Neile K. Edens

Effects of estradiol on tissue distribution of newly-synthesized fatty acids in rats and hamsters

Estradiol treatment decreases body weight and adiposity in ovariectomized (OVX) rats and hamsters partly by increasing energy expenditure. Other manipulations which increase energy expenditure (e.g., cold exposure or overfeeding) enhance thermogenesis in brown adipose tissue (BAT) and stimulate BAT fatty acid synthesis/uptake. We examined the effect of estradiol treatment on the in vivo distribution of newly-synthesized fatty acids in OVX rats and hamsters. In both species estradiol treatment increased BAT fatty acid synthesis/uptake (incorporation of tritium from (3H)2O into lipid), consistent with the possibility that enhanced thermogenesis in BAT may contribute to estradiol-induced energy expenditure. Estradiol treatment increased BAT lipoprotein lipase (LPL) activity in hamsters, but not in rats. Thus, hamsters may utilize fatty acids synthesized in other tissues as a fuel for BAT thermogenesis, whereas rats may rely more on in situ lipogenesis. Estradiol-induced decreases in carcass adiposity (white adipose tissue mass) may be accomplished by different means in rats and hamsters. Estradiol treatment reduced white adipose tissue LPL activity and fatty acid synthesis/uptake in rats, but not in hamsters. While there are some species differences in the effects of estradiol on lipid metabolism, it appears that in both rats and hamsters estradiol acts to direct metabolic fuels (especially lipids) away from white adipose tissue storage depots and into tissues where they are oxidized (e.g., BAT). Finally, cold acclimation and estradiol had similar effects in OVX hamsters including increases in BAT fatty acid synthesis/uptake, BAT LPL activity, and energy expenditure. These findings, too, are consistent with a role for BAT in estradiol-induced thermogenesis.

Molecular and metabolomic effects of voluntary running wheel activity on skeletal muscle in late middle‐aged rats

We examined the molecular and metabolomic effects of voluntary running wheel activity in late middle‐aged male Sprague Dawley rats (16–17 months). Rats were assigned either continuous voluntary running wheel access for 8 weeks (RW+) or cage‐matched without running wheel access (RW−). The 9 RW+ rats averaged 83 m/day (range: 8–163 m), yet exhibited both 84% reduced individual body weight gain (4.3 g vs. 26.3 g, P = 0.02) and 6.5% reduced individual average daily food intake (20.6 g vs. 22.0 g, P = 0.09) over the 8 weeks. Hindlimb muscles were harvested following an overnight fast. Muscle weights and myofiber cross‐sectional area showed no difference between groups. Western blots of gastrocnemius muscle lysates with a panel of antibodies suggest that running wheel activity improved oxidative metabolism (53% increase in PGC1α, P = 0.03), increased autophagy (36% increase in LC3B‐II/‐I ratio, P = 0.03), and modulated growth signaling (26% increase in myostatin, P = 0.04). RW+ muscle also showed 43% increased glycogen phosphorylase expression (P = 0.04) and 45% increased glycogen content (P = 0.04). Metabolomic profiling of plantaris and soleus muscles indicated that even low‐volume voluntary running wheel activity is associated with decreases in many long‐chain fatty acids (e.g., palmitoleate, myristoleate, and eicosatrienoate) relative to RW− rats. Relative increases in acylcarnitines and acyl glycerophospholipids were also observed in RW+ plantaris. These data establish that even modest amounts of physical activity during late middle‐age promote extensive metabolic remodeling of skeletal muscle.

Cellular and Physiological Effects of Dietary Supplementation with β-Hydroxy-β-Methylbutyrate (HMB) and β-Alanine in Late Middle-Aged Mice.

There is growing evidence that severe decline of skeletal muscle mass and function with age may be mitigated by exercise and dietary supplementation with protein and amino acid ingredient technologies. The purposes of this study were to examine the effects of the leucine catabolite, beta-hydroxy-beta-methylbutyrate (HMB), in C2C12 myoblasts and myotubes, and to investigate the effects of dietary supplementation with HMB, the amino acid β-alanine and the combination thereof, on muscle contractility in a preclinical model of pre-sarcopenia. In C2C12 myotubes, HMB enhanced sarcoplasmic reticulum (SR) calcium release beyond vehicle control in the presence of all SR agonists tested (KCl, P<0.01; caffeine, P = 0.03; ionomycin, P = 0.03). HMB also improved C2C12 myoblast viability (25 μM HMB, P = 0.03) and increased proliferation (25 μM HMB, P = 0.04; 125 μM HMB, P<0.01). Furthermore, an ex vivo muscle contractility study was performed on EDL and soleus muscle from 19 month old, male C57BL/6nTac mice. For 8 weeks, mice were fed control AIN-93M diet, diet with HMB, diet with β-alanine, or diet with HMB and β-alanine. In β-alanine fed mice, EDL muscle showed a 7% increase in maximum absolute force compared to the control diet (202 ± 3vs. 188± 5 mN, P = 0.02). At submaximal frequency of stimulation (20 Hz), EDL from mice fed HMB plus β-alanine showed an 11% increase in absolute force (88.6 ± 2.2 vs. 79.8 ± 2.4 mN, P = 0.025) and a 13% increase in specific force (12.2 ± 0.4 vs. 10.8 ± 0.4 N/cm2, P = 0.021). Also in EDL muscle, β-alanine increased the rate of force development at all frequencies tested (P<0.025), while HMB reduced the time to reach peak contractile force (TTP), with a significant effect at 80 Hz (P = 0.0156). In soleus muscle, all experimental diets were associated with a decrease in TTP, compared to control diet. Our findings highlight beneficial effects of HMB and β-alanine supplementation on skeletal muscle function in aging mice.

Dietary HMB and β-alanine co-supplementation does not improve in situ muscle function in sedentary, aged male rats

This study evaluated the effects of dietary β-hydroxy-β-methylbutyrate (HMB) combined with β-alanine (β-Ala) in sedentary, aged male rats. It has been suggested that dietary HMB or β-Ala supplementation may mitigate age-related declines in muscle strength and fatigue resistance. A total of 20 aged Sprague-Dawley rats were studied. At age 20 months, 10 rats were administered a control, purified diet and 10 rats were administered a purified diet supplemented with both HMB and β-Ala (HMB+β-Ala) for 8 weeks (approximately equivalent to 3 and 2.4 g per day human dose). We measured medial gastrocnemius (MG) size, force, fatigability, and myosin composition. We also evaluated an array of protein markers related to muscle mitochondria, protein synthesis and breakdown, and autophagy. HMB+β-Ala had no significant effects on body weight, MG mass, force or fatigability, myosin composition, or muscle quality. Compared with control rats, those fed HMB+β-Ala exhibited a reduced (41%, P = 0.039) expression of muscle RING-finger protein 1 (MURF1), a common marker of protein degradation. Muscle from rats fed HMB+β-Ala also exhibited a 45% reduction (P = 0.023) in p70s6K phosphorylation following fatiguing stimulation. These data suggest that HMB+β-Ala at the dose studied may reduce muscle protein breakdown by reducing MURF1 expression, but has minimal effects on muscle function in this model of uncomplicated aging. They do not, however, rule out potential benefits of HMB+β-Ala co-supplementation at other doses or durations of supplementation in combination with exercise or in situations where extreme muscle protein breakdown and loss of mass occur (e.g., bedrest, cachexia, failure-to-thrive).

Effects of running wheel activity and dietary HMB and B—alanine co-supplementation on muscle quality in aged male rats

ObjectiveLoss of skeletal muscle function is linked to increased risk for loss of health and independence in older adults. Dietary interventions that can enhance aging muscle function, alone or in combination with exercise, may offer an effective way to reduce these risks. The goal of this study was to evaluate the muscular effects of beta-hydroxy-beta-methylbutyrate (HMB) and beta-alanine (β-Ala) co-supplementation in aged Sprague-Dawley rats with voluntary access to running wheels (RW).MethodsAged (20 months) rats were housed with ad libitum access to RW while on a purified diet for 4 weeks, then balanced for RW activity and assigned to either a control or an experimental diet (control + HMB and β-Ala) for the next 4 weeks (n = 10/group). At the end of the study, we assessed muscle size, in situ force and fatigability in the medial gastrocnemius muscles, as well as an array of protein markers related to various age- and activity-responsive signaling pathways.ResultsDietary HMB+β-Ala did not improve muscle force or fatigue resistance, but a trend for increased muscle cross-sectional area (CSA) was observed (P = 0.077). As a result, rats on the experimental diet exhibited reduced muscle quality (force/CSA; P = 0.032). Dietary HMB+β-Ala reduced both the abundance of PGC1-α (P = 0.050) and the ratio of the lipidated to non-lipidated forms of microtubule-associated protein 1 light chain 3 beta (P = 0.004), markers of mitochondrial biogenesis and autophagy, respectively. Some alterations in myostatin signaling also occurred in the dietary HMB+β-Ala group. There was an unexpected difference (P = 0.046) in RW activity, which increased throughout the study in the animals on the control diet, but not in animals on the experimental diet.ConclusionsThese data suggest that the short-term addition of dietary HMB+β-Ala to modest physical activity provided little enhancement of muscle function in this model of uncomplicated aging.