Comunicação ao VIII International Symposium on Experimental Tecnics -Yeast as eukaryotic model to detect metal response - glutathione as marker of intracellular redox status and V2O5 as stress agent

Abstract

Glutathione a thiol-containing tripeptide is implicated in the cellular defence against xenobiotics and endogenous deleterious compounds, such as free radicals and hydroperoxides. Usually glutathione status is a highly sensitive indicator of cell functionality and viability. In Human its depletion is linked to aging, cancer, neurodegenerative and cardiovascular diseases. Vanadium, a transition metal, is used as therapeutic agent in diabetes and cancer. Although Vanadium oxide (V2O5) exhibits the most biological activity among vanadium compounds its molecular effects are poorly studied. In this work we use yeast cells as model to evaluate stress effects of V2O5 on cell viability and glutathione homeostasis. Here we compare the effects of five medium V2O5 (0.5, 1.0, 1.5 and 2.0 mM) on growth rate, GSH, GSSG and GSH/GSSG ratio of wild Saccharomyces cerevisiae UE-ME3. The cells were grown to OD 0.8, at 28ºC, in YEPD medium with 2% (w/v) glucose. Cells (106 cells ml-1) at mid-exponential phase were exposed to V2O5, during 200 min. CFU were determined, and cells were harvested for ultra-sonic disrupt. The post-peroxissomal supernatants, obtained by differential centrifugation, where used for glutathione and glutathione disulfide determination by OPA fluorescence method, using NEM as scavenger of endogenous GSH in the GSSG procedure. The results show a bimodal response of S. cerevisiae to V2O5, with slightly effect on yeast cells growth rate for 0.5 and 1.0 mM, and a significantly decrease (58% and 14% of control) for 1.5 and 2.0 mM, respectively. Conversely, we observed a significant increase of GSH/GSSG ratio for 0.5 mM V2O5, and a decrease of GSH/GSSG ratio for V2O5 higher than 1.0 mM in culture medium. We interpret the increase of GSH/GSSG as an adaptative response to the stress making yeast cells more resistant to oxidative effects. Conversely the decrease of reducing power observed for V2O5 higher than 1.0 mM in culture medium may be linked to apoptosis.