Avaliação da citotoxicidade e da participação de citocinas e óxido nítrico na ação anti-inflamatória do extrato hidroetanólico de Macrosiphonia longiflora (Desf.) Mull. Arg.

Abstract

Macrosiphonia longiflora (A. St.-Hil.) Müll. Arg (Apocynaceae) popularly known as ‘velame’ and ‘velame branco’, is a native subshrub that grows in the Brazilian Cerrado. The plant is widely used in traditional medicine in the form of decoction and infusion, particularly as anti-inflammatory, depurative, anti-rheumatic, antisyphilitic and antiulcer agent. There is available information in the literature that has addressed the pharmacological activity of Macrosiphonia longiflora in experimental animals. This study aimed to evaluate the antiinflammatory pharmacological profile of the hydroethanolic extract of Macrosiphonia longiflora (Desf.), using in vivo and in vitro acute inflammation experimental models. As well, as investigate the cytokines and nitric oxide participation in its mechanism of action. Hydroethanolic (70%) extract of Macrosiphonia longiflora (HEMl) was prepared by maceration. The cytotoxic potential of HEMl in Chinese hamster ovary (CHO-K1) epithelial cell lines was evaluated using Alamar Blue method. Anti-inflammatory activity was evaluated with carrageenan and dextran –induced paw edemas and carrageenan-induced pleurisy in rats and lipopolysaccharide (LPS)-induced peritonitis in mice. Effects of HEMl on the inflammatory cytokines (IL-1β, IL-10, IL-17, INF-γ and TNF-α) concentrations in the pleural and peritoneal fluid and nitric oxide in the anti-inflammatory effects were assessed using ELISA kits and Griess method respectively. In the cytotoxicity study, IC50 of HEMl was 174.36 ± 8.35 mg / mL. It effectively inhibited (p<0.05) paw edema by carrageenan (3rd h), and by dextran (2nd h) after induction with the phlogistic agents. Furthermore, HEMl also significantly reduced exudates volume and leukocyte migration in the carrageenan-induced pleurisy and LPS-induced peritonitis, neutrophils counts in LPS-induced peritonitis. Evaluations of involvement of the inflammatory cytokines demonstrated significant decrease in the levels of IL-1β and IL-10 levels in the peritoneal lavage, IL-17 level was not affected in the peritonitis model, but rather in the pleurisy model. HEMl had no effect on the levels of tumor necrosis factor alpha (TNF-α) present in the lavages. The concentration of nitric oxide (NO) as assessed by measurement of nitrite (NO2 - ), showed that pretreatment with HEMl reduced significantly NO in the peritoneal and pleural lavage and in RAW 264.7 cells stimulated with LPS and interferon gamma (INF-γ). The results obtained in this study indicate lack of cytotoxic effects of HEMl coupled with its evident anti-inflammatory effects in both the in vivo and in vitro models of acute inflammation. The anti-inflammatory effect is partly related to inhibition of IL-1β, IL-10, IL-17 and nitric oxide releases and does not depend on the inhibition of TNF-α. The current study provided supportive evidence for the popular use of HEMl in the treatment of inflammatory conditions, and further shed more light on the possible roles of the inflammatory cytokines in its mechanisms of action as antiinflammatory agents.