Estudo comparativo dos efeitos moduladores da (-)-cubebina sobre a mutagenicidade / recombinogenicidade induzida por diferentes agentes químicos

Abstract

(-)-Cubebin (CUB) is dibenzylbutyrolactolic lignan isolated from dry seeds of Piper cubeba L. (Piperaceae). CUB possesses anti-inflammatory, analgesic and antimicrobial activities. In the present study were examined the mutagenicity and recombinogenicity of different concentrations (0.25; 0.5; 1.0; 2.0 ou 4.0 mM) of CUB alone or in combination with DXR (DXR 0.2 mM) using standard (ST) and high bioactivation (HB) crosses of the wing Somatic Mutation And Recombination Test (SMART) in Drosophila melanogaster. The results from both crosses were rather similar. CUB alone did not induce mutation or recombination. Doxorubicin (DXR) used as a positive control and also to antimutagenic evaluation of CUB, is a chemotherapeutic agent that inhibits topoisomerase inducing single- and double-strand breaks, intercalate into the DNA and generate oxygen free radicals. Recombination is the major effect observed in wing somatic cells of D. melanogaster when treated with DXR. When associated (CUB + DXR), at lower concentrations, CUB significantly reduced the frequencies of mutant spots induced by DXR. However, increasing the concentrations, CUB potentiated the effects of DXR, raising the frequencies of mutant spots and at higher concentrations, due to toxicity, the frequencies of mutant spots reduced. The combined treatments reduced DXR mutagenicity but did not affect DXR recombinogenicity. To the assessment of the interference of CUB on the chromosomal damage (clastogenic and aneugenic effects) induced by DXR (2 μM), were used lung fibroblasts of Chinese hamster (cells V79). For this purpose, the frequency of micronuclei (MN) in 6000 binucleated cells/treatment was used as a parameter of analysis. The chosen concentrations (2, 4, 8 or 16 μM) were obtained after evaluation of the nuclear division index (3000 cells/treatment), used to evaluated cytotoxicity potential. No significant difference in the induction of MN was observed between the groups treated with the different concentrations of CUB and the negative control (P>0.05). There was no significant difference on MN induction observed between groups treated with different concentrations of CUB when compared to negative control (P> 0,05). These findings indicate the absence of mutagenic effects of CUB at the concentrations tested. The results also showed that CUB significantly reduced the frequency of MN induced by DXR, with a mean reduction of 63.88%. In order to evaluate the interference of CUB on the enzymatic detoxification complex cytochrome P450 (CYP), ethyl carbamate (urethane URE) was associated with CUB. URE have been proved to be a promutagen and procarcinogenic substance due to its metabolization through CYP increases significantly the frenquencies of mutant spots in D. melanogaster. The ST and HB versions of the SMART were used to estimate the antimutagenic / antirecombinogenic effects of CUB (0.25; 0.5; 1.0; 2.0 or 4.0 mM) associated to URE (10 mM). The data obtained showed that in the MH individuals from the ST cross, significant inhibitory effect was found only at concentrations of 1.0 and 2.0 mM, while in HB cross all concentrations of CUB statistically inhibit URE-induced DNA damage. At lower concentrations, the recombination level decreases, but at highest concentration the recombination level increases. In conclusion, our data suggest that depending on the concentration, CUB may act as a free radical scavenger at low concentrations; a pro-oxidant at higher concentrations, when it interacts with the enzymatic system that catalyzes the metabolic detoxification of DXR or URE; and/or an inducer of recombinational DNA repair. These studies should collaborate with the better understanding of mechanisms of action of CUB and, consequently, to the production of effective and safe drugs with low toxicity.