Methanolic extract of Cariniana rubra Gardner ex Miers stem bark negatively regulate the leukocyte migration and TNF-α and up-regulate the annexin-A1 expression.

ABSTRACT

ETHNOPHARMACOLOGICAL RELEVANCE Cariniana rubra Gardner ex Miers (Lecythidaceae), is a native and endemic tree in Brazil, whose inner stem bark decoction preparation is used in folk medicine to treat various inflammatory disorders. Previous scientific reports confirmed its popular use as an anti-inflammatory, without, however, evaluating its action mechanisms. AIM: The objective of this study was to determine the cytotoxicity and anti-inflammatory mechanism of action of the methanolic extract of Cariniana rubra (MECr), using experimental models in vivo and in vitro, as well as to identify secondary metabolites present in the extract. MATERIAL AND METHODS: The MECr was prepared by maceration of inner stem bark powder in methanol (110 w/v). The in vitro cytotoxicity effect was evaluated in CHO-k1 cells. The Hippocratic screening test was conducted to evaluate the acute toxicity of MECr in mice. The actions of MECr on leukocyte migration, cytokine levels (IL-1ß and TNF-α) and annexin-A1 (AnxA1) expression, were carried out on lambda-type carrageenan air pouch inflammation model in Swiss mice. Additionally, the phytochemical analysis of MECr was carried out by thin-layer chromatography (TLC) and spectrometric mass analysis with electrospray ionization ESI(-)/MS and gas chromatography-mass spectrometry (GC-MS). RESULTS: Treatment of CHO-k1 cells for 24 h with MECr did not cause cytotoxicity (IC50 > 200 µg/mL), however, the MECr was shown to be cytotoxic after 72 h of cell exposure (IC50 = 19.90 ± 3.51 µg/mL). In the Hippocratic test, oral treatment of mice with 750, 1500, or 3000 mg/kg of MECr did not show any histopathological changes and mortality during the 14 days of observation. In the carrageenan air pouch inflammation model, MECr reduced (p < 0.001) polymorphonuclear migration (57.7% and 57.8%), leukocyte monocyte migration (74.5% and 61.8%) in the air pouch cavity and in the skin tissue, respectively. MECr also inhibited TNF-α concentration in the air cavity wash (3.2%, p < 0.01) and increased expression of the AnxA1 protein (26.9%, p < 0.01) in the skin tissue, particularly in neutrophils. ß-sitosterol (1.95%), gallic acid (1.24%), ß-amyrin (0.87%) and stigmasterol (0.66%) were identified as the major constituents in methanolic extract. CONCLUSION: MECr exhibits significant anti-inflammatory action at least by increasing AnxA1 expression and by inhibiting the release of TNF-α pro-inflammatory cytokine and leukocyte migration, which is probably linked to the presence of identified biologically active compounds, especially gallic acid and terpenes. We believe that the results of this study provide a pharmacological basis for the MECr to be considered as a potential therapeutic agent for the treatment of inflammatory diseases.