Activation of J77A.1 Macrophages by Three Phospholipases A2Isolated fromBothrops atroxSnake Venom

Juliana L Furtado,Anderson M Kayano,Sulamita Da S Setúbal,Juliana P Zuliani,Adriana S Pontes,Fabianne Lacouth-Silva,Andreimar M Soares,Kayena D Zaqueo,Caroline V Xavier,Leonardo A Calderon,Rodrigo G Stábeli,Beatriz F Lima,George A Oliveira

doi:10.1155/2014/683123

Abstract

In the present study, we investigated the in vitro effects of two basic myotoxic phospholipases A2 (PLA2), BaTX-I, a catalytically inactive Lys-49 variant, and BaTX-II, a catalytically active Asp-49, and of one acidic myotoxic PLA2, BaPLA2, a catalytically active Asp-49, isolated from Bothrops atrox snake venom, on the activation of J774A.1 macrophages. At noncytotoxic concentrations, the toxins did not affect the adhesion of the macrophages, nor their ability to detach. The data obtained showed that only BaTX-I stimulated complement receptor-mediated phagocytosis. However, BaTX-I, BaTX-II, and BaPLA2 induced the release of the superoxide anion by J774A.1 macrophages. Additionally, only BaTX-I raised the lysosomal volume of macrophages after 15 min of incubation. After 30 min, all the phospholipases increased this parameter, which was not observed within 60 min. Moreover, BaTX-I, BaTX-II, and BaPLA2 increased the number of lipid bodies on macrophages submitted to phagocytosis and not submitted to phagocytosis. However, BaTX-II and BaPLA2 induced the release of TNF-α by J774A.1 macrophages. Taken together, the data show that, despite differences in enzymatic activity, the three toxins induced inflammatory events and whether the enzyme is acidic or basic does not seem to contribute to these effects.

Highlights

Bothrops atrox is responsible for most cases of envenoming in the Northern region of Brazil [1]
The enzymatically inactive toxin BaTX-I did not induce any significant increase in the production of Tumor Necrosis Factor-α (TNF-α) in the studied period. Both acidic and basic Phospholipases A2 (PLA2) can be found in snake venoms in variable proportions depending on the snake species
All acidic PLA2s purified from Viperidae venoms present an Asp residue at position 49 [37, 38]

Summary

Introduction

Bothrops atrox is responsible for most cases of envenoming in the Northern region of Brazil [1]. Phospholipases A2 (PLA2; EC 3.1.1.4) belong to a diverse super family of lipolytic enzymes that catalyse the hydrolysis of the sn-2 ester bond of membrane glycerophospholipids to produce free fatty acids and lysophospholipids. These enzymes can differ in structure, molecular weight, substrate specificity, requirement for Ca2+, cell localization, and mechanism of action [2]. Snake venom PLA2s of the Viperidae family, belong to the subgroup IIA, which share structural features with PLA2s present in inflammatory exudates in mammals [7, 8] These snake venom PLA2s display a variety of pharmacological activities, such as myotoxic, neurotoxic, anticoagulant, hypotensive, hemolytic, platelet aggregation inhibiting, bactericidal, and proinflammatory activities [9,10,11,12]. Proteins of this subgroup can be further subdivided into two types: (i) catalytically active PLA2s, which have conserved residues at BioMed Research International the catalytic network and at the calcium-binding loop, including Asp, and (ii) catalytically inactive variants having a Lys instead of Asp at position 49 [10, 13, 14]

Methods

Results

Conclusion