Myostatin mRNA Levels Research Articles

In ovo betaine injection improves breast muscle growth in newly hatched goslings through FXR/IGF-2 pathway

Betaine has been shown to enhance growth performance and increase breast muscle yield in ducks and broilers through various mechanisms, including the modification of DNA methylation. However, the impact of in ovo betaine injection on muscle growth in newly hatched goslings remains unclear. In this study, fifty eggs were injected with saline or betaine at 7.5 mg/egg prior to incubation, and the subsequent effects on breast muscle growth in the newly hatched goslings were investigated. Betaine significantly increased (P < 0.05) the hatch weight, breast muscle weight, and breast muscle index, accompanied by an augmentation in muscle bundle cross-sectional area. Concurrently, betaine significantly upregulated (P < 0.05) the expression levels of myogenic regulatory factors, including myogenin (MyoG) and paired box 7 (Pax7) both mRNA and protein, while downregulating (P < 0.05) the mRNA and protein levels of myostatin (MSTN). Histological analysis revealed a higher abundance of proliferating cell nuclear antigen (PCNA) and Pax7 immune-positive cells in the breast muscle of the betaine group, consistent with elevated PCNA and Pax7 mRNA and protein levels. Additionally, significantly increased (P < 0.05) contents of insulin-like growth factor 1 (IGF-1) and insulin-like growth factor 2 (IGF-2) were observed in the breast muscle of the betaine group, so was mRNA expression of IGF-1, IGF-2, and insulin-like growth factor 1 receptor (IGF-1R). Betaine also significantly in8creased (P < 0.05) global DNA methylation of the breast muscle, accompanied by enhanced mRNA and protein levels of methionine cycle and DNA methylation-related enzymes, Interestingly, the promoter regions of IGF-1, IGF-2, and IGF-1R genes were significantly hypomethylated (P < 0.05). Moreover, in ovo betaine injection significantly upregulated (P < 0.05) the protein level of farnesoid X receptor (FXR) in breast muscle and FXR binding to the promoter of IGF-2 gene. These findings suggest that in ovo betaine injection promotes breast muscle growth during embryonic development in goslings through the FXR-mediated IGF-2 pathway, ultimately improving hatch weight and breast muscle weight.

Long-term fasting induced basal thermogenesis flexibility in female Japanese quails

Male Japanese quails (Coturnix japonica) have been found to exhibit a three-phase metabolic change when subjected to prolonged fasting, during which basal thermogenesis is significantly reduced. A study had shown that there is a significant difference in the body temperature between male and female Japanese quails. However, whether female Japanese quails also show the same characteristic three-phase metabolic change during prolonged fasting and the underlying thermogenesis mechanisms associated with such changes are still unclear. In this study, female Japanese quails were subjected to prolonged starvation, and the body mass, basal metabolic rate (BMR), body temperature, mass of tissues and organs, body fat content, the state-4 respiration (S4R) and cytochrome c oxidase (CCO) activity in the muscle and liver of these birds were measured to determine the status of metabolic changes triggered by the starvation. In addition, the levels of glucose, triglyceride (TG) and uric acid, and thyroid hormones (T3 and T4) in the serum and the mRNA levels of myostatin (MSTN) and avian uncoupling protein (av-UCP) in the muscle were also measured. The results revealed the existence of a three-phase stage similar to that found in male Japanese quails undergoing prolonged starvation. Fasting resulted in significantly lower body mass, BMR, body temperature, tissues masses and most organs masses, as well as S4R and CCO activity in the muscle and liver. The mRNA level of av-UCP decreased during fasting, while that of MSTN increased but only during Phase I and II and decreased significantly during Phase III. Fasting also significantly lowered the T3 level and the ratio of T3/T4 in the serum. These results indicated that female Japanese quails showed an adaptive response in basal thermogenesis at multiple hierarchical levels, from organismal to biochemical, enzyme and cellular level, gene and endocrine levels and this integrated adjustment could be a part of the adaptation used by female quails to survive long-term fasting.

Cysteine-cysteine Chemokine Receptor Type 5 Plays a Critical Role in Exercise Performance by Regulating Mitochondrial Content in Skeletal Muscle.

Cysteine-cysteine chemokine receptor type 5 (CCR5) is thought to play an important role in the trafficking of lymphoid cells but has recently also been associated with AMPK signaling pathways that are implicated in energy metabolism in skeletal muscle. We hypothesized that genetic deletions of CCR5 would alter mitochondria content and exercise performance in mice. CCR5-/- and wild-type mice with the same genetic background were subjected to endurance exercise and grip strength tests. The soleus muscle was stained with immunofluorescence for myosin heavy chain 7 (MYH7) and succinate dehydrogenase (SDH) analysis as well as the expression of genes associated with muscle atrophy and mitochondrial oxidative phosphorylation were measured using qPCR. Although there were no differences in the weight of the soleus muscle between the CCR5-/- group and the wild-type mice, the CCR5-/- mice showed the following muscular dysfunctions: (i) decreased MYH7 percentage and cross-section area, (ii) higher myostatin and atrogin-1 mRNA levels, (iii) dropped expression of mitochondrial DNA-encoded electron respiratory chain genes (cytochrome B, cytochrome c oxidase subunit III, and ATP synthase subunit 6) as well as mitochondrial generation genes (PPARγandPGC-1α), and (iv) lower SDH activity and exercise performance when compared with wild-type mice. In addition, genes associated with mitochondrial biogenesis (PGC-1α, PPARγ, and MFN2) and mitochondrial complex (ND4 and Cytb) were upregulated when the skeletal muscle cell line C2C12 was exposed to cysteine-cysteine chemokine ligand 4 (a ligand of CCR5) in vitro. These findings suggested that attenuation of endurance exercise performance is related to the loss of mitochondrial content and lower SDH activity of soleus muscle in CCR5 knockout mice. The present study provides evidence indicating that the chemokine receptor CCR5 might modulate the skeletal muscle metabolic energy system during exercise.

A new insight on copper: Promotion of collagen synthesis and myofiber growth and development in juvenile grass carp (Ctenopharyngodon idella)

Copper (Cu) is a trace element, essential for fish growth. In the current study, in addition to growth performance, we first explored the effects of Cu on collagen synthesis and myofiber growth and development in juvenile grass carp (Ctenopharyngodon idella). A total of 1080 fish (11.16 ± 0.01 g) were randomly divided into 6 treatments (3 replicates per treatment) to receive five doses of organic Cu, which were Cu citrate (CuCit) at 0.99 (basal diet), 2.19, 4.06, 6.15, and 8.07 mg/kg, and one dose of inorganic Cu (CuSO4·5H2O at 3.15 mg/kg), for 9 weeks. The results showed appropriate Cu level (4.06 mg/kg) enhanced growth performance, improved nutritional Cu status, and downregulated Cu-transporting ATPase 1 mRNA levels in the hepatopancreas, intestine, and muscle of juvenile grass carp. Meanwhile, collagen content in fish muscle was increased after Cu intake, which was probably due to the following pathways: (1) activating CTGF/TGF-β1/Smads signaling pathway to regulate collagen transcription; (2) upregulating of La ribonucleoprotein domain family 6 (LARP6) mRNA levels to regulate translation initiation; (3) increasing proline hydroxylase, lysine hydroxylase, and lysine oxidase activities to regulate posttranslational modifications. In addition, optimal Cu group increased myofiber diameters and the frequency of myofibers with diameter >50 μm, which might be associated with upregulation of cyclin B, cyclin D, cyclin E, proliferating cell nuclear antigen, myogenic determining factor (MyoD), myogenic factor 5, myogenin (MyoG), myogenic regulatory factor 4 and myosin heavy chain (MyHC) and downregulation of myostatin mRNA levels, increasing protein levels of MyoD, MyoG and MyHC in fish muscle. Finally, based on percentage weight gain (PWG), serum ceruloplasmin (Cp) activity and collagen content in fish muscle, Cu requirements were determined as 4.74, 4.37 and 4.62 mg/kg diet (CuCit as Cu source) of juvenile grass carp, respectively. Based on PWG and Cp activity, compared to CuSO4·5H2O, the efficacy of CuCit were 131.80% and 115.38%, respectively. Our findings provide new insights into Cu supplementation to promote muscle growth in fish, and help improve the overall productivity of aquaculture.

Effects of elastase-induced emphysema on muscle and bone in mice.

Chronic obstructive pulmonary disease (COPD) causes sarcopenia and osteoporosis. However, the mechanisms underlying muscle and bone loss as well as the interactions between muscle and bone in the COPD state remain unclear. Therefore, we herein investigated the effects of the COPD state on muscle and bone in mice intratracheally administered porcine pancreatic elastase (PPE). The intratracheal administration of PPE to mice significantly reduced trabecular bone mineral density (BMD), trabecular bone volume, trabecular number, cortical BMD and cortical area. It also significantly decreased grip strength, but did not affect muscle mass or the expression of myogenic differentiation-, protein degradation- or autophagy-related genes in the soleus and gastrocnemius muscles. Among the myokines examined, myostatin mRNA levels in the soleus muscles were significantly elevated in mice treated with PPE, and negatively related to grip strength, but not bone parameters, in mice treated with or without 2 U PPE in simple regression analyses. Grip strength positively related to bone parameters in mice treated with or without PPE. In conclusion, we showed that a PPE model of COPD in mice exerts dominant effects on bone rather than skeletal muscles. Increased myostatin expression in the soleus muscles of mice in the COPD state may negatively relate to a reduction in grip strength, but not bone loss.

Dietary (−)-Epicatechin supplementation regulates myofiber development, fillet quality, and antioxidant status of Yellow River carp (Cyprinus carpio)

(−)-Epicatechin (EC) exhibits antioxidant effects and strengthens skeletal muscle growth and differentiation in mammals. However, there are no reports about the effects of EC in fish. This study evaluated the effects of EC on muscle antioxidant capacity, myofiber development, texture characteristics, and histomorphology were also investigated through feeding trial. 300 fish (16.27 ± 0.24 g) were fed four diets with different levels of EC (0, 100, 500, and 1000 mg/kg) for 60 days. The result showed that 1000 mg/kg EC supplementation enhanced antioxidant capacity and protein content in the muscle. Meanwhile, higher expression levels of proliferating cell nuclear antigen (pcna), myogenic differentiation antigen (myod), myogenin, myosin myosin heavy chain (myhc), and myosin light chain (mylc) were detected in the muscle of EC treatment, while the myostatin (mstnb) mRNA level was decreased in the EC 1000 group, the similar trends were observed in the protein expression levels of MyoD, Myogenin, MyLc (P < 0.05). Muscle hardness and pH value were increased significantly in EC 500 and 1000 groups (P < 0.05). Additionally, myofiber density, small-sized fiber percentage, and expression of genes related to AMP-activated protein kinase (AMPK) signaling pathway was promoted in the EC groups. These results indicated that dietary EC improved fillet quality and promoted myofiber development in Yellow River carp, which might be related to AMPK pathway.

Effects of Chinese yam Polysaccharides on the Muscle Tissues Development-Related Genes Expression in Breast and Thigh Muscle of Broilers.

This experiment was conducted to evaluate the effects of dietary Chinese yam polysaccharides (CYP) on myogenic differentiation 1 (MYOD1), myogenin (MYOG), and myostatin (MSTN) mRNA expression of breast and thigh muscle tissues in broilers. A total of 360 (1-day-old, gender-balanced) crossbred broilers chicks with similar body weight (BW) were randomly distributed into four groups, with three replicates in each group and each replicate included 30 broilers. The feeding trial lasted for 48 days. Experimental broilers were fed 0.00 mg/kg basal diet (control group), 250 mg/kg, 500 mg/kg, and 1000 mg/kg CYP, respectively. The results showed that CYP250 and CYP500 groups had higher thigh muscle percentage (TMP) compared to the control group (p &lt; 0.05). Meanwhile, the expression of MYOD1, MYOG mRNA in breast muscle tissues of CYP500 and CYP1000 groups was higher (p &lt; 0.05), and the expression of MSTN mRNA in thigh muscle of CYP250, CYP500, and CYP1000 groups was lower than that of the control group (p &lt; 0.05). In addition, there was no significant difference in the expression of MYOD1 mRNA in the thigh muscle tissue of each group (p &gt; 0.05). Bivariate correlation analysis showed that the expression levels of MYOD1, MYOG, and MSTN mRNA in the thigh muscle tissue of broiler chickens in the CYP500 group were positively correlated with TMP. However, the expression of MYOG mRNA in thigh muscle tissue of the CYP1000 group was negatively correlated with TMP. In general, this study indicated that appropriate dietary CYP supplementation influenced the growth and development of thigh muscle tissue in broilers by altering TMP and muscle tissue development-related genes expression. Therefore, CYP could be used as a potential feed additive to promote the development of muscle tissues in broilers.

Modification of beneficial fatty acid composition and physicochemical qualities in the muscle of sub-adult grass carp (Ctenopharyngodon idella): The role of lipids

Flesh quality, including beneficial fatty acid composition and physicochemical qualities, is the most important factor for many consumers. The actions and mechanisms of promotion of polyunsaturated fatty acid (PUFA) contents, muscle hardness, and related signalling pathways by lipids were investigated in sub-adult grass carp. In this experiment, 450 healthy grass carp (initial body weight 756.57 ± 1.28 g) were selected and randomly divided into 6 treatments, and fish in each group were fed diets with different lipid levels (0.79, 1.58, 2.72, 4.06, 5.16, and 6.33%) for 60 days. The results showed that compared with the control group (0.79% lipid diet), the optimal lipid levels in the diet increased the percentage of weight gain (PWG), feed intake (FI) and flesh rate (P < 0.05), indicating that lipids improved the growth performance of sub-adult grass carp. The optimal levels of lipids also increased muscle PUFA content, including eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) (P < 0.05), which demonstrated that lipids improved beneficial fatty acid composition in the muscle. Meanwhile, optimal lipid levels increased muscle shear force, collagen content and myofibre density (P < 0.05), which indicated that lipids improved muscle physicochemical qualities. We further explored the mechanism of lipid-increased muscle PUFA content and muscle hardness in fish. We observed that the increase in muscle PUFA content by optimal lipid levels may be related to the promotion of LC-PUFA biosynthesis and catabolism signalling pathways (LXRα/SREBP-1, Sp1, and PPARα) in fish. Meanwhile, the enhancement of muscle hardness by optimal levels of lipids may be associated with the increase in collagen content as well as the improvement of the muscle histological characteristics (diameter and density) in fish. The increase in the muscle collagen content may be related to lipids motivating collagen synthesis pathways including the TGF-β1/Smads pathway at the transcriptional level and the PI3K/Akt/LARP6 signalling pathway at the initial stage of translation. In addition, the increase in muscle fibre density may be attributed to lipid-upregulated MRFs (MyoD, MyoG and Myf5) mRNA levels and decreased myostatin (MSTN1 and MSTN2) mRNA levels (P < 0.05), which were involved in myofibre development. Collectively, lipid-improved flesh quality, including beneficial muscle fatty acid composition and physicochemical qualities, is connected with the activation of related signalling pathways and regulatory factors. Moreover, based on PWG and muscle shear force, the dietary lipid requirements for sub-adult grass carp (756–1459 g) were 4.01% and 3.65% of the diet, respectively.

Gene markers of dietary macronutrient composition and growth in the skeletal muscle of gilthead sea bream (Sparus aurata)

To increase our current knowledge on the nutritional regulation of growth and gene expression pattern in fish skeletal muscle, the effect of dietary macronutrient composition was assessed on digestibility, nutrient retention, growth performance, and the mRNA levels of key genes involved in functionality, growth and development of the skeletal muscle in gilthead sea bream (Sparus aurata). Long-term starvation decreased the expression of myogenic regulatory factors such as Myod2, Myf5, myogenin (Myog) and Myf6 in the skeletal muscle of S. aurata. The supply of high or medium protein, low carbohydrate diets enhanced growth parameters, feed efficiency ratio, feed conversion ratio and significantly upregulated myod2. However, the supply of low protein, high carbohydrate diets restricted growth and stimulated the mRNA levels of myostatin, while downregulated follistatin (fst), igf1, mtor and rps6. Microarray analysis revealed igfals, tnni2, and gadd45a as gene markers upregulated by diets enriched with protein, lipids and carbohydrates, respectively. The results of the present study show that in addition to myod2, fst, igf1, mtor and rps6, the expression levels of igfals, tnni2 and remarkably gadd45a in the skeletal muscle can be used as markers to evaluate the effect of dietary macronutrient changes on fish growth and muscle development in S. aurata.

Renal failure suppresses muscle irisin expression, and irisin blunts cortical bone loss in mice.

BackgroundChronic renal failure induces bone mineral disorders and sarcopenia. Skeletal muscle affects other tissues, including bone, by releasing myokines. However, the effects of chronic renal failure on the interactions between muscle and bone remain unclear.MethodsWe investigated the effects of renal failure on bone, muscle, and myokines linking muscle to bone using a mouse 5/6 nephrectomy (Nx) model. Muscle mass and bone mineral density (BMD) were analysed by quantitative computed tomography 8 weeks after Nx.ResultsNephrectomy significantly reduced muscle mass in the whole body (12.1% reduction, P < 0.05), grip strength (10.1% reduction, P < 0.05), and cortical BMD at the femurs of mice (9.5% reduction, P < 0.01) 8 weeks after surgery, but did not affect trabecular BMD at the femurs. Among the myokines linking muscle to bone, Nx reduced the expression of irisin, a proteolytic product of fibronectin type III domain‐containing 5 (Fndc5), in the gastrocnemius muscles of mice (38% reduction, P < 0.01). Nx increased myostatin mRNA levels in the gastrocnemius muscles of mice (54% increase, P < 0.01). In simple regression analyses, cortical BMD, but not trabecular BMD, at the femurs was positively related to Fndc5 mRNA levels in the gastrocnemius muscles of mice (r = 0.651, P < 0.05). The weekly administration of recombinant irisin to mice ameliorated the decrease in cortical BMD, but not muscle mass or grip strength, induced by Nx (6.2% reduction in mice with Nx vs. 3.3% reduction in mice with Nx and irisin treatment, P < 0.05).ConclusionsThe present results demonstrated that renal failure decreases the expression of irisin in the gastrocnemius muscles of mice. Irisin may contribute to cortical bone loss induced by renal failure in mice as a myokine linking muscle to bone.

Phytoecdysteroids Do Not Have Anabolic Effects in Skeletal Muscle in Sedentary Aging Mice.

Skeletal muscle mass and strength are lost with aging. Phytoecdysteroids, in particular 20-hydroxyecdysone (20E), increase protein synthesis in C2C12 skeletal muscle cells and muscle strength in young rats. The objective of this study was to determine whether an extract from Ajuga turkestanica (ATE), enriched in phytoecdysteroids, and 20E affect skeletal muscle mass and fiber size, fiber type, activation of the PI3K–Akt signaling pathway, and the mRNA levels of MAFbx, MuRF-1, and myostatin in sedentary aging mice. Aging male C57BL/6 mice (20 months old) received ATE, 20E, or vehicle (CT) once per day for 28 days or a single acute dose. Treatment did not alter body, muscle, or organ mass; fiber cross-sectional area; or fiber type in the triceps brachii or plantaris muscles. Likewise, protein synthesis signaling markers (i.e., phosphorylation of AktSer473 and p70S6kThr389) measured after either 28 days or acutely were unchanged. Neither ATE nor 20E treatment for 28 days affected the mRNA levels of MAFbx, MuRF-1, and myostatin. In conclusion, these data indicate that phytoecdysteroid treatment does not alter muscle mass or fiber type, nor does it activate protein synthesis signaling in the skeletal muscle of sedentary aging mice.

PO-306 The effect of four weeks hypoxic resistance training on myostatin and follistatin expression in skeletal muscle of rats

Objective Loss of skeletal muscle weight is a common phenomenon in hypoxic environment. It has been recognized that resistance training can reduce hypoxia-induced skeletal muscle atrophy, but its molecular mechanism is still unclear. Myostatin is a major factor that inhibits muscle growth and differentiation, and Follistatin can inhibit Myostatin. Therefore, this study is to clarify the effect of 4-week hypoxic resistance exercise on Myostatin and Follistatin gene expression in skeletal muscle of rats.&#x0D; Methods Twenty four 8-week-old male SD rats were randomly divided into normoxic control group (group C: 6 rats), normoxic exercise group (group R: 6 rats), hypoxic control group (group H: 6 rats) and hypoxic exercise group (group HR: 6 rats). Rats in each hypoxic group were fed in a hypoxic chamber (atmospheric hypoxia) with oxygen concentration of 12.7% (simulated 4000m altitude). Rats in each exercise group were trained according to the rat's resistance training program developed in our laboratory. After all the intervention, DEXA was used to analyze the body composition. The soleus, extensor digitorum longus and biceps brachii muscles of rats were taken and the wet weight of individual muscles was measured. The data were processed by SPSS17.0 statistical software. The expression level of skeletal muscle mRNA was expressed as "median (25-75%)" and the data of body composition and muscle wet weight were expressed as"mean±standard deviation". The differences between the groups were evaluated using a one-way analysis of variance (ANOVA) test. The significance level for the study was less than 0.05.&#x0D; Results Body composition analysis after 4 weeks of hypoxic intervention showed that the body weight of rats in group H decreased significantly (p=0.012), and the muscle mass decreased more significantly (p&lt;0.001). But resistance exercises obviously reduced the muscle atrophy (p&lt;0.01) caused by hypoxia. After analyzing the changes of the wet weight of individual muscles, it was found that the wet weight of biceps brachii in HR group was significantly higher than that in H group (p=0.048). After 4 weeks of hypoxic intervention and hypoxic resistance exercise, the expression of Myostatin mRNA in individual muscles of each group changed differently. The expression of Myostatin mRNA in soleus muscle of H group was significantly higher than that of C group (371.2%) after 4 weeks of hypoxia intervention. Myostatin mRNA expression in soleus and biceps brachii of HR group was significantly lower than that of H group (591.1% and 478.4% respectively). However, there was no significant difference in the expression level of Myostatin mRNA in the extensor digitorum longus between each group (p=0.259). The change of Follistatin mRNA expression in different groups also showed a different trend. The expression of Follistatin mRNA in soleus muscle and biceps brachii muscle was significantly different among groups (p=0.003, p=0.004, respectively). However, there was no significant difference in the expression level of Follistatin mRNA in the extensor digitorum longus between each group (p=0.734). Myostatin mRNA/Follistatin mRNA ratio (M/F) showed a more significant difference. The M/F ratio of soleus muscle in group H was significantly lower than that in group C (p&lt;0.001), but the M/F ratio in group HR was significantly higher than that in group H (p&lt;0.001). The M/F ratio of biceps brachii in group H was significantly lower than that in group C (p&lt;0.001), but the M/F ratio in group HR showed a higher trend than that in group H (p=0.051).&#x0D; Conclusions Hypoxic exposure results in an increase in Myostatin mRNA expression in skeletal muscle, but hypoxic resistance exercise reduces such an increase. On the contrary, the level of Follistatin mRNA expression in skeletal muscle decreased after hypoxic exposure, and hypoxic resistance exercise could slow down the decline. As a result, rat resistance exercise significantly slowed down hypoxia-induced muscle atrophy. In conclusion, the mutual restriction between Myostatin and Follistatin is one of the main links of resistance exercise to reduce hypoxia-induced skeletal muscle atrophy. However, the process of resistance training to reduce the hypoxia-induced skeletal muscle atrophy is very complex. There are many molecular signaling pathways involved, which need further study.

β-hydroxy-β-methyl butyrate promotes leucine metabolism and improves muscle fibre composition in growing pigs.

The aim of this study was to investigate the effects of excess leucine (Leu) vs. its metabolites α-ketoisocaproate (KIC) and β-hydroxy-β-methyl butyrate (HMB) on Leu metabolism, muscle fibre composition and muscle growth in growing pigs. Thirty-two pigs with a similar initial weight (9.55±0.19kg) were fed 1 of 4 diets for 45days: basal diet, basal diet+1.25% L-Leu, basal diet+1.25% KIC-Ca, basal diet+0.62% HMB-Ca. Results indicated that relative to the basal diet and HMB groups, Leu and KIC groups exhibited increased Leu concentrations and decreased concentrations of isoleucine, valine and EAAs in selected muscle (p<0.05) and had lower mRNA levels of MyHC I and higher expression of MyHC IIx/IIb (p<0.05), and there was no significant difference between the basal and HMB-supplemented groups. Moreover, the mRNA expression levels of AMPKα and UCP3 were higher but the myostatin mRNA levels were lower in the soleus muscle of the HMB group than those from other groups (p<0.05). These findings demonstrated that doubling dietary Leu content exerted growth-depressing effects in growing pigs; dietary KIC supplementation induced muscular branched-chain amino acid imbalance and promoted muscle toward a more glycolytic phenotype; while dietary HMB supplementation promoted the generation of more oxidative muscle types and increased muscle growth specially in oxidative skeletal muscle, and these effects of HMB might be associated with the AMPKα-Sirt1-PGC-1α axis and mitochondrial biogenesis.

MiRNA-23a/27a attenuates muscle atrophy and renal fibrosis through muscle-kidney crosstalk.

BackgroundThe treatment of muscle wasting is accompanied by benefits in other organs, possibly resulting from muscle–organ crosstalk. However, how the muscle communicates with these organs is less understood. Two microRNAs (miRs), miR‐23a and miR‐27a, are located together in a gene cluster and regulate proteins that are involved in the atrophy process. MiR‐23a/27a has been shown to reduce muscle wasting and act as an anti‐fibrotic agent. We hypothesized that intramuscular injection of miR‐23a/27a would counteract both muscle wasting and renal fibrosis lesions in a streptozotocin‐induced diabetic model.MethodsWe generated an adeno‐associated virus (AAV) that overexpresses the miR‐23a∼27a∼24‐2 precursor RNA and injected it into the tibialis anterior muscle of streptozotocin‐induced diabetic mice. Muscle cross‐section area (immunohistology plus software measurement) and muscle function (grip strength) were used to evaluate muscle atrophy. Fibrosis‐related proteins were measured by western blot to monitor renal damage. In some cases, AAV‐GFP was used to mimic the miR movement in vivo, allowing us to track organ redistribution by using the Xtreme Imaging System.ResultsThe injection of AAV‐miR‐23a/27a increased the levels of miR‐23a and miR‐27a as well as increased phosphorylated Akt, attenuated the levels of FoxO1 and PTEN proteins, and reduced the abundance of TRIM63/MuRF1 and FBXO32/atrogin‐1 in skeletal muscles. It also decreased myostatin mRNA and protein levels as well as the levels of phosphorylated pSMAD2/3. Provision of miR‐23a/27a attenuates the diabetes‐induced reduction of muscle cross‐sectional area and muscle function. Curiously, the serum BUN of diabetic animals was reduced in mice undergoing the miR‐23a/27a intervention. Renal fibrosis, evaluated by Masson trichromatic staining, was also decreased as were kidney levels of phosphorylated SMAD2/3, alpha smooth muscle actin, fibronectin, and collagen. In diabetic mice injected intramuscularly with AAV‐GFP, GFP fluorescence levels in the kidneys showed linear correlation with the levels in injected muscle when examined by linear regression. Following intramuscular injection of AAV‐miR‐23a∼27a∼24‐2, the levels of miR‐23a and miR‐27a in serum exosomes and kidney were significantly increased compared with samples from control virus‐injected mice; however, no viral DNA was detected in the kidney.ConclusionsWe conclude that overexpression of miR‐23a/27a in muscle prevents diabetes‐induced muscle cachexia and attenuates renal fibrosis lesions via muscle–kidney crosstalk. Further, this crosstalk involves movement of miR potentially through muscle originated exosomes and serum distribution without movement of AAV. These results could provide new approaches for developing therapeutic strategies for diabetic nephropathy with muscle wasting.

Experimental Increases in Foraging Costs Affect Pectoralis Muscle Mass and Myostatin Expression in Female, but Not Male, Zebra Finches (Taeniopygia guttata).

Skeletal muscle remodeling is an important component of phenotypic flexibility in birds and impacts organismal metabolism and performance, which could potentially influence fitness. One regulator of skeletal muscle remodeling is myostatin, an autocrine/paracrine muscle growth inhibitor that may be down-regulated under conditions promoting heavier muscle masses. In this study, we employed protocols requiring hovering while foraging to increase foraging costs and modify phenotypes of zebra finches (Taeniopygia guttata). We examined the effects of high-cost foraging (HF) on skeletal muscle masses and used real-time reverse-transcription polymerase chain reaction and Western blots to measure gene and protein expression of myostatin and its metalloproteinase activators tolloid-like proteases TLL-1 and TLL-2 in pectoralis muscle. Female finches average shorter wings and higher wing loading than males, so increased flight costs might be expected to disproportionately affect females. Indeed, HF female finches exhibited reduced total fat masses, increased pectoralis muscle masses, and lower myostatin protein levels than controls. Male finches showed no significant differences in pectoralis muscle masses or myostatin protein levels between HF and control birds. Myostatin, TLL-1, and TLL-2 messenger RNA (mRNA) expression remained stable between treatments for both female and male finches. Myostatin mRNA and protein levels showed variable directions of correlations with pectoralis mass residuals among treatments. Thus, these data offer only mixed support for a regulatory role for myostatin in mediating the flexibility of pectoralis muscle phenotypes of small birds.

Agaro-Oligosaccharides Prevent Myostatin Hyperexpression and Myosin Heavy Chain Protein Degradation in C2C12 Myotubes Induced by Tumor Necrosis Factor-&amp;lt;i&amp;gt;α&amp;lt;/i&amp;gt;

Myostatin is a major factor involved in the regulation of skeletal muscle protein mass. High myostatin levels have been associated with an increase in myotube shrinkage. Enhanced myostatin expression is caused by pro-catabolic reactions involving compounds such as tumor necrosis factor (TNF)-α. The present study investigated the effects of agaro-oligosaccharides (AOSs) on hypercatabolism of myotubes exposed to TNF-α. C2C12 myotubes exposed to TNF-α in the presence or absence of AOSs. Myotube exposure to TNF-α resulted in a reduction in the amount of myosin heavy chain (MyHC) protein and a decrease in myotube diameter, which was associated with increased myostatin mRNA expression. AOSs prevented TNF-α-induced MyHC protein loss and restored normal myostatin mRNA levels, with agarobiose and agarotetraose effectively suppressing the hyperexpression of the mRNA. In addition, expression levels of the known myostatin inhibitors, latent transforming growth factor beta binding protein 3 (Ltbp3) and growth and differentiation factor-associated serum protein 1 (Gasp1) mRNAs, decreased more in TNF-α-induced myotubes than in the TNF-α-free control, possibly resulting in myostatin upregulation. However, AOSs restored nearly normal expression levels of Ltbp3 and Gasp1 mRNA, potentially suppressing myostatin expression. These findings suggest that AOSs could prevent myotube shrinkage induced by TNF-α.

Effects of dietary methionine on breast muscle growth, myogenic gene expression and IGF-I signaling in fast- and slow-growing broilers

This study investigated the responses of fast- (FG) and slow- (SG) growing broilers to dietary methionine (Met) status. The broilers were subjected to low (LM, 0.38 and 0.28 g/100 g), adequate (AM, 0.51 and 0.42 g/100 g) and high (HM, 0.65 and 0.52 g/100 g) Met during 1–21 and 22–42 d, respectively. Compared with the LM diets, the AM and HM diets increased body weight gain only in the FG broilers. The HM diets increased breast muscle yield only in the FG broilers, although insulin-like growth factor-I (IGF-I) concentration was increased in both strains of broilers. The HM diets increased mRNA levels of myogenic regulatory factors (MRF4, Myf5) and myocyte enhancer factor 2 (MEF2A and MEF2B) in the FG broilers, and increased MEF2A and decreased myostatin mRNA level in the SG broilers. Extracellular signal-regulated kinase (ERK) phosphorylation of breast muscle was increased by the HM diets in both strains of broilers, but mechanistic target of rapamycin (mTOR) phosphorylation was increased by the AM and HM diets only in the FG broilers. These results reflect a strain difference in broiler growth and underlying mechanism in response to dietary Met.

A comparative examination of cortisol effects on muscle myostatin and HSP90 gene expression in salmonids

Cortisol, the primary corticosteroid in teleost fishes, is released in response to stressors to elicit local functions, however little is understood regarding muscle-specific responses to cortisol in these fishes. In mammals, glucocorticoids strongly regulate the muscle growth inhibitor, myostatin, via glucocorticoid response elements (GREs) leading to muscle atrophy. Bioinformatics methods suggest that this regulatory mechanism is conserved among vertebrates, however recent evidence suggests some fishes exhibit divergent regulation. Therefore, the aim of this study was to evaluate the conserved actions of cortisol on myostatin and hsp90 expression to determine if variations in cortisol interactions have emerged in salmonid species. Representative salmonids; Chinook salmon (Oncorhynchus tshawytscha), cutthroat trout (Oncorhynchus clarki), brook trout (Salvelinus fontinalis), and Atlantic salmon (Salmo salar); were injected intraperitoneally with a cortisol implant (50μg/g body weight) and muscle gene expression was quantified after 48h. Plasma glucose and cortisol levels were significantly elevated by cortisol in all species, demonstrating physiological effectiveness of the treatment. HSP90 mRNA levels were elevated by cortisol in brook trout, Chinook salmon, and Atlantic salmon, but were decreased in cutthroat trout. Myostatin mRNA levels were affected in a species, tissue (muscle type), and paralog specific manner. Cortisol treatment increased myostatin expression in brook trout (Salvelinus) and Atlantic salmon (Salmo), but not in Chinook salmon (Oncorhynchus) or cutthroat trout (Oncorhynchus). Interestingly, the VC alone increased myostatin mRNA expression in Chinook and Atlantic salmon, while the addition of cortisol blocked the response. Taken together, these results suggest that cortisol affects muscle-specific gene expression in species-specific manners, with unique Oncorhynchus-specific divergence observed, that are not predictive solely based upon mammalian stress responses.

Akt/mTOR pathway contributes to skeletal muscle anti-atrophic effect of aerobic exercise training in heart failure mice

BackgroundExercise intolerance is one of the main clinical symptoms of heart failure (HF) and is associated with skeletal muscle wasting due to an imbalance between proteolysis and protein synthesis. In this study, we tested whether aerobic exercise training (AET) would counteract skeletal muscle atrophy by activating IGF-I/Akt/mTOR pathway in HF mice. MethodsSympathetic hyperactivity induced HF mice were assigned into 8-week moderate intensity AET. Untrained wild type and HF mice were used as control. Soleus cross sectional area was evaluated by histochemistry and motor performance by rotarod. 26S proteasome activity was assessed by fluorimetric assay, and components of IGF-I/Akt/mTOR pathway or myostatin pathway by qRT-PCR or immunoblotting. A different subset of mice was used to evaluate the relative contribution of mTOR inhibition (rapamycin) or activation (leucine) on AET-induced changes in muscle mass regulation. ResultsAET prevented exercise intolerance and impaired motor performance in HF mice. These effects were associated with attenuation of soleus atrophy. Rapamycin treatment precluded AET effects on soleus mass in HF mice suggesting the involvement of IGF signaling pathway in this response. In fact, AET increased IGF-I Ea and IGF-I Pan mRNA levels, while it reduced myostatin and Smad2 mRNA levels in HF mice. At protein levels, AET prevented reduced expression levels of IGF-I, pAkt (at basal state), as well as, p4E-BP1 and pP70S6K (leucine-stimulated state) in HF mice. Additionally, AET prevented 26S proteasome hyperactivity in HF mice. ConclusionsTaken together, our data provide evidence for AET-induced activation of IGF-I/Akt/mTOR signaling pathway counteracting HF-induced muscle wasting.

Loss of Ovarian Function Results in Increased Loss of Skeletal Muscle in Arthritic Rats.

We studied the effects of loss of ovarian function (ovariectomy) on muscle mass of gastrocnemius and the mRNA levels of IGF-1, atrogin-1, MuRF-1, and myostatin in an experimental model of rheumatoid arthritis in rats. We randomly allocated 24 female Wistar rats (9 weeks, 195.3 ± 17.4 grams) into four groups: control (CT-Sham; n = 6); rheumatoid arthritis (RA; n = 6); ovariectomy without rheumatoid arthritis (OV; n = 6); ovariectomy with rheumatoid arthritis (RAOV; n = 6). We performed the ovariectomy (OV and RAOV) or Sham (CT-Sham or RA) procedures at the same time, fifteen days before the rheumatoid arthritis induction. The RA and RAOV groups were immunized and then were injected with Met-BSA in the tibiotarsal joint. After 15 days of intra-articular injections the animals were euthanized. We evaluated the external manifestations of rheumatoid arthritis (perimeter joint) as well as animal weight, and food intake throughout the study. We also analyzed the cross-sectional areas (CSA) of gastrocnemius muscle fibers in 200 fibers (H&E method). In the gastrocnemius muscle, we analyzed mRNA expression by quantitative real time PCR followed by the Livak method (ΔΔCT). The rheumatoid arthritis induced reduction in CSA of gastrocnemius muscle fibers. The RAOV group showed a lower CSA of gastrocnemius muscle fibers compared to RA and CT-Sham groups. Skeletal muscle IGF-1 mRNA increased in arthritics and ovariectomized rats. The increased IGF-1 mRNA was higher in OV groups than in the RA and RAOV groups. Antrogin-1 mRNA also increased in the gastrocnemius muscle of arthritic and ovariectomized rats. However, the increased atrogin-1 mRNA was higher in RAOV groups than in the RA and OV groups. Gastrocnemius muscle MuRF-1 mRNA increased in the OV and RAOV groups, but not in the RA and Sham groups. However, the RAOV group showed higher MuRF-1 mRNA than the OV group. The myostatin gene expression was similar in all groups. Loss of ovarian function results in increased loss of skeletal muscle-related ubiquitin ligases atrogin-1 and MuRF-1 in arthritic rats.