Estudo do potencial anti-inflamatório e antiartrítico de Pterodon polygalaeflorus e dos seus mecanismos de ação

Abstract

The Pterodon genus belongs to the Fabaceae family and includes four native species of Brazil: P. emarginatus Vog, P. apparicioi Pedersoli, P. abruptus Benth. and P. polygalaeflorus Benth., the species studied in this work. Its fruits are used in folk medicine due to anti-rheumatic, analgesic and anti-inflammatory properties, among others. The aim of this study was to evaluate the anti-inflammatory, anti-arthritic and toxicology potential of the Pterodon polygalaeflorus specie, through the analysis of its effects on in vitro and in vivo models. The EEPpg, EHPpg e EDPpg extracts reduced (p <0.01) the in vitro NO production by macrophages activated by LPS, with low cytotoxicity, and decreased the cellularity (p<0.05) in the inflammatory exudate of the in vivo inflammation model known as air pouch. The most active extract (EHPpg) was selected and fractionated on silica gel 60 column generating four fractions. All fractions (Fr I-IV) reduced: NO production (p<0.001) by activated macrophages, with little or no cytotoxicity; in vitro (p<0.01, wound healing assay) and in vivo (p<0.05, thioglycollate induced-peritonitis) migration of macrophages; and proliferation of splenocytes stimulated with Con A (p<0.001). Fractions III and IV, the most active ones, showed anti-arthritic action (0.02 mg/kg), using the in vivo model of arthritis induced by Freund's complete adjuvant (AIA), as demonstrated by reduction of: the paw edema index in 23.7% (Fr III) and 43.95% (Fr IV); typical histopathological lesions on tibial-tarsal region of joints with AIA (p <0.05); the lymph nodes and spleen weight and cellularity, but not in bone marrow cellularity; in lymphocyte subpopulations (CD4+, CD8+ and CD19+) in the inguinal lymph nodes, but with a significant increase in CD4+CD69+ activated subpopulation, reduction of CD8+CD69+ cells and increased CD19+CD69+ (only Fr III); and reducing the population of macrophages in the spleen (p <0.001). Fractions III and IV showed immunosuppressive activities in the cellular and molecular level, decreasing (p <0.001) the iNOS mRNA levels, as well as the IL-1β, TNF-α and IL-10 cytokines at the mRNA and protein levels in macrophage culture. The expression of the CD14 receptor, by LPS stimulated macrophages (31.74%), was inhibited only by Fr III. The osteoclastogenesis was inhibited by the fractions III and IV, suggesting an anti-arthritic action for these fractions at level of cellular differentiation into osteoclasts (inhibition). This effect may result from inhibition of the NFATc1 transcription factor expression in the case of Fr III. Finally, Fr III and Fr IV did not present subacute toxicity, mutagenic (Ames test) or genotoxic (micronucleus test) potential. The absence of in vivo toxicity of the fractions was demonstrated by the absence of change in body weight and organs, serum concentrations of creatinine, uric acid, triglycerides, cholesterol, ALT and ALP, or in the CYP1A1 and GST activities in the liver. Phytochemical analysis (GC-MS and TLC) showed a large variety of terpenes in the fractions III and IV, being the furan-diterpenes vouacapan derivatives as major compounds. The isolated diterpene Ppg-01, present in fractions III (5.12%) and IV (18.47%), reduced the in vitro NO production and the paw edema induced by carrageenan (P <0.001). Together, the data suggest that Ppg-01 is contributing to the anti-inflammatory and immunomodulatory actions of the fractions III and IV and that these properties are associated with the observed anti-arthritic effects.