Estudos comparativos da atividade anticoagulante de fosfolipases A2 isoladas da peçonha de Bothrops pirajai e inibidores diretos do fator X

Abstract

The Brazilian’s venomous snakes belongs to Viperidae and Elapidae families. The Viperidae family is responsible for the majority of snakebites. Bothrops pirajai belongs to the Viperidae family, this snake can be found in the south region of Bahia and in the northeast of Minas Gerais. The venom of snakes from the Viperidae family causes local effects and systemic manifestations associated with bleeding, edema, coagulopathies and hypovolemic shock. Among the various proteins that compose the snake venoms, phospholipases A2 (PLA2s) are responsible for several toxic and pharmacological effects. Some of these effects are cardiotoxicity, myotoxicity and pre and / or post-synaptic neurotoxicity. In addition, to these effects, we have anticoagulant, bactericidal, convulsive, edema inducing, hypotensive and hemolytic activities. Although commercial drugs are effective in preventing and treating thromboembolic disorders, they produce a variable anticoagulant response, requiring monitoring of clotting time and constant adjustment of the dose. This study aimed to evaluate in vitro the anticoagulant effect of three proteins from Bothrops pirajai snake venom and commercial anticoagulant drugs (directly inhibit coagulation X factor activated) on the coagulation factors of secondary hemostasis using citrated human plasma. Likewise, the in sílico structural analysis of the possible interactions between ophidian proteins and X factor activated that interfere in secondary hemostasis. The presence of Piratoxin-I (PrTX-I), as well as Piratoxin-II (PrTX-II), did not cause significant change in Prothrombin Time (TP), but increased Partial Thromboplastin Time activated (aPTT), while Piratoxin-III (PrTX-III) promoted the increase of both Prothrombin Time and Partial Thromboplastin Time activated in a directly proportional dose-dependent manner. PrTX-I and PrTX-II can be classified as weakly anticoagulant A2 phospholipases because they act in a non-specific way on factor X. In addition, they are classified as a PLA2, catalytically inactive (not acting on phospholipid hydrolysis). PrTX-III can be classified as a strongly anticoagulant phospholipases A2, due to the anticoagulant mechanism action of this Asp-49-PLA2 could involve the direct action of FXa, either by not allowing its activation and / or by direct inhibition of FXa, as nonactivation of factor X. As well as inhibiting the formation of the extrinsic tenase and / or prothrombinase complexes. In addition, the PrTX-III shows the activity of phospholipases A2 (acting in the hydrolysis of membrane phospholipids). The development of specific methodologies to assess the toxic effects caused by snakebites and their constituents makes it possible to propose new pharmaceutical products as well as biotechnological products. The three phospholipases A2 analyzed in this study act in secondary hemostasis and can be use for the design of new anticoagulant drugs that are more effective in the treatment of hemostatic disorders. Since these phospholipases A2 show versatility of action involving the coagulation factor X. Additionally, evaluative and descriptive studies on the biological effects triggered by components of snake venoms can be used in the development of complementary treatments to serotherapy and / or in the treatment of hemostatic disorders.