Components From Tityus discrepans Scorpion Venom With Fibrino(geno)lytic Activity; Plasmin And Factor Xa Inhibitory Activity.

^{1Brazón J., 2Guerrero B., 1D´Suze G., 1Sevcik C.} and ²Arocha-Piñango C.L.

1 Lab. Neurofarmacología Celular. CBB. 2 Lab. Fisiopatología. CME. IVIC. Apartado 21827 Caracas 1020A. Venezuela.

In vitro studies from our laboratory have demonstrated that Tityus discrepans (Td) scorpion venom is active on the coagulation and fibrinolysis system. Recent work using a S-2288 chromogenic substrate specific for tissue-type plasminogen activator (t-PA) demonstrated that this venom has a t-PA-like activity. The venom was also able to degrade plasminogen rich fibrin plates leaving a lysis-free center. Td venom incubated 7 h 1:30 with fibrinogen (Fg) degraded chain A, but was unable to clot fibrinogen; incubating for 2 h under the same conditions the thrombin/tris time was prolonged. The venom inhibited the amidolytic activity of plasmin (Plm) and factor Xa (FXa) on chromogenic substrates S-2251 and S-2222, respectively. The poison also had FXa-like activity but did not inhibit the thrombin amidolytic activity on chromogenic substrate S-2238. The aim of this work was to characterize the Td venom fractions responsible of these activities. Venom gel filtration chromatography (D'Suze et al, 1997) produced 5 fractions (Fr I, II, III, IV, V). Fraction I (100 µg) degraded fibrin in presence of plasminogen after incubation for 24 h at 37°C. This fraction had a t-PA-like activity of 0.82 U/mg, on substrate S-2288 (0.6 mM), this is ´12.5 the raw venom effect. Fr-I also presented an FXa-like activity of 0.25 U/mg on S-2222 (0.8 mM). Incubating 30:1 Fg:Fr-I, fibrinogen chains A (>2h) and B (>6h) were degraded; after 4 h, Fg degradation fragments ranging from 31 to 21.5 kDa were detected (tricine-SDS PAGE 7.5% gels under reduced conditions). Fr-I diminished Fg functionality after 2h incubation, prolonging the thrombin/tris time to >4 s. Fr-IV (100 µg) inhibited 50% of the FXa (3 nKat/ml) and Plm (3 nKat/ml) activity on S-2222 and S-2251 (0.8 mM) respectively, but Fr-V inhibited 80% of Plm activity on S-2251. Conclusions: Fr-I containing toxins having molecular weights >30 kDa showed a FXa- and t-PA-like and fibrino(geno)lytic activities. It is highly likely that the raw venom fibrinolytic activity is due to Fr-I’s t-PA-like activity. Fr-IV containing components with molecular weights 4 kDa, inhibited FXa activity.  Fr-V contains a plasmin inhibitor; this finding explains the lysis-free center observed on fibrin plates incubated with whole venom.

Support: FONACIT Grant S1-2001000908 to G.D’S.

Correspondence to: gdsuze@ivic.ve