Body mass and composition in rapamycin-fed mice (filled circles) versus controls (hollow circles).

<p>P-values shown on individual panels only if there is a significant treatment effect independent of age. Sample sizes varied, depending on age, control females, n = 32–2; rapamycin females, n = 30–4; control males, n = 33–2; rapamycin males, n = 36–3. <b>A, B: Total body mass.</b> Highly significant differences (p << 0.001) exist in treatment x age effects in body for both sexes. Although they weighed less than controls by 16 months of age, rapamycin-fed females retained body mass longer, whereas rapamycin-fed males were similar to controls at 16 months but lost body mass earlier and remained lighter as they aged. <b>C, D: Percent body fat.</b> Highly significant differences (p << 0.001) in treatment x age effects exist for both sexes. As in with body mass, aging rapamycin-fed females retained body fat longer and lost body fat more slowly than age-matched controls. In contrast, rapamycin-fed males initially had a higher percentage of body fat, but lost fat mass earlier than controls. <b>E, F: Fat-free mass,</b> sometimes referred to as lean mass. Although obscured by the scaling, rapamycin-fed females had lower fat-free mass than controls at all ages measured. Fat-free mass declined more slowly with age in rapamycin-fed females than males.</p

Body weight, skeletal muscle mass, and testis weight of mitochondrial mutator mice.

<p>Body weights (A), vastus lateralis (VL) and gastrocnemius (GN) muscle mass (B) of 13–16 months old +/+, +/D257A, and D257A/D257A males (left) and females (right) under control diet or calorie restricted conditions. (C) Testes weights of 10–18 months old +/+, +/D257A, and D257A/D257A males under control diet or calorie restricted conditions. ‡P < 0.05 control diet vs CR diet within genotype. n = 8–16. *P < 0.05 +/+ vs D257A/D257A within diet. #P < 0.05 +/+ vs +/D257A within diet. §P < 0.05 +/D257A vs D257A/D257A within diet. +/+ = <i>wild-type</i>, +/D257A = <i>Polg</i>^+/D257A, and D257A/D257A = <i>Polg</i>^D257A/D257A.</p

Echocardiography of 13–16 months old wild-type and <i>Polg</i>^D257A/D257A mice on control and calorie-restricted diets.

<p>Echocardiography of 13–16 months old wild-type and <i>Polg</i>^D257A/D257A mice on control and calorie-restricted diets.</p

Age-related changes in inner ear histology was not altered by rapamycin treatment.

<p><b>A, B: The number of cochlear neurons</b> in male and female mice were not statistically different between control and rapamycin-fed animals. <b>C,D: The number of outer hair cells</b> in male and female mice were not statistically different between control and rapamycin-fed animals. <b>E,F: The number of inner hair cells</b> in male and female mice were not statistically different between control and rapamycin-fed animals.</p

Kaplan-Meier survival curves of D257A/D257A males and females.

<p>Survival curves of control diet (solid black line) and calorie restricted (red dashed line) D257A/D257A males (left) and females (right). n = 23–29. D257A/D257A = <i>Polg</i>^D257A/D257A.</p

Survival parameters of PolgD257A/D257A mice on control and calorie-restricted diets.

<p>Survival parameters of PolgD257A/D257A mice on control and calorie-restricted diets.</p

Body mass and composition in rapamycin-fed mice (filled circles) versus controls (hollow circles).

<p>P-values shown on individual panels only if there is a significant treatment effect independent of age. Sample sizes varied, depending on age, control females, n = 32–2; rapamycin females, n = 30–4; control males, n = 33–2; rapamycin males, n = 36–3. <b>A, B: Total body mass.</b> Highly significant differences (p << 0.001) exist in treatment x age effects in body for both sexes. Although they weighed less than controls by 16 months of age, rapamycin-fed females retained body mass longer, whereas rapamycin-fed males were similar to controls at 16 months but lost body mass earlier and remained lighter as they aged. <b>C, D: Percent body fat.</b> Highly significant differences (p << 0.001) in treatment x age effects exist for both sexes. As in with body mass, aging rapamycin-fed females retained body fat longer and lost body fat more slowly than age-matched controls. In contrast, rapamycin-fed males initially had a higher percentage of body fat, but lost fat mass earlier than controls. <b>E, F: Fat-free mass,</b> sometimes referred to as lean mass. Although obscured by the scaling, rapamycin-fed females had lower fat-free mass than controls at all ages measured. Fat-free mass declined more slowly with age in rapamycin-fed females than males.</p

Strength, coordination and movement in rapamycin-fed mice (filled circles) compared to controls (hollow circles).

<p>P-values shown on individual panels only if there is a significant treatment effect independent of age. <b>A, B: Grip strength</b> declined significantly with age in all animals, regardless of treatment (p << 0.001); however rapamycin treatment affected males and females differently (treatment x sex, p = 0.003). Rapamycin-fed females had greater grip strength than controls at all ages measured; whereas grip strength in control and rapamycin treated males did not differ. Sample sizes varied, depending on age, control females, n = 17–7; rapamycin females, n = 27–4; control males, n = 22–4; rapamycin males, n = 31–3. <b>C, D: Stride length</b> increased in males and females until 27 months of age and then declined with increasing age (p << 0.001). Rapamycin treatment had no effect on stride length in either sex. Sample sizes varied, depending on age, control females, n = 15–6; rapamycin females, n = 21–5; control males, n = 19–4; rapamycin males, n = 26–9. <b>E, F: Rotarod performance,</b> measured as maximum latency to fall, was significantly affected by body mass (p << 0.001) and so body mass was included as a covariate in the analysis. With the effects of body size removed, females showed no effects of rapamycin treatment and males showed a marginally significant negative effect of rapamycin treatment on rotarod performance. The y-axis shows the residuals of rotarod performance (latency to fall) regressed against body mass. Sample sizes varied, depending on age, control females, n = 11–6; rapamycin females, n = 19–6; control males, n = 13–7; rapamycin males, n = 21–8.</p

Differences in two rapamycin studies.

<p>↑ Increased ↓ Decreased, ↑↓ Maintained with age,—No difference from controls</p><p>*p < 0.05</p><p>** p < 0.01</p><p>*** p < 0.001</p><p>^§ Treatment X Age effect</p><p>Healthspan measures: young-fed measured between ages 16 and 29 months; old-fed measured at 25, 31 and 32 months.</p><p>Young-fed mice (rapamycin feeding begun at 4 mo, this study) compared with old-fed (rapamycin feeding begun at 19 mo) [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0126644#pone.0126644.ref011" target="_blank">11</a>].</p><p>Differences in two rapamycin studies.</p

Metabolic activity in rapamycin-fed mice (filled circles) compared to controls (hollow circles).

<p>P-values shown on individual panels only if there is a significant treatment effect independent of age. Sample sizes varied, depending on age, control females, n = 8–4; rapamycin females, n = 11–3; control males, n = 9–5; rapamycin males, n = 16–2. <b>A, B: Mass-specific metabolic rate during the light (= inactive) phase.</b> Males and females showed no effects of rapamycin treatment on mass-specific metabolic rate during the inactive phase of their daily 24-hour cycle, although both sexes showed highly significant (p << 0.001) sex x age treatment effects. <b>C, D: Mass-specific metabolic rate during the dark (= active) phase.</b> Aging rapamycin-fed females, but not males, maintained significantly higher metabolic rates between measures taken at 24 and 28 months of age compared to controls during the dark (= active) phase of the 24-hour light cycle. Both males and females showed highly significant (p <<0.001) decline dark-phase metabolic rate with age irrespective of treatment. <b>E, F: Resting mass-specific metabolic rate.</b> Resting metabolic rate declined with age in females, but aging rapamycin-fed females had higher resting metabolic rates compared to age-matched controls. Resting metabolic rate declined significantly in aging rapamycin-fed males but not in age-matched controls (treatment x age, p << 0.001).</p