Figure 4

Rapamycin treatment extends lifespan, improves barrier function and reduces TOR signaling in germ-free flies. (A) Survival curves of w^Dahomey female flies rendered germ free as embryos, and either maintained germ free (AX, dashed lines) or refed with a bacterial homogenate (HF, solid lines) and treated with (red) or without (black) rapamycin from day 4 onwards (post-eclosion). ***p < 0.0001, log rank test; n ≥ 200 flies. Representative result of 3 separate lifespan trials shown. (B) Intestinal integrity of day 10, 35 and 42 w^Dahomey flies with (HF, solid bars) or without homogenate feeding (AX, dashed bars) and with (red) or without rapamycin (black) treatment from day 4 onwards (post-eclosion). **p < 0.01 and ***p < 0.001, one-way ANOVA/Bonferroni’s multiple comparisons test; n = 300 flies at day 10 per treatment. (C) Western blot detection and densitometry of p-S6K (T398) and total S6K levels from day 10 homogenate fed (HF) w^Dahomey flies treated with or without rapamycin from day 4 onwards. n = 5 replicates, 10 flies per replicate; **p < 0.01; Mann-Whitney U-test. (D) Western blot detection and densitometry of p-S6K (T398) and total S6K levels from day 10 axenic (AX) w^Dahomey flies treated with or without rapamycin from day 4 onwards. n = 5 replicates, 10 flies per replicate; *p < 0.05; Mann-Whitney U-test. Rapamycin was provided in the media at a concentration of 10 μg/ml. All error bars represent SEM.