Skeletal Muscle Gene Expression Profile in Response to Caloric Restriction and Aging: A Role for SirT1.

SirT1 plays a crucial role in the regulation of some of the caloric restriction (CR) responsive biological pathways. Aging suppresses SirT1 gene expression in skeletal muscle, suggesting that aging may affect the role of CR in muscle. To determine the role of SirT1 in the regulation of CR regulated pathways in skeletal muscle, we performed high-throughput RNA sequencing using total RNA isolated from the skeletal muscles of young and aged wild-type (WT), SirT1 knockout (SirT1-KO), and SirT1 overexpression (SirT1-OE) mice fed to 20 wk ad libitum (AL) or 40% CR diet. Our data show that aging repressed the global gene expression profile, which was restored by CR via upregulating transcriptional and translational process-related pathways. CR inhibits pathways linked to the extracellular matrix and cytoskeletal proteins regardless of aging. Mitochondrial function and muscle contraction-related pathways are upregulated in aged SirT1 KO mice following CR. SirT1 OE did not affect whole-body energy expenditure or augment skeletal muscle insulin sensitivity associated pathways, regardless of aging or diet. Overall, our RNA-seq data showed that SirT1 and CR have different functions and activation of SirT1 by its activator or exercise may enhance SirT1 activity that, along with CR, likely have a better functional role in aging muscle.

The role of SIRT1 in skeletal muscle function and repair of older mice.

BACKGROUND: Sirtuin 1 (SIRT1) is a NAD+ sensitive deacetylase that has been linked to longevity and has been suggested to confer beneficial effects that counter aging-associated deterioration. Muscle repair is dependent upon satellite cell function, which is reported to be reduced with aging; however, it is not known if this is linked to an aging-suppression of SIRT1. This study tested the hypothesis that Sirtuin 1 (SIRT1) overexpression would increase the extent of muscle repair and muscle function in older mice. METHODS: We examined satellite cell dependent repair in tibialis anterior, gastrocnemius, and soleus muscles of 13 young wild-type mice (20-30 weeks) and 49 older (80+ weeks) mice that were controls (n = 13), overexpressed SIRT1 in skeletal muscle (n = 14), and had a skeletal muscle SIRT1 knockout (n = 12) or a satellite cell SIRT1 knockout (n = 10). Acute muscle injury was induced by injection of cardiotoxin (CTX), and phosphate-buffered saline was used as a vector control. Plantarflexor muscle force and fatigue were evaluated before or 21 days after CTX injection. Satellite cell proliferation and mitochondrial function were also evaluated in undamaged muscles. RESULTS: Maximal muscle force was significantly lower in control muscles of older satellite cell knockout SIRT1 mice compared to young adult wild-type (YWT) mice (P < 0.001). Mean contraction force at 40 Hz stimulation was significantly greater after recovery from CTX injury in older mice that overexpressed muscle SIRT1 than age-matched SIRT1 knockout mice (P < 0.05). SIRT1 muscle knockout models (P < 0.05) had greater levels of p53 (P < 0.05 MKO, P < 0.001 OE) in CTX-damaged tissues as compared to YWT CTX mice. SIRT1 overexpression with co-expression of p53 was associated with increased fatigue resistance and increased force potentiation during repeated contractions as compared to wild-type or SIRT1 knockout models (P < 0.001). Muscle structure and mitochondrial function were not different between the groups, but proliferation of satellite cells was significantly greater in older mice with SIRT1 muscle knockout (P < 0.05), but not older SIRT1 satellite cell knockout models, in vitro, although this effect was attenuated in vivo after 21 days of recovery. CONCLUSIONS: The data suggest skeletal muscle structure, function, and recovery after CTX-induced injury are not significantly influenced by gain or loss of SIRT1 abundance alone in skeletal muscle; however, muscle function is impaired by ablation of SIRT1 in satellite cells. SIRT1 appears to interact with p53 to improve muscle fatigue resistance after repair from muscle injury.

Effects of hindlimb suspension and reloading on gastrocnemius and soleus muscle mass and function in geriatric mice.

Reloading of atrophied muscles after hindlimb suspension (HLS) can induce muscle injury and prolong recovery after disuse in old rats, especially in fast contracting muscles. Less is known about the responses in mice and whether fast and slow muscles from geriatric mice will respond in a similar fashion to HLS unloading and recovery (HLS + R). Furthermore, while slow muscles undergo atrophy with disuse, they typically are more resistant to sarcopenia than fast contracting muscles. Geriatric (28 mo. of age) male C57BL/6 mice were randomly placed into 3 groups. These included HLS for 14 days n = 9, and HLS followed by 14 days of reloading recovery (HLS + R; n = 9), or normal ambulatory cage controls (n = 9). Control mice were not exposed to unloading. Electrically evoked maximal muscle function was assessed in vivo in anesthetized mice at baseline, after 14 days of HLS or HLS + R. As expected, HLS significantly reduced body weight, wet weight of gastrocnemius and soleus muscles and in vivo maximal force. There were no differences in vivo fatigability of the plantar flexor muscles and overall fiber size. There were only minor fiber type distribution and frequency distribution of fiber sizes that differ between HLS + R and control gastrocnemius and soleus muscles. Soleus muscle wet weight had recovered to control levels after reloading, but type I/IIA fibers in the soleus muscles were significantly smaller after HLS + R than control muscles. In contrast, gastrocnemius muscle wet weight did not recover to control levels after reloading. Plantar flexion muscle force (primarily influenced by the gastrocnemius muscles) did not recover in HLS + R conditions as compared to HLS conditions and both were lower than control force production signaling for apoptosis, autophagy and anabolic markers were not different between control and HLS + R gastrocnemius and soleus muscles in geriatric mice. These results suggest that molecular signaling does not explain attenuated ability to regain muscle wet weight, fiber size or muscle force production after HLS in geriatric mice. It is possible that fluid shifts, reduced blood flow, or shortened muscle fibers which failed to regain control lengths contributed to the attenuation of muscle wet weight after HLS and reloading and this affected force production. Further work is needed to determine if altered/loss of neural activity contributed to the inability of geriatric mice to regain gastrocnemius muscle weight and function after HLS and reloading.

Mitochondria Initiate and Regulate Sarcopenia.

We present the hypothesis that an accumulation of dysfunctional mitochondria initiates a signaling cascade leading to motor neuron and muscle fiber death and culminating in sarcopenia. Interactions between neural and muscle cells that contain dysfunctional mitochondria exacerbate sarcopenia. Preventing sarcopenia will require identifying mitochondrial sources of dysfunction that are reversible.

Dysregulation of SIRT-1 in aging mice increases skeletal muscle fatigue by a PARP-1-dependent mechanism.

Accumulation of reactive oxygen species (ROS) in skeletal muscles and the resulting decline in muscle performance are hallmarks of sarcopenia. However, the precise mechanism by which ROS results in a decline in muscle performance is unclear. We demonstrate that isometric-exercise concomitantly increases the activities of Silent information regulator 1 (SIRT-1) and Poly [ADP-ribose] polymerase (PARP-1), and that activated SIRT-1 physically binds with and inhibits PARP-1 activity by a deacetylation dependent mechanism in skeletal muscle from young mice. In contrast, skeletal muscle from aged mice displays higher PARP-1 activity and lower SIRT-1 activity due to decreased intracellular NAD+ content, and as a result reduced muscle performance in response to exercise. Interestingly, injection of PJ34, a PARP-1 inhibitor, in aged mice increased SIRT-1 activity by preserving intracellular NAD+ content, which resulted in higher skeletal muscle mitochondrial biogenesis and performance. We found that the higher activity of PARP-1 in H2O2-treated myotubes or in exercised-skeletal muscles from aged mice is due to an elevated level of PARP-1 acetylation by the histone acetyltransferase General control of amino acid synthesis protein 5-like 2 (GCN-5). These results suggest that activation of SIRT-1 and/or inhibition of PARP-1 may ameliorate skeletal muscle performance in pathophysiological conditions such as sarcopenia and disuse-induced atrophy in aging.

Regulation of satellite cell function in sarcopenia.

The mechanisms contributing to sarcopenia include reduced satellite cell (myogenic stem cell) function that is impacted by the environment (niche) of these cells. Satellite cell function is affected by oxidative stress, which is elevated in aged muscles, and this along with changes in largely unknown systemic factors, likely contribute to the manner in which satellite cells respond to stressors such as exercise, disuse, or rehabilitation in sarcopenic muscles. Nutritional intervention provides one therapeutic strategy to improve the satellite cell niche and systemic factors, with the goal of improving satellite cell function in aging muscles. Although many elderly persons consume various nutraceuticals with the hope of improving health, most of these compounds have not been thoroughly tested, and the impacts that they might have on sarcopenia and satellite cell function are not clear. This review discusses data pertaining to the satellite cell responses and function in aging skeletal muscle, and the impact that three compounds: resveratrol, green tea catechins, and ß-Hydroxy-ß-methylbutyrate have on regulating satellite cell function and therefore contributing to reducing sarcopenia or improving muscle mass after disuse in aging. The data suggest that these nutraceutical compounds improve satellite cell function during rehabilitative loading in animal models of aging after disuse (i.e., muscle regeneration). While these compounds have not been rigorously tested in humans, the data from animal models of aging provide a strong basis for conducting additional focused work to determine if these or other nutraceuticals can offset the muscle losses, or improve regeneration in sarcopenic muscles of older humans via improving satellite cell function.

The effects of age, sex, season and geographic region on circulating serum cortisol concentrations in threatened and endangered Steller sea lions (Eumetopias jubatus).

The role of stress in the decline of Steller sea lions has become of interest in the identification of factors leading to their drastic decline and subsequent failure to recover. Serum cortisol was validated as a potential indicator of stress in three sub-populations inhabiting the majority of the Steller sea lions range (n=941). Additionally, seasonal variability in serum cortisol measured in captive animals (n=9) of different age classes and sex was investigated. Significant interactions were found between region and year, as well as year and sex (p=0.001 and p=0.005, respectively). Samples from pups in Russia (151.5+/-3.2 ng ml(-1)) and southeast Alaska (152.7+/-2.7 ng ml(-1)) were consistently higher than concentrations measured in pups from southwest Alaska (133.0+/-3.2 ng ml(-1)). Serum cortisol concentrations measured in 2001 (134.9+/-2.0 ng ml(-1)) were significantly lower than 2002 (156.5+/-2.4 ng ml(-1), p<0.001) for all regions. Seasonal effects indicated significant interactions between age, year and sex (p=0.011). Serum cortisol was elevated in months associated with the breeding season and annual molt (86.5+/-5.0 and 92.4+/-5.1 ng ml(-1), respectively) compared to the other seasons. Our results suggest that cortisol concentrations in Steller sea lions can be a useful diagnostic tool to compare the physiology between groups of sea lions, but factors such as animal sex, age, season and year must be considered.

Organochlorine contaminants in endangered Steller sea lion pups (Eumetopias jubatus) from western Alaska and the Russian Far East.

Investigations into the cause of the Steller sea lion population decline have focused on numerous factors, including exposure to toxic contaminants such as organochlorines (OCs). OCs, such as polychlorinated biphenyls (PCBs), 1,1,1-trichloro-2,2-bis(p-chlorophenyl)ethane, or dichlorodiphenyltrichloroethane (DDT), have been associated with various biological effects in marine mammals. We measured these compounds in whole blood of free-ranging Steller sea lion pups in order to determine the extent and magnitude of contamination across their geographical range. Of 212 pups analyzed for OCs, 76 pups (36 females and 40 males) were from western Alaska and the other 136 (63 females and 73 males) were from the Russian Far East. Concentrations of summation SigmaPCBs in the whole blood of pups from western Alaska ranged from 0.21 to 13 ng/g wet weight with a mean of 2.1+/-0.27 ng/g wet weight. In the Russian animals, summation SigmaPCB concentrations in the whole blood of pups ranged from 0.33 to 36 ng/g wet weight with a mean of 4.3+/-0.44 ng/g wet weight. summation SigmaDDT concentrations in the whole blood of pups from western Alaska ranged from 0.18 to 11 ng/g wet weight with a mean of 1.6+/-0.23 ng/g wet weight. In Russia, summation SigmaDDT in the whole blood of pups ranged from undetectable to 26 ng/g wet weight with a mean of 3.3+/-0.36 ng/g wet weight. Average OC concentrations were significantly higher in the blood of Russian animals compared to western Alaska (for PCBs and DDTs, p<0.001) and in both areas females had higher concentrations than males. Male pups from western Alaska had significantly lower levels of summation SigmaPCBs and summation SigmaDDT when compared to male pups from Russia (for PCBs and DDTs p<0.001). Female pups from western Alaska were significantly lower in summation SigmaPCBs than Russian female pups (for PCBs p=0.009) as were female pups for summation SigmaDDT levels between areas (for DDTs p=0.026). OC contaminants data indicate that Steller sea lion pups have measurable concentrations of these synthetic chemicals. While any physiological effect and the specific role these chemicals may have in either the decline or the failure of the endangered Steller sea lion population to recover needs to be further investigated, this study indicates specific areas and animals that may be most at risk.

The effects of age, season and geographic region on thyroid hormones in Steller sea lions (Eumetopias jubatus).

The purpose of this study was to investigate thyroid hormone concentrations, thyroxine (T4) and triiodothyronine (T3), in order to determine basal levels in Steller sea lions of different ages and over seasons. Serum concentrations of total T4 were highest in Steller sea lions followed by total T3 concentrations. Concentrations of free T4 and free T3 were three to four orders of magnitude lower. Concentrations for all four thyroid hormone measurements tended to a lower level as animals matured beyond the neonatal stage. When thyroid hormones from captive sea lions were evaluated across seasons, all thyroid hormones were highest in the July to September period. When compared across the geographic range, animals in southeast Alaska tended to have lower thyroid hormone levels, while the Steller sea lions west of Prince William Sound and animals from the Russian Far East had significantly higher concentrations. Significant inter-annual differences in concentrations were also observed across the geographic range. With an understanding of the basic changes in thyroid hormone concentrations, changes in plane of nutrition or life history states (i.e. fasting, pregnancy or lactation) can now be evaluated for their effect on the overall health of this endangered species.