Muscle Gene Expression Research Articles

Effect of the Aerobic and Resistance Training on Follistatin-Like 1 and Leukemia Inhibitory Factor Muscle Gene Expression in Rats Fed With a High-Fat Diet

Background: Follistatin-like 1 (FSTL-1) and leukemia inhibitory factor (LIF) are two myokines that are affected by overweight and have inflammatory and damaging effects. Considering that exercise reduces excess weight, this study aimed to evaluate the effect of aerobic and resistance training on FSTL-1 and LIF muscle gene expression in rats fed with a high-fat diet. Materials and Methods: In this experimental study, 32 rats were randomly divided into healthy control, obese control, obese+aerobic exercise, and obese+resistance exercise groups. The training was performed for 4 weeks at aerobic moderate intensity (50-65% VO2max). For resistance training, rats were also trained to climb the ladder (height 110 cm, slope 80%, and the distance between the bars of the ladder 2 cm), which is based on the determination of one repetition maximum. A high-fat diet was prepared with 40% fat, 13% protein, and 47% carbohydrates and continued until the rats reached the obesity range. The tissue sample was taken from the gluteus muscle. Results: The expression of FSTL-1 and LIF in the obese control group increased significantly compared to the healthy control group (P=0.044 and P=0.039, respectively). The expression of FSTL-1 and LIF in the resistance training group significantly decreased in comparison to the obese control group (P=0.049 and P=0.046, respectively). There was no significant difference between the aerobic exercise group and the obese control group (P=0.053 and P=0.059, respectively). However, a significant difference was observed between aerobic and resistance training groups in terms of FSTL-1 (P=0.042). Conclusion: Resistance exercise seems to have a greater and better effect on FSTL-1 and LIF in the muscles of obese samples compared to aerobic exercise.

The time-course of cancer cachexia onset reveals biphasic transcriptional disruptions in female skeletal muscle distinct from males

BackgroundCancer-cachexia (CC) is a debilitating condition affecting up to 80% of cancer patients and contributing to 40% of cancer-related deaths. While evidence suggests biological sex differences in the development of CC, assessments of the female transcriptome in CC are lacking, and direct comparisons between sexes are scarce. This study aimed to define the time course of Lewis lung carcinoma (LLC)-induced CC in females using transcriptomics, while directly comparing biological sex differences.ResultsWe found the global gene expression of the gastrocnemius muscle of female mice revealed biphasic transcriptomic alterations, with one at 1 week following tumor allograft and another during the later stages of cachexia development. The early phase was associated with the upregulation of extracellular-matrix pathways, while the later phase was characterized by the downregulation of oxidative phosphorylation, electron transport chain, and TCA cycle. When DEGs were compared to a known list of mitochondrial genes (MitoCarta), ~ 47% of these genes were differently expressed in females exhibiting global cachexia, suggesting transcriptional changes to mitochondrial gene expression happens concomitantly to functional impairments previously published. In contrast, the JAK-STAT pathway was upregulated in both the early and late stages of CC. Additionally, we observed a consistent downregulation of Type-II Interferon signaling genes in females, which was associated with protection in skeletal muscle atrophy despite systemic cachexia. Upregulation of Interferon signaling was noted in the gastrocnemius muscle of cachectic and atrophic male mice. Comparison of female tumor-bearing mice with males revealed ~ 70% of DEGs were distinct between sexes in cachectic animals, demonstrating dimorphic mechanisms of CC.ConclusionOur findings suggest biphasic disruptions in the transcriptome of female LLC tumor-bearing mice: an early phase associated with ECM remodeling and a late phase, accompanied by the onset of systemic cachexia, affecting overall muscle energy metabolism. Notably, ~ 2/3 of DEGs in CC are biologically sex-specific, providing evidence of dimorphic mechanisms of cachexia between sexes. Downregulation of Type-II Interferon signaling genes appears specific to CC development in females, suggesting a new biological sex-specific marker of CC not reliant on the loss of muscle mass, that might represent a protective mechanism against muscle loss in CC in female mice.

Effects of linseed oil supplementation duration on fatty acid profile and fatty acid metabolism-related genes in the muscles of Chinese crested white ducks

Meat rich in polyunsaturated fatty acids is considered beneficial to health. Supplementing the diet with linseed oil promotes the deposition of polyunsaturated fatty acids (PUFAs) in poultry, a conclusion that has been confirmed multiple times in chicken meat. However, fewer studies have focused on the effects of dietary fatty acids on duck meat. Therefore, this study aims to evaluate the effects of the feeding time of a linseed oil diet on duck meat performance and gene expression, including meat quality performance, plasma biochemical indicators, fatty acid profile, and gene expression. For this study, we selected 168 Chinese crested ducks at 28 days old and divided them into three groups, with 56 birds in each group. The linseed oil content in the different treatment groups was as follows: the control group (0% flaxseed oil), the 14d group (2% linseed oil), and the 28d group (2% linseed oil). Ducks in the two experimental groups were fed a linseed oil diet for 28 and 14 days at 28 and 42 days of age, respectively. The results showed that linseed oil had no negative effect on duck performance (slaughter rate, breast muscle weight, and leg muscle weight) or meat quality performance (pH, meat color, drip loss, and shear force) (P > 0.05). The addition of linseed oil in the diet increased plasma total cholesterol and high-density lipoprotein cholesterol levels (P < 0.05), while decreasing triglyceride content (P < 0.05). Furthermore, the supplementation of linseed oil for four weeks affected the composition of muscle fatty acids. Specifically, levels of α-linolenic acid, eicosapentaenoic acid, and docosahexaenoic acid were increased (P < 0.05), while eicosatetraenoic acid content was negatively correlated with flaxseed oil intake (P < 0.05). qRT-PCR analysis further revealed that the expression of FATP1, FABP5, and ELOVL5 genes in the breast muscle, as well as FABP3 and FADS2 genes in the thigh muscle, increased after four weeks of linseed oil supplementation (P < 0.05). However, after two weeks of feeding, CPT1A gene expression inhibited fatty acid deposition, suggesting an increase in fatty acid oxidation (P < 0.05). Overall, the four-week feeding time may be a key factor in promoting the deposition of n-3 PUFAs in duck meat. However, the limitation of this study is that it remains unknown whether longer supplementation time will continue to affect the deposition of n-3 PUFAs. Further experiments are needed to explain how prolonged feeding of linseed oil will affect the meat quality traits and fatty acid profile of duck meat.

Deletion of Nmnat1 in Skeletal Muscle Leads to the Reduction of NAD+ Levels but Has No Impact on Skeletal Muscle Morphology and Fiber Types.

Nicotinamide adenine dinucleotide (NAD+) is a coenzyme that mediates many redox reactions in energy metabolism. NAD+ is also a substrate for ADP-ribosylation and deacetylation by poly (ADP-ribose) polymerase and sirtuin, respectively. Nicotinamide mononucleotide adenylyltransferase 1 (Nmnat1) is a NAD+ biosynthesizing enzyme found in the nucleus. Recent research has shown that the maintaining NAD+ levels is critical for sustaining muscle functions both in physiological and pathological conditions. However, the role of Nmnat1 in skeletal muscle remains unexplored. In this study, we generated skeletal muscle-specific Nmnat1 knockout (M-Nmnat1 KO) mice and investigated its role in skeletal muscle. We found that NAD+ levels were significantly lower in the skeletal muscle of M-Nmnat1 KO mice than in control mice. M-Nmnat1 KO mice, in contrast, had similar body weight and normal muscle histology. Furthermore, the distribution of muscle fiber size and gene expressions of muscle fiber type gene expression were comparable in M-Nmnat1 KO and control mice. Finally, we investigated the role of Nmnat1 in muscle regeneration using cardiotoxin-induced muscle injury model, but muscle regeneration appeared almost normal in M-Nmnat1 KO mice. These findings imply that Nmnat1 has a redundancy in the pathophysiology of skeletal muscle.

The isolated effects of local cold application on proteolytic and myogenic signaling

Post-exercise cooling studies reveal inhibitory effects on markers of skeletal muscle growth. However, the isolated effect of local cold application has not been adequately addressed. It is unclear if the local cold or the combination of local cold and exercise is driving negatively altered skeletal muscle gene expression. The purpose was to determine the effects of a 4 h local cold application to the vastus lateralis on the myogenic and proteolytic response. Participants (n = 12, 27 ± 6 years, 179 ± 9 cm, 82.8 ± 13.0 kg, 18.4 ± 7.1 %BF) rested with a thermal wrap placed on each leg with either circulating cold fluid (10 °C, COLD) or no fluid circulation (room temperature, RT). Muscle samples were collected to quantify mRNA (RT-qPCR) and proteins (Western Blot) associated with myogenesis and proteolysis. Temperatures in COLD were lower than RT at the skin (13.2 ± 1.0 °C vs. 34.8 ± 0.9 °C; p < 0.001) and intramuscularly (20.5 ± 1.3 °C vs. 35.6 ± 0.8 °C, p < 0.001). Myogenic-related mRNA, MYO-G and MYO-D1, were lower in COLD (p = 0.001, p < 0.001, respectively) whereas myogenic-mRNA, MYF6, was greater in COLD (p = 0.002). No other myogenic associated genes were different between COLD and RT (MSTN, p = 0.643; MEF2a, p = 0.424; MYF5, p = 0.523; RPS3, p = 0.589; RPL3-L, p = 0.688). Proteolytic-related mRNA was higher in COLD (FOXO3a, p < 0.001; Atrogin-1, p = 0.049; MURF-1, p < 0.001). The phosphorylation:total protein ratio for the translational repressor of muscle mass, 4E-BP1Thr37/46, was lower in COLD (p = 0.043), with no differences in mTORser2448 (p = 0.509) or p70S6K1Thr389 (p = 0.579). Isolated local cooling over 4 h exhibits inhibited myogenic and higher proteolytic skeletal muscle molecular response.

The pyruvate kinase of the whiteleg shrimp Litopenaeus vannamei: Gene structure and responses to short term hypoxia

The shrimp Litopenaeus vannamei is the main farmed crustacean worldwide. This shrimp suffers environmental changes in oxygen availability that affect its energy metabolism. Pyruvate kinase (PK) catalyzes the last reaction of glycolysis and is key for the regulation of glycolysis and gluconeogenesis. There is ample knowledge about mammalian PK, but in crustaceans, the information is very scarce. In this study, we analyzed in silico the structures of the PK gene and protein. Also, the effects of hypoxia on gene expression, enzymatic activity, glucose, and lactate in hepatopancreas and muscle were analyzed. The PK gene is 15,103 bp and contains 11 exons and 10 introns, producing four mRNA variants by alternative splicing and named PK1, PK2, PK3 and PK4, that results in two proteins with longer C-terminus and two with a 12 bp insertion. The promoter contains putative binding sites for transcription factors (TF) that are typically involved in stress responses. The deduced amino acid sequences contain the classic domains, binding sites for allosteric effectors and potential reversible phosphorylation residues. Protein modeling indicates a homotetramer with highly conserved structure. The effect of hypoxia for 6 and 12 h showed tissue-specific patterns, with higher expression, enzyme activity and lactate in muscle, but higher glucose in hepatopancreas. Changes in response to hypoxia were detected at 12 h in expression with induction in muscle and reduction in hepatopancreas, while enzyme activity was maintained, and glucose and lactate decreased. These results show rapid changes in expression and metabolites, while enzyme activity was maintained to cope with short-term hypoxia.

Zinc supplementation and ractopamine hydrochloride impact gene expression of zinc transporters in finishing beef steers

Zinc is a trace mineral of interest for optimizing growth in feedlot cattle due to its roles in many physiological functions, including growth. Twenty-four Angus-cross steers (467 ± 13 kg) were used to assess the effects of supplemental Zn and ractopamine hydrochloride (RAC) on trace mineral concentrations and muscle gene expression. Four GrowSafe-equipped pens were randomly assigned to treatments (1 pen of six steers/treatment): 0 (CON), 60 (LOW), 120 (MED) or 180 (HI) mg supplemental Zn/kg DM (Availa-Zn, Zinpro). Dietary Zn treatments were initiated on d 0 and RAC supplementation (300 mg·steer·-1·d-1; Actogain45, Zoetis) began on d 53. Blood, liver and muscle (longissimus thoracis) samples were collected from all steers on d -4, 48, and 67. The LOW treatment was removed from gene expression analyses due to &amp;lt; 3 steers being represented for 14 of 22 genes. Data were analyzed using ProcMixed of SAS with the fixed effect of treatment and steer as the experimental unit; orthogonal linear and quadratic contrast statements were used to compare treatments. On d 48 and 67, there were linear and quadratic trends for plasma Zn to be greater in Zn-supplemented steers than CON (P ≤ 0.10). On d 48, there was a tendency for a quadratic decrease on the expression of SLC30A4 (P ≤ 0.07) but no other differences due to treatment. On d 67, several genes involved in Zn transport and storage (MTA1, SLC39A7, SLC39A8, SLC39A9, SLC39A10, SLC39A13) were decreased (P ≤ 0.08), suggesting increased growth influences intracellular Zn trafficking and demands.

33-OR: Epigenetic Reader BRD4 Directs Fast-Twitch Myofiber Gene Programs Influencing Skeletal Muscle Endurance Capacity and Systemic Glucose Homeostasis

Skeletal myofibers exhibit remarkable diversity and plasticity in contractile and metabolic functions and play a key role in controlling systemic metabolism. Previous studies of muscle fiber-type switching have centered on the specification of slow-twitch myofibers. However, mechanisms that drive fast-twitch myofibers formation remain unclear. Here, we showed that BRD4, an epigenetic reader enriched in fast myofibers, plays a critical role in controlling fast-twitch myofiber identity as well as systemic glucose homeostasis. BRD4 levels are associated with fast-twitch myofiber gene expression in human muscle. Skeletal muscle-specific ablation of BRD4 in mice (Brd4f/fHSA-Cre, BRD4 mKO) resulted in downregulation of the fast-twitch myofiber gene program, increased number (~100%) of type I myofibers and mitochondrial oxidative capacity. This led to enhanced muscle endurance capacity. In adult mice, acute and inducible BRD4 deletion using a tamoxifen-inducible Cre elicited a similar fast-to-slow fiber type switch in muscle. Mechanically, genome-wide ChIP-seq and mRNA-seq analyses revealed that BRD4 directly binds to and activates transcriptional of fast-twitch myofiber genes through cooperating with MEF2. Furthermore, mice lacking muscle BRD4 were protected against high-fat-diet (HFD)-induced obesity, glucose intolerance and insulin resistance. HFD-fed BRD4 mKO mice had significantly less fat mass (∼50%) compared with HFD-fed WT controls. Histological and biochemical measurements confirmed a pronounced (~60%) decrease in fat deposition in BRD4 mKO livers. Thus, our results uncover a previously unrecognized mechanism underlying the determination of fast-twitch myofiber and provide new therapeutic opportunities for driving muscle fiber-type switch to combat metabolic diseases. Disclosure Z.Zhou: None. L.Liu: None. Z.Xu: None. T.Fu: None. L.Yang: None. M.Yan: None. Z.Gan: None.

458-P: Proinflammatory Macrophage Phenotype Modulates Vascular Smooth Muscle Cell Differentiation via the Histone Demethylase JMJD3

Patients with diabetes are at increased risk of a multitude of vascular diseases, including atherosclerosis, which is driven by macrophage-mediated inflammation. While it is known that atherosclerosis is characterized by a proliferative, synthetic smooth muscle phenotype that underlies disease, the exact role of other cell types, namely myeloid cells and their interaction with smooth muscle cells is less understood. Furthermore, how diabetes modifies atherosclerotic severity is another important question. To address this, we performed an epigenetic superarray to identify key chromatin modifying enzymes in vascular smooth muscle cells (vSMC) from normal diet (ND) and diabetic-induced obese (DIO) aortas. We identified the histone demethylase, JMJD3, which was significantly decreased in DIO compared to ND vSMC (p&amp;lt;0.05). Furthermore, in our single cell RNA-seq data set from human atherosclerotic lesions, JMJD3 was expressed in smooth muscle cells. It is known that macrophages in atherosclerotic lesions exhibit a pro-inflammatory phenotype. To model this pro-inflammatory process, we stimulated bone marrow derived macrophages with LPS and found that supernatant from LPS-stimulated BMDMs significantly inhibited smooth muscle specific gene expression (SMA, SM22, CNN1, SM-MHC) as well as expression of JMJD3 compared to control media (p&amp;lt;0.05). Additionally, siRNA-mediated knockdown of JMJD3 in vSMC reduced SMC-specific gene expression, and treatment of mouse aortic vSMC with the JMJD3 selective inhibitor, GSK4J, decreased smooth muscle gene expression (p&amp;lt;0.05). Our data shows that pro-inflammatory macrophages drive reduced vascular smooth muscle gene expression and that JMJD3 in SMC is a regulator of this process. These results implicate JMJD3 in macrophage-smooth muscle cell crosstalk in the pro-inflammatory state and identify it as a potential therapeutic target in the management of atherosclerosis. Disclosure K.D.Mangum: None. A.Joshi: None. P.Mazumder: None. S.Wolf-fortune: None. E.Barrett: None. B.Moore: None. F.M.Davis: None. K.Gallagher: None. Funding National Institutes of Health (1F32DK131799-01); American College of Surgeons; Vascular &amp; Endovascular Surgery Society

High-Intensity Interval Training’s Effect on Cognitive Functions

HIIT has been known to improve cognitive function. In addition to its effect on the hippocampal area. After HIIT training, BDNF levels also increase in the spinal cord, cerebellum and several cortical areas through increasing levels of Insulin-like Growth Factor-1 (IGF-1). In neurons, BDNF is present not only in the cytoplasm, but also near the dendritic spines, which influences their development. BDNF stimulates the process of neuroplasticity, which is manifested in neurogenesis, stimulation of the plasticity of serotoninergic, dopaminergic, cholinergic or noradrenergic neurons, dendritogenesis, and synaptogenesis. Moreover, BDNF facilitates the growth and survival of neurons and microglial cells. It also participates in cell differentiation, potentiation of signal transmission, induction, and maintenance of long-term potentiation of the synapse enhancement. Because of these properties, BDNF enhances cognition and takes part in emotional processes, spatial orientation and learning, as well as body coordination. Evidence suggests that IGF-1 is a major determinant of the effect of physical exercise on BDNF levels and thus on cognition more generally. There is an upregulation of Fndc5 gene expression in skeletal muscle and an increase in irisin after prolonged resistance training in rats and humans following HIIT training. When hippocampal Fndc5 was upregulated during exercise, BDNF and other neuroprotective genes were also activated in the rat hippocampus. Exercise-induced adult hippocampal neurogenesis is associated with increased Fndc5 and BDNF genes thus enhancing cognition. Then, stroke is associated with neuroinflammation that affects the processes of neuroplasticity in the lesion core, penumbra and small areas such as the spinal cord.

Thyroarytenoid Oxidative Metabolism and Synaptic Signaling Dysregulation in the Female Pink1-/- Rat.

Vocal dysfunction, including hypophonia, in Parkinson disease (PD) manifests in the prodromal period and significantly impacts an individual's quality of life. Data from human studies suggest that pathology leading to vocal deficits may be structurally related to the larynx and its function. The Pink1-/- rat is a translational model used to study pathogenesis in the context of early-stage mitochondrial dysfunction. The primary objective of this work was to identify differentially expressed genes in the thyroarytenoid muscle and examine the dysregulated biological pathways in the female rat. RNA sequencing was used to determine thyroarytenoid (TA) muscle gene expression in adult female Pink1-/- rats compared with controls. A bioinformatic approach and the ENRICHR gene analysis tool were used to compare the sequencing dataset with biological pathways and processes, disease relationships, and drug-repurposing compounds. Weighted Gene Co-expression Network Analysis was used to construct biological network modules. The data were compared with a previously published dataset in male rats. Significant upregulated pathways in female Pink1-/- rats included fatty acid oxidation and muscle contraction, synaptic transmission, and neuromuscular processes. Downregulated pathways included anterograde transsynaptic signaling, chemical synaptic transmission, and ion release. Several drug treatment options including cetuximab, fluoxetine, and resveratrol are hypothesized to reverse observed genetic dysregulation. Data presented here are useful for identifying biological pathways that may underlie the mechanisms of peripheral dysfunction including neuromuscular synaptic transmission to the TA muscle. These experimental biomarkers have the potential to be targeted as sites for improving the treatment for hypophonia in early-stage PD. NA Laryngoscope, 133:3412-3421, 2023.

Dietary ribose supplementation improves flesh quality through purine metabolism in gibel carp (Carassius auratus gibelio)

Since the aquaculture industry is currently observing a deterioration in the flesh quality of farmed fish, the use of nutrients as additives to improve the flesh quality of farmed fish species is a viable strategy. The aim of this study was to investigate the effect of dietary D-ribose (RI) on the nutritional value, texture and flavour of gibel carp (Carassius auratus gibelio). Four diets were formulated containing exogenous RI at 4 gradient levels: 0 (Control), 0.15% (0.15RI), 0.30% (0.30RI) and 0.45% (0.45RI). A total of 240 fish (150±0.31g) were randomly distributed into 12 fibreglass tanks (150L per tank). Triplicate tanks were randomly assigned to each diet. The feeding trial was carried out in an indoor recirculating aquaculture system for 60d. After the feeding trial, the muscle and liver of gibel carp were analysed. The results showed that RI supplementation did not result in any negative impact on the growth performance and 0.30RI supplementation significantly increased the whole-body protein content compared to the control group. The contents of collagen and glycogen in muscle were enhanced by RI supplementation. The alterations in the flesh indicated that RI supplementation improved the texture of the flesh in terms of its water-holding capacity and hardness, therefore improving the taste. Dietary RI facilitated the deposition of amino acids and fatty acids in the muscle that contributed to the meaty taste and nutritional value. Furthermore, a combination of metabolomics and expression of key genes in liver and muscle revealed that 0.30RI activated the purine metabolism pathways by supplementing the substrate for nucleotide synthesis and thereby promoting the deposition of flavour substance in flesh. This study offers a new approach for providing healthy, nutritious and flavourful aquatic products.

Effect of crossbreeding and sex on slaughter performance and meat quality in Xingguo gray goose based on multiomics data analysis

Here, we examined the effects of crossbreeding and sex on growth performance, slaughter performance, and meat quality in Xingguo gray (XG) goose, using transcriptomic and metabolomic techniques. The experiment was conducted using 400 goslings (1-day old) of 2 genotypes: the XG breed and its ternary hybrids [F2 geese; (XG Goose♂×Yangzhou Goose♀)♀×Shitou Goose♂]. The goslings were divided into 4 groups: female XG, male XG, female F2 geese, and male F2 geese, and growth parameters were examined at 70 d of age, using 30 birds from each group. Following slaughter, samples of breast and thigh muscles were collected from each group for chemical, metabolome, and transcriptome analyses. Growth rate, live body and slaughter weights, meat chemical composition, and muscle fiber diameter were affected by crossbreeding and sex. Crossbreeding significantly improved the dressing percentage, semieviscerated rate, eviscerated yield, and abdominal fat yield of XG geese. To clarify the potential regulatory network affected by crossbreeding and sex, we used RNA-seq and nontargeted metabolomics to detect changes in male and female goose breast muscle. The transcriptome results showed that there were 534, 323, 297, and 492 differently expressed genes (DEGs) among the 4 comparison groups (XG-Female vs. F2-Female, XG-Male vs. F2-Male, F2-Male vs. F2-Female, and XG-Male vs. XG-Female, respectively) that were mainly related to muscle growth and development and fatty acid metabolism pathways. A total of 141 significantly differentially accumulated metabolites (DAMs) were enriched in serine and threonine, propionate, and pyruvate metabolism. Finally, we comprehensively analyzed the metabolome and transcriptome data and found that many DEGs and DAMs played crucial roles in lipid metabolism and muscle growth and development. In summary, crossbreeding can improve XG goose production performance and affect breast muscle gene expression and metabolites in both female and male geese.

SPSB1-mediated inhibition of TGF-β receptor-II impairs myogenesis in inflammation.

Sepsis-induced intensive care unit-acquired weakness (ICUAW) features profound muscle atrophy and attenuated muscle regeneration related to malfunctioning satellite cells. Transforming growth factor beta (TGF-β) is involved in both processes. We uncovered an increased expression of the TGF-β receptor II (TβRII)-inhibitor SPRY domain-containing and SOCS-box protein 1 (SPSB1) in skeletal muscle of septic mice. We hypothesized that SPSB1-mediated inhibition of TβRII signalling impairs myogenic differentiation in response to inflammation. We performed gene expression analyses in skeletal muscle of cecal ligation and puncture- (CLP) and sham-operated mice, as well as vastus lateralis of critically ill and control patients. Pro-inflammatory cytokines and specific pathway inhibitors were used to quantitate Spsb1 expression in myocytes. Retroviral expression plasmids were used to investigate the effects of SPSB1 on TGF-β/TβRII signalling and myogenesis in primary and immortalized myoblasts and differentiated myotubes. For mechanistical analyses we used coimmunoprecipitation, ubiquitination, protein half-life, and protein synthesis assays. Differentiation and fusion indices were determined by immunocytochemistry, and differentiation factors were quantified by qRT-PCR and Western blot analyses. SPSB1 expression was increased in skeletal muscle of ICUAW patients and septic mice. Tumour necrosis factor (TNF), interleukin-1β (IL-1β), and IL-6 increased the Spsb1 expression in C2C12 myotubes. TNF- and IL-1β-induced Spsb1 expression was mediated by NF-κB, whereas IL-6 increased the Spsb1 expression via the glycoprotein 130/JAK2/STAT3 pathway. All cytokines reduced myogenic differentiation. SPSB1 avidly interacted with TβRII, resulting in TβRII ubiquitination and destabilization. SPSB1 impaired TβRII-Akt-Myogenin signalling and diminished protein synthesis in myocytes. Overexpression of SPSB1 decreased the expression of early (Myog, Mymk, Mymx) and late (Myh1, 3, 7) differentiation-markers. As a result, myoblast fusion and myogenic differentiation were impaired. These effects were mediated by the SPRY- and SOCS-box domains of SPSB1. Co-expression of SPSB1 with Akt or Myogenin reversed the inhibitory effects of SPSB1 on protein synthesis and myogenic differentiation. Downregulation of Spsb1 by AAV9-mediated shRNA attenuated muscle weight loss and atrophy gene expression in skeletal muscle of septic mice. Inflammatory cytokines via their respective signalling pathways cause an increase in SPSB1 expression in myocytes and attenuate myogenic differentiation. SPSB1-mediated inhibition of TβRII-Akt-Myogenin signalling and protein synthesis contributes to a disturbed myocyte homeostasis and myogenic differentiation that occurs during inflammation.

Remodeling of Neuromuscular Junctions in Target Muscle Following Nerve Regeneration in Mice After Delayed Peripheral Nerve Repair

Peripheral nerve injury (PNI) induces severe functional loss in extremities. Progressive denervation and atrophy occur in the muscles if the nerve repair is delayed for long periods of the time. To overcome these difficulties, detailed mechanisms should be determined for neuromuscular junction (NMJ) degeneration in target muscles after PNI and regeneration after nerve repair. We established two models of end-to-end neurorrhaphy and allogeneic nerve grafting in the chronic phase after common peroneal nerve injury in female mice (n = 100 in total). We evaluated motor function, histology, and gene expression in the target muscles during their regeneration processes and compared the models. We found that the functional recovery with allogeneic nerve grafting was superior to that with end-to-end neurorrhaphy, and the number of reinnervated NMJs and Schwann cells was increased at 12 weeks after allograft. In addition, NMJ- and Schwann cell-related molecules showed high expression in the target muscle in the allograft model. These results suggest that Schwann cell migrating from the allograft might play a crucial role in nerve regeneration in the chronic phase after PNI. The relationship between the NMJ and Schwann cells should be further investigated in the target muscle.

Effect of muscle fibre types and carnosine levels on the expression of carnosine-related genes in pig skeletal muscle

It is generally accepted that carnosine (β-alanyl-l-histidine) content is higher in glycolytic than in oxidative muscle fibres, but the underlying mechanisms responsible for this difference remain to be elucidated. A first study to better understand potential mechanisms involved was undertaken (1) to determine whether differences in the expression of carnosine-related enzymes (CARNS1, CNDP2) and transporters (SLC6A6, SLC15A3, SLC15A4, SLC36A1) exist between oxidative and glycolytic myofibres and (2) to study the effect of carnosine on myoblast proliferative growth and on carnosine-related gene expression in cultured myoblasts isolated from glycolytic and oxidative muscles. Immunohistochemistry analyses were conducted to determine the cellular localization of carnosine-related proteins. Laser-capture microdissection and qPCR analyses were performed to measure the expression of carnosine-related genes in different myofibres isolated from the longissimus dorsi muscle of ten crossbred pigs. Myogenic cells originating from glycolytic and oxidative muscles were cultured to assess the effect of carnosine (0, 10, 25 and 50 mM) on their proliferative growth and on carnosine-related gene expression. The mRNA abundance of CNDP2 and of the studied carnosine transporters was higher in oxidative than in glycolytic myofibres. Since carnosine synthase (CARNS1) mRNA abundance was not affected by either the fibre type or the addition of carnosine to myoblasts, its transcriptional regulation would not be the main process by which carnosine content differences are determined in oxidative and glycolytic muscles. The addition of carnosine to myoblasts leading to a dose-dependent increase in SLC15A3 transcripts, however, suggests a role for this transporter in carnosine uptake and/or efflux to maintain cellular homeostasis.

The eQTL colocalization and transcriptome-wide association study identify potentially causal genes responsible for economic traits in Simmental beef cattle

BackgroundA detailed understanding of genetic variants that affect beef merit helps maximize the efficiency of breeding for improved production merit in beef cattle. To prioritize the putative variants and genes, we ran a comprehensive genome-wide association studies (GWAS) analysis for 21 agronomic traits using imputed whole-genome variants in Simmental beef cattle. Then, we applied expression quantitative trait loci (eQTL) mapping between the genotype variants and transcriptome of three tissues (longissimus dorsi muscle, backfat, and liver) in 120 cattle.ResultsWe identified 1,580 association signals for 21 beef agronomic traits using GWAS. We then illuminated 854,498 cis-eQTLs for 6,017 genes and 46,970 trans-eQTLs for 1,903 genes in three tissues and built a synergistic network by integrating transcriptomics with agronomic traits. These cis-eQTLs were preferentially close to the transcription start site and enriched in functional regulatory regions. We observed an average of 43.5% improvement in cis-eQTL discovery using multi-tissue eQTL mapping. Fine-mapping analysis revealed that 111, 192, and 194 variants were most likely to be causative to regulate gene expression in backfat, liver, and muscle, respectively. The transcriptome-wide association studies identified 722 genes significantly associated with 11 agronomic traits. Via the colocalization and Mendelian randomization analyses, we found that eQTLs of several genes were associated with the GWAS signals of agronomic traits in three tissues, which included genes, such as NADSYN1, NDUFS3, LTF and KIFC2 in liver, GRAMD1C, TMTC2 and ZNF613 in backfat, as well as TIGAR, NDUFS3 and L3HYPDH in muscle that could serve as the candidate genes for economic traits.ConclusionsThe extensive atlas of GWAS, eQTL, fine-mapping, and transcriptome-wide association studies aid in the suggestion of potentially functional variants and genes in cattle agronomic traits and will be an invaluable source for genomics and breeding in beef cattle.

Crush Injury and Simulated Flight Effects on Muscle Gene Expression in Female Mice.

Aeromedical evacuation provides critical care during long-distance transport of injured victims between medical facilities. Often, these victims sustain muscle trauma related to mechanical insults, such as crush. Understanding the effects of flight on injured muscle is important because the aircraft cabin represents an external environment with mild hypoxia-the cabin's altitude is 2,438 m instead of sea level. Because mild hypobaric hypoxia can alter gene expression in normal muscle and affect recovery patterns, it is beneficial to examine whether this type of hypoxia may also alter injury-related genes. The objective of this study was to verify the hypothesis that differential gene expression occurs in response to mild hypobaric hypoxia exposure in crush-injured muscle during two early recovery (preregeneration stage) time points. Twenty-four female mice were anesthetized, and the right gastrocnemius muscle underwent crush injury. Approximately 24 hours later, mice were exposed to normobaric normoxia or hypobaric hypoxia for 8-9 hours. After 32 or 48 hours of recovery, the mice were euthanized, and the right and left lateral gastrocnemius muscles were collected for microarray and bioinformatics analyses. The study hypothesis was verified. There were 353 highly upregulated, differentially expressed genes identified in the injured muscle compared to the uninjured muscle. Mid1 was upregulated in both pressure conditions regardless of injury status. There were 52 and 15 differentially expressed genes at 32 and 48 hours postinjury, respectively, in the hypobaric hypoxia-exposed, injured muscle compared to the normobaric normoxia-exposed, injured muscle. The macrophage gene Cd68 correlated with other leukocyte-related genes. These findings expand our understanding of the genetic changes that occur in muscle in response to a crush injury, including those related to the macrophage protein CD68. Nursing interventions addressing adequate functioning after crush muscle injury may need to consider the effects on Cd68 and its closely related genes. In addition, our results suggest a responsiveness of the gene Mid1 to flight-relevant hypobaric hypoxia. Changes in the expression of Mid1 may be appropriate in assessing the long-term health of flight crew members.

Estrogen-related Receptor Signaling in Skeletal Muscle Fitness.

Skeletal muscle is a highly plastic tissue that can alter its metabolic and contractile features, as well as regenerative potential in response to exercise and other conditions. Multiple signaling factors including metabolites, kinases, receptors, and transcriptional factors have been studied in the regulation of skeletal muscle plasticity. Recently, estrogen-related receptors (ERRs) have emerged as a critical transcriptional hub in control of skeletal muscle homeostasis. ERRα and ERRγ - the two highly expressed ERR sub-types in the muscle respond to various extracellular cues such as exercise, hypoxia, fasting and dietary factors, in turn regulating gene expression in the skeletal muscle. On the other hand, conditions such as diabetes and muscular dystrophy suppress expression of ERRs in the skeletal muscle, likely contributing to disease progression. We highlight key functions of ERRs in the skeletal muscle including the regulation of fiber type, mitochondrial metabolism, vascularization, and regeneration. We also describe how ERRs are regulated in the skeletal muscle, and their interaction with important muscle regulators (e. g. AMPK and PGCs). Finally, we identify critical gaps in our understanding of ERR signaling in the skeletal muscle, and suggest future areas of investigation to advance ERRs as potential targets for function promoting therapeutics in muscle diseases.

Nutritional status affects Igf1 regulation of skeletal muscle myogenesis, myostatin, and myofibrillar protein degradation pathways in gopher rockfish (Sebastes carnatus)

Insulin-like growth factor-1 (Igf1) regulates skeletal muscle growth in fishes by increasing protein synthesis and promoting muscle hypertrophy. When fish experience periods of insufficient food intake, they undergo slower muscle growth or even muscle wasting, and those changes emerge in part from nutritional modulation of Igf1 signaling. Here, we examined how food deprivation (fasting) affects Igf1 regulation of liver and skeletal muscle gene expression in gopher rockfish (Sebastes carnatus), a nearshore rockfish of importance for commercial and recreational fisheries in the northeastern Pacific Ocean, to understand how food limitation impacts Igf regulation of muscle growth pathways. Rockfish were either fed or fasted for 14 d, after which a subset of fish from each group was treated with recombinant Igf1 from sea bream (Sparus aurata). Fish that were fasted lost body mass and had lower body condition, reduced hepatosomatic index, and lower plasma Igf1 concentrations, as well as a decreased abundance of igf1 gene transcripts in the liver, increased hepatic mRNAs for Igf binding proteins igfbp1a, igfbp1b, and igfbp3a, and decreased mRNA abundances for igfbp2b and a putative Igf acid labile subunit (igfals) gene. In skeletal muscle, fasted fish showed a reduced abundance of intramuscular igf1 mRNAs but elevated gene transcripts encoding Igf1 receptors A (igf1ra) and B (igf1rb), which also showed downregulation by Igf1. Fasting increased skeletal muscle mRNAs for myogenin and myostatin1, as well as ubiquitin ligase F-box only protein 32 (fbxo32) and muscle RING-finger protein-1 (murf1) genes involved in muscle atrophy, while concurrently downregulating mRNAs for myoblast determination protein 2 (myod2), myostatin2, and myogenic factors 5 (myf5) and 6 (myf6 encoding Mrf4). Treatment with Igf1 downregulated muscle myostatin1 and fbxo32 under both feeding conditions, but showed feeding-dependent effects on murf1, myf5, and myf6/Mrf4 gene expression indicating that Igf1 effects on muscle growth and atrophy pathways is contingent on recent food consumption experience.