Effects of Bothrops atrox venom and two isolated toxins on the human complement system: Modulation of pathways and generation of anaphylatoxins

The complement system plays important biological roles, including the activation of inflammatory processes in response to the generation of proteolytic fragments of its components. Here we evaluated the effects of Bothrops atrox venom and two of its toxins (the P-I metalloprotease Batroxase and the acidic phospholipase A2 BatroxPLA2) on the human complement system, evaluating their effects on the classical (CP), lectin (LP) and alternative (AP) pathways, as well as on different complement components associated to the generation of anaphylatoxins. Primarily, the venom and both toxins modulated the hemolytic activity of the complement CP, with the venom and Batroxase reducing this activity and BatroxPLA2 increasing it. ELISA deposition assays indicated that B. atrox venom and Batroxase were also capable of modulating all three activation pathways (CP, LP and AP), reducing their activity after incubation with normal human serum (NHS), while BatroxPLA2 apparently only interfered with AP. Additionally, the venom and Batroxase, but not BatroxPLA2, promoted significant degradation of the components C3, C4, Factor B and C1-Inhibitor, as shown by Western blot and SDS-PAGE analyses, also generating anaphylatoxins C3a, C4a and C5a. Therefore, B. atrox venom and Batroxase were able to activate the complement system by direct proteolytic action on several components, generating anaphylatoxins and affecting the activation pathways, while BatroxPLA2 only interfered with the hemolysis induced by CP and the C3 deposition related to AP. Our results indicate that Batroxase and possibly other metalloproteases should be the main toxins in B. atrox venom to induce pronounced effects on the complement system.