Chronic high fat feeding attenuates load-induced hypertrophy in mice

Abstract

The incidence of obesity and obesity-related conditions, such as metabolic syndrome and insulin resistance, is on the increase. The effect of obesity on skeletal muscle function, especially the regulation of muscle mass, is poorly understood. In this study we investigated the effect of diet-induced obesity on the ability of skeletal muscle to respond to an imposed growth stimulus, such as increased load. Male C57BL/6 mice were randomized into two diet groups: a low fat, high carbohydrate diet (LFD) and a high fat, low carbohydrate diet (HFD) fed ad libitum for 14 weeks. Mice from each diet group were divided into two treatment groups: sedentary control or bilateral functional overload (FO) of the plantaris muscle. Mice were evaluated at 3, 7, 14 or 30 days following FO. By 14 days of FO, there was a 10% reduction (P < 0.05) in absolute growth of the plantaris in response to overload in HFD mice vs. LFD mice. By 30 days the attenuation in growth increased to 16% in HFD mice compared to LFD mice. Following FO, there was a reduction in the formation of polysomes in the HFD mice relative to the LFD mice, suggesting a decrease in protein translation. Further, activation of Akt and S6K1, in response to increased mechanical loading, was significantly attenuated in the HFD mice relative to the LFD mice. In conclusion, chronic high fat feeding impairs the ability of skeletal muscle to hypertrophy in response to increased mechanical load. This failure coincided with a failure to activate key members of the Akt/mTOR signalling pathway and increase protein translation.

Figures

Figure 1. Growth curves of male C57BL/6 mice fed either a low fat (LFD) or a high fat diet (HFD) for a period of up to 18 weeks

The average (mean ±s.e.m.) body weight of mice fed either a LFD (•) or HFD (ô) was measured weekly over the 14 weeks of feeding (n= 60/diet). *Statistical difference between LFD and HFD mice at a specific age (P < 0.05).

Figure 2. Effect of diet on load-induced growth of the plantaris muscle following 3, 7, 14 and 30 days of functional overload (FO)

Histograms representing the average (mean ±s.e.m.) wet weight, in milligrams, of the plantaris of mice fed either a low fat diet (LFD) or a high fat diet (HFD) and subjected to normal cage activity (control (CON), filled bars) or FO (open bars) for a period of 3, 7, 14 or 30 days. *Statistical difference between CON and FO groups at a specific time (P < 0.05). #Statistical difference between LFD and HFD FO groups at a specific time (P < 0.05). Sample size: n= 10 mice/group at 3 and 7 days; n= 14, n= 20 mice/group at 14 and 30 days.

Figure 3. Effect of the diet duration on the growth response of the plantaris to 14 days of functional overload (FO)

Histograms representing the average (mean ±s.e.m.) wet weight, in milligrams, of the plantaris of mice fed either a low fat diet (LFD) or a high fat diet (HFD) for a total of 16 or 32 weeks, and then subjected to normal cage activity (control (CON), filled bars) or FO (open bars) for a period of 14 days. *Statistical difference between CON and FO groups for a specific diet group (P < 0.05). #Statistical difference between LFD FO and HFD FO groups for a specific diet duration (P < 0.05). §Statistical difference between 16 and 32 weeks HFD FO groups (P < 0.05).

Figure 5. Quantitative analysis of polysome aggregation in the plantaris

Histograms representing the average (mean ±s.e.m.) area under the curve for the entire polysome trace (A), and for the region after the 80S peak (B) of mice (n= 3–5/group) fed either a low fat diet (LFD) or a high fat diet (HFD), and then subjected to normal cage activity (control (CON), filled bars) or FO (open bars) for a period of 3, 7, 14 or 30 days. *Statistical difference between CON and FO groups for a specific diet group (P < 0.05). #Statistical difference between LFD FO and HFD FO groups for a specific diet duration (P < 0.05).

Figure 4. Qualitative analysis of polysome aggregation in the plantaris

Polysome profiles were produced using sucrose density gradient ultracentrifugation. A representative polysome profile is presented for sedentary control mice (CON) and mice subjected to 3, 7, 14 and 30 days of functional overload (FO) in both the low fat diet (LFD) and high fat diet (HFD) groups.

Figure 6. Effect of diet on Akt/PKB activation following functional overload (FO)

The phosphorylation status of Akt on Ser473 was measured in the plantaris muscle of mice fed either a low fat diet (LFD) or high fat diet (HFD), and then subjected to functional overload (FO) for 3, 7, 14 or 30 days or sedentary control (CON). Data expressed as mean ±s.e.m., n= 3–5/group. A, Akt phosphorylation on residue Ser473 expressed as level of phosphorylation per total protein for FO (open bars) and CON (filled bars) mice in each diet group and at each time point. B, relative activation of Akt on Ser473 in response to FO, expressed as fold change in Akt phosphorylation relative to control, for LFD (filled bars) and HFD (open bars) groups. *Statistical difference between CON and FO at a specific time point (P < 0.05). #Statistical difference between LFD and HFD groups (P < 0.05). Western blots of phosphorylated and native proteins from the plantaris of a single mouse in each group and time point (C, control).

Figure 7. Effect of diet on S6K1 activation following functional overload (FO)

The phosphorylation status of S6K1 on Thr389 was measured in the plantaris muscle of mice fed either a low fat diet (LFD) or high fat diet (HFD), and then subjected to functional overload (FO) for 3, 7, 14 or 30 days or sedentary control (CON). Data expressed as mean ±s.e.m., n= 3–5/group. A, S6K1 phosphorylation on residue Thr389 expressed as level of phosphorylation per total protein for FO (open bars) and CON (filled bars) mice in each diet group and at each time point. B, relative activation of S6K1 on Thr389 in response to FO, expressed as fold change in S6K1 phosphorylation relative to control, for LFD (filled bars) and HFD (open bars) groups. *Statistical difference between CON and FO at a specific time point (P < 0.05). #Statistical difference in FO between LFD and HFD groups (P < 0.05). Western blots of phosphorylated and native proteins from the plantaris of a single mouse in each group and time point (C, control).

Figure 8. Effect of diet on GSK-3β inactivation following functional overload (FO)

The phosphorylation status of GSK-3β on Ser9 was measured in the plantaris muscle of mice fed either a low fat diet (LFD) or high fat diet (HFD), and then subjected to functional overload (FO) for 3, 7, 14 or 30 days or sedentary control. Data expressed as mean ±s.e.m., n= 3–5/group. GSK-3β phosphorylation on residue Ser9 expressed as level of phosphorylation per total protein for FO (open bars) and control (filled bars) mice in each diet group and at each time point. Western blots of phosphorylated and native proteins from the plantaris of a single mouse in each group and time point (C, control).