Bile acids, such as cholic acid (CA) and ursodeoxycholic acid (UDCA) have shown to decrease or increase the enzymatic activity of group IB pancreatic PLA(2), depending on the concentration used. Studies suggest that the inhibition of hydrolysis rate of the substrate is due to formation in aqueous phase of a complex between bile acid and PLA(2), which is catalytically inert. For this reason, we tested the inhibition of the enzymatic activity of group IIA snake venom PLA(2) by bile acids, using an aqueous phase model. In addition, we measured the ability of bile acids to inhibit the toxic effects caused by the mentioned toxin. UDCA and CA inhibited the enzymatic activity of the PLA(2) in a competitive mode. Moreover, these compounds inhibited myotoxic, cytotoxic and edema-forming activities induced by the toxin, but UDCA was more efficient than CA. It was demonstrated that bile acids interact directly with this protein by causing slight changes in the intrinsic fluorescence spectra. Preliminary molecular docking studies suggest that bile acids interact with amino acids at the active site of the PLA(2) through different interactions, CA showed hydrogen bonds with His48, whereas, UDCA displayed with Asp49. Results obtained herein may turn UDCA and CA into promising models for the development of new molecules with anti-inflammatory and anti-snake venom PLA(2) properties.
RESUMO:Veneno de cobra continua a ser um problema importante de saúde em muitos países da América Latina. Apesar dos avanços na terapia antiveneno, os efeitos locais causados por fosfolipases A 2 miotóxica (PLA 2 ) presentes no veneno, ainda persistem. Em busca de alternativas para antagonizar a atividade da PLA 2 do veneno de Bothrops asper, foram selecionados 36 extratos pertencentes a dezessete famílias de plantas vasculares e briófitas. Uma inibição significativa da atividade enzimática de PLA 2 presente no veneno de B. asper foi observada em onze extratos. Além disso, a atividade antioxidante dos extratos foi avaliada. Os resultados evidenciaram uma correlação estatisticamente significativa entre os extratos com ação inibitória contra a PLA 2 e aqueles com atividade antioxidante. Também, a quantidade de fenóis foi avaliada e foi encontrada uma relação entre a atividade biológica e a presença dessas substâncias. Nove extratos foram testados contra uma fração do veneno rico em PLA 2 básica (Fx-V B. asper), resultando em um efeito inibitório na atividade desta fração da PLA 2 na faixa de 30-80%. Esta atividade foi apoiada pela inibição que esses extratos apresentaram na citotoxicidade causada pelo Fx-V B. asper sobre mioblastos C2C12 de músculo esquelético de murino. Os resultados podem indicar a minimização dos esforços na busca de inibidores da PLA 2 , com foco nas amostras com propriedades antioxidantes conhecidas.Unitermos: fosfolipase A 2 , veneno de cobra, Bothrops asper, extratos de plantas, antioxidantes, DPPH.ABSTRACT: Snakebite continues to be a significant health problem in many countries of Latin America. Even though, there has been an improvement in the antivenom therapy, the local effects caused by myotoxic phospholipases A 2 (PLA 2 ) present in the venoms, still persist. In search for alternatives to antagonize the PLA 2 activity of Bothrops asper's venom, 36 extracts belonging to seventeen families of vascular plants and bryophytes were screened. A significant inhibition of the enzymatic activity of PLA 2 present in B. asper's whole venom was seen in eleven of these extracts. In addition, the antioxidant activity of all the extracts was evaluated. The results evidenced a significant statistical correlation between extracts with an inhibitory effect against PLA 2 and those with an antioxidant activity. Moreover, the amount of phenols was quantified finding a relationship between the bioactivity and the presence of these compounds. Nine extracts were screened against a fraction of the venom rich in basic PLA 2 (Fx-V B. asper), exhibiting an inhibitory effect on PLA 2 activity of this fraction in a range of 30-80%. This activity was supported by the inhibition that these extracts presented on the cytotoxicity caused by Fx-V B. asper on murine skeletal muscle C2C12 myoblasts. The results obtained, could point to minimize efforts in the search of PLA 2 inhibitors by focusing in samples with known antioxidant properties.
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