Effect of Furostanol Glycosides from Dioscorea deltoidea on Redox State of Alfalfa Cells In Vitro

The effect of exogenous furostanol glycosides (FG) on the activity of redox enzymes was investigated in suspension cell culture of alfalfa (Medicago sativa L.). It was shown that 60-min-long treatment of alfalfa cells with FG at a concentration of 10(-5) M triggered formation of ROS and activated enzymes of the antioxidant complex: superoxide dismutase and guaiacol-dependent peroxidase where the highest activity was associated with ion-bound fraction. Application of inhibitor of NADPH oxidase diphenyliodonium chloride showed that ROS are generated in the presence of FG by NADPH oxidase of plasma membrane. It was found that treatment of alfalfa in vitro cells with FG elevated activity of the key enzyme of pentose phosphate pathway: glucose-6-phosphate dehydrogenase (G-6-P DH) and glutathione reductase. A relationship was revealed between the operation of G-6-P DH, NADPH oxidase, and glutathione reductase. It is assumed that, owing to suppression of one of the main consumers of NADPH (NADPH oxidase), a rise in the activity of glutathione reductase may eliminate the inhibition of G-6-P DH. Under hyperosmotic stress, FG improved the viability of alfalfa cells in vitro to 70%, whereas it was only 24% in control material. At the same time, viability reached 90% in reference cells without treatment. Such an effect of FG became apparent as a result of elevation in activity of aldehyde dehydrogenase, reduction in lipid peroxidation (by 24%), and activation of antioxidant enzymes. Adaptation mechanisms operating on the level of redox systems are discussed.