Evaluation of the geroprotective effects of withaferin A in Drosophila melanogaster

Abstract

Withanolides are a class of compounds usually found in plant extracts which are an attractive geroprotective drug design starting point. We evaluated the geroprotective properties of Withaferin A (WA) in vivo using the Drosophila model. Flies were supplemented by nutrient medium with WA (at a concentration of 1, 10, or 100 μM dissolved in ethanol) for the experiment group and 30 μM of ethanol for the control group. WA treatment at 10 and 100 μM concentrations prolong the median life span of D. melanogaster's male by 7.7, 9.6% (respectively) and the maximum life span (the age of death 90% of individuals) by 11.1% both. Also WA treatment at 1, 10 and 100 μM improved the intestinal barrier permeability in older flies and affected an expression of genes involved in antioxidant defense (PrxV), recognition of DNA damage (Gadd45), heat shock proteins (Hsp68, Hsp83), and repair of double-strand breaks (Ku80). WA was also shown to have a multidirectional effect on the resistance of flies to the prooxidant paraquat (oxidative stress) and 33° C hyperthermia (heat shock). WA treatment increased the resistance to oxidative stress in males at 4 and 7 week old and decreased it at 6 weeks old. It increased the male's resistance to hyperthermia at 2, 4 and 7 weeks old and decreased it at 3, 5 and 8 weeks old. WA treatment decreased the resistance to hyperthermia in females at 1, 2 and 3 weeks old and not affected on their resistance to oxidative stress.

Keywords: Drosophila melanogaster; lifespan; stress response; stress-resistance; withaferin A.

Figure 1

Basic skeleton of withanolides. Withaferin A R₁ = OH, R₂ = H, R₃ = H; Withaferin D R₁ = H, R₂ = OH, R₃ = H; 27-Deoxywithaferin A R₁ = H, R₂ = H, R₃ = H; 27-hydroxywithanolide D R₁ = OH, R₂ = OH, R₃ = H; Dihydrodeoxywithaferin A R₁ = H, R₂ = H, R₃ = H; Dihydrowithaferin A R₁ = H, R₂ = OH, 17-hydroxywithaferin A, R₁ = R₃ = OH, R₂ = H.

Figure 2

Effect of Withaferin A on the life span in males (A, B) and females (C, D) of Drosophila melanogaster wild type Canton-S. Results of two independent repeats are combined. dM and d90% are the percentage of change in median life span and age of death of 90% of individuals (respectively); * p <0.01, ** p <0.001, *** p <0.0001.

Figure 3

Results of the intestinal barrier permeability test (smurf test) in males (A–C) and females (D–F) of Drosophila melanogaster wild type Canton-S at the age of 4, 6 and 8 weeks. * p <0.01, ** p <0.001.

Figure 4

Changing of stress response genes expression in males (A) and females (B) of Drosophila melanogaster wild type Canton-S after WA treatment. Results were normalized to control. * p <0.01, ** p <0.001, *** p <0.0001.