M
authors.initials: VM
authors.lastname: Rumjanek
authors.forename: R Q
authors.initials: RQ
authors.lastname: Monteiro
mesh_terms: D000818:Animals; D001777:Blood Coagulation; D001779:Blood Coagulation Factors / Q000378:metabolism; D045744:Cell Line, Tumor; D003546:Cysteine Endopeptidases / Q000378:metabolism; D005434:Flow Cytometry; D005910:Glioma / Q000737:chemistry / Q000473:pathology* / Q000503:physiopathology; D006801:Humans; D009363:Neoplasm Proteins / Q000378:metabolism; D010718:Phosphatidylserines / Q000032:analysis; D051381:Rats; D013917:Thrombin / Q000096:biosynthesis; D013925:Thromboplastin / Q000032:analysis / Q000378:metabolism
chemical_list: D001779:Blood Coagulation Factors; D009363:Neoplasm Proteins; D010718:Phosphatidylserines; D013925:Thromboplastin; D013917:Thrombin; D003546:Cysteine Endopeptidases; C030910:cancer procoagulant