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Context Heating of meat leads to structural changes reflected in the juiciness and the tenderness of the cooked meat. Aims This study aimed to characterise the effect of prolonged ageing and cooking on pork-quality traits. Methods Longissimus lumborum samples from 12 carcasses were aged 3 days (conventional ageing) or 15 days (prolonged ageing) and pork cuboids were cooked at 50–80°C for 30 min. Cooking loss, total water content (TWC), Warner–Bratzler shear force (WBSF) and shrinkage (longitudinal, transverse and estimated volume) of the pork loin cuboids were measured. Key results Prolonged ageing for 15 days reduced the WBSF of samples cooked at 50°C, and the cooking loss for samples cooked at 70°C and 80°C, relative to conventional ageing for 3 days. The WBSF of pork aged for 15 days was not different from that of pork aged for 3 days. Prolonged ageing reduced longitudinal shrinkage of cuboids, but TWC and transverse/volume shrinkage of cuboids were not affected by ageing. The diameter of cooked muscle fibre fragments was smaller in pork subjected to prolonged ageing. Conclusions Prolonged ageing was favourable for minimising cooking loss at higher cooking temperatures but was only favourable for tenderness at the lowest cooking temperature. Low pH of the samples is likely to have caused the lack of tenderisation with ageing.

Automated peptide and oligonucleotide synthesizers enabled a revolution in molecular biology and helped pave the way to modern synthetic biology. Similarly, fully automated synthetic chemistry coul...

Meat shrinks and assumes an irregular shape during heating due to the varying distribution of connective tissue and extracellular spaces. The terrestrial 3D laser scanning technology is proposed as an alternative method to manual measurements to estimate the volume of irregularly shaped meat cuboids and to predict the cooking loss based on the heating induced volume shrinkage. Cuboids from aged pork loins (longissimus lumborum, n = 12) were heated at 50, 60, 70 or 80 °C for 30 min. Two methods of 3D reconstruction and volume estimation of the pork cuboids by laser scanning were used; without a base scan (laser-B) and with inclusion of a base scan (laser+B), as well as two methods based on manual caliper measurements of all twelve edges (caliper-12) or of three edges in each direction (caliper-3). Both laser scanning methods (Laser+B and Laser-B) resulted in greater volume estimates for the raw samples than the caliper-12 and caliper-3 measurements (38.3, 39.4 compared to 33.9, 34.8 cm3, respectively). Cooking loss across the different temperatures could be best predicted by the caliper-based perimeter shrinkage (r = 0.94, P 0.05) with the cooking loss. 3D laser scanning technologies can be considered by the food industry for 3D reconstruction and volume estimation, however improvements are needed in the data processing to allow for better predictability of meat quality attributes.

Duchenne muscular dystrophy involves an absence of dystrophin, a cytoskeletal protein which supports cell structural integrity and scaffolding for signalling molecules in myocytes. Affected individuals experience progressive muscle degeneration that leads to irreversible loss of ambulation and respiratory diaphragm function. Although clinical management has greatly advanced, heart failure due to myocardial cell loss and fibrosis remains the major cause of death. We examined cardiac morphology and function in D2.B10-Dmdmdx/J (D2-mdx) mice, a relatively new mouse model of muscular dystrophy, which we compared to their wild-type background DBA/2J mice (DBA/2). We also tested whether drug treatment with a specific blocker of mitochondrial permeability transition pore opening (Debio-025), or ACE inhibition (Perindopril), had any effect on dystrophy-related cardiomyopathy. D2-mdx mice were treated for six weeks with Vehicle control, Debio-025 (20 mg/kg/day), Perindopril (2 mg/kg/day), or a combination ( n = 8 /group). At 18 weeks, compared to DBA/2, D2-mdx hearts displayed greater ventricular collagen, lower cell density, greater cell diameter, and greater protein expression levels of IL-6, TLR4, BAX/Bcl2, caspase-3, PGC-1α, and notably monoamine oxidases A and B. Remarkably, these adaptations in D2-mdx mice were associated with preserved resting left ventricular function similar to DBA/2 mice. Compared to vehicle, although Perindopril partly attenuated the increase in heart weight and collagen at 18 weeks, the drug treatments had no marked impact on dystrophic cardiomyopathy.

Ultraviolet-visible (UV-Vis) absorption spectra are routinely collected as part of high-performance liquid chromatography (HPLC) analysis systems and can be used to identify chemical reaction products by comparison to the reference spectra. Here, we present UV-adVISor as a new computational tool for predicting the UV-Vis spectra from a molecule's structure alone. UV-Vis prediction was approached as a sequence-to-sequence problem. We utilized Long-Short Term Memory and attention-based neural networks with Extended Connectivity Fingerprint Diameter 6 or molecule SMILES to generate predictive models for the UV spectra. We have produced two spectrum datasets (dataset I, N = 949, and dataset II, N = 2222) using different compound collections and spectrum acquisition methods to train, validate, and test our models. We evaluated the prediction accuracy of the complete spectra by the correspondence of wavelengths of absorbance maxima and with a series of statistical measures (the best test set median model parameters are in parentheses for model II), including RMSE (0.064), R2 (0.71), and dynamic time warping (DTW, 0.194) of the entire spectrum curve. Scrambling molecule structures with the experimental spectra during training resulted in a degraded R2, confirming the utility of the approaches for prediction. UV-adVISor is able to provide fast and accurate predictions for libraries of compounds.

Ultraviolet-visible (UV-Vis) absorption spectra are routinely collected as part of high-performance liquid chromatography (HPLC) analysis systems and can be used to identify chemical reaction products by comparison to reference spectra. Here, we present UV-adVISor as a new computational tool for predicting UV-Vis spectra from a molecule’s structure alone. UV-Vis prediction was approached as a sequence-to-sequence problem. We utilized Long-Short Term Memory and attention-based neural networks with Extended Connectivity Fingerprint diameter 6 or molecule SMILES to generate predictive models for UV-spectra. We have produced two spectrum datasets (Dataset I, N = 949 and Dataset II, N = 2222) using different compound collections and spectrum acquisition methods to train, validate, and test our models. We evaluated the prediction accuracy of the complete spectra by the correspondence of wavelengths of absorbance maxima and with a series of statistical measures (the best test set median model parameters are in parentheses for Model II), including RMSE (0.064), R2 (0.71), and dynamic time warping (DTW, 0.194) of the entire spectrum curve. Scrambling molecule structures with experimental spectra during training resulted in a degraded R2, confirming the utility of the approaches for prediction. UV-adVISor is able to provide fast and accurate predictions for libraries of compounds.

We carried out laboratory experiments to study the decomposition of carbon fibers at high temperature and studied the material used in charring carbon phenolic ablators for planetary probe heatshields. Porous plug samples were exposed to controlled rates of molecular oxygen, carbon dioxide, and carbon monoxide at furnace-heated flow-tube reactor temperatures between 700 and 1500 K and pressures between 2000 and 6200 Pa. We used calibrated mass spectroscopy to make time-resolved quantitative measurements of decomposition products of the gases downstream of the test sample. Absolute and differential pressure measurements were used to determine the high-temperature permeability of the material and to monitor changes during material decomposition, and we characterized the microstructure of the porous sample using scanning electron microscopy. We found substantial carbon fiber oxidation for O2 flows, resulting in increasing CO/CO2 production ratios with increasing temperature. The CO2 was shown to react with carbon fibers following a Boudouard reaction at temperatures above 1200 K.

The effect of thermal protein denaturation on the structure and quality of muscles of different fibre types is not well understood. Unaged masseter (100% type I fibres) and cutaneous trunci (93% type II fibres) muscles (N = 10) were assessed for their characteristics, protein denaturation, cooking loss, Warner- Bratzler shear force (WBSF) and shrinkage after heating at 50 °C - 85 °C with a rate of 5 °C/ min. Raw masseter had a higher pH, collagen and water content, shorter sarcomere, comparable fibre diameter, and shorter and wider fragments upon homogenization, than cutaneous trunci. In cutaneous trunci, at 55 °C - 60 °C, the lower transition temperature of myosin and the greater cumulative enthalpy resulted in greater cooking loss in muscle cuboids, and greater transverse, longitudinal and volume shrinkage in fibres and fibre fragments, than in masseter. Protein denaturation explained 71% variability in fibre fragment volume and 58% in cooking loss of both muscles, as well as 47% variability in WBSF of masseter.

Duchenne muscular dystrophy (DMD) is a progressive fatal neuromuscular disorder with no cure. Therapies to restore dystrophin deficiency have been approved in some jurisdictions but long-term effectiveness is yet to be established. There is a need to develop alternative strategies to treat DMD. Resveratrol is a nutraceutical with anti-inflammatory properties. Previous studies have shown high doses (100–400 mg/kg bodyweight/day) benefit mdx mice. We treated 4-week-old mdx and wildtype mice with a lower dose of resveratrol (5 mg/kg bodyweight/day) for 15 weeks. Voluntary exercise was used to test if a lower dosage than previously tested could reduce exercise-induced damage where a greater inflammatory infiltrate is present. We found resveratrol promoted skeletal muscle hypertrophy in wildtype mice. In dystrophic muscle, resveratrol reduced exercise-induced muscle necrosis. Gene expression of immune cell markers, CD86 and CD163 were reduced, however, signalling targets associated with resveratrol’s mechanism of action including Sirt1 and NF-κB were unchanged. In conclusion, a lower dose of resveratrol compared to the dosage used by other studies reduced necrosis and gene expression of inflammatory cell markers in dystrophic muscle suggesting it as a therapeutic candidate for treating DMD.

As myosin heavy chain (MyHC) profile of muscle fibres is heavily influenced by neural input, changes in MyHC expression are expected in horses clinically affected with recurrent laryngeal neuropathy (RLN) yet, this has not been thoroughly investigated.To describe the changes in MyHC and fibre diameter in left cricoarytenoideus dorsalis (L-CAD) muscle of horses with clinical signs of RLN.Observational cohort study.Immunohistochemistry was used to assess the MyHC-based fibre-type proportion, size and grouping in the L-CAD of 10 Thoroughbred horses, five clinically affected with RLN and five unaffected controls based on resting endoscopic examination. The Mann-Whitney U test was used to compare the two groups.Compared to controls (of mean age 3.0 ± 1.7 years) which only expressed type I, IIA and IIX MyHC, the L-CAD of affected horses (of mean age 2.8 ± 0.8 years) had obvious fibre-type grouping, and despite apparent compensatory hypertrophy of a small number of fibres, a decrease in overall fibre diameter (median difference -35.2 µm, 95% CI -47.4 to -7.9, P = .02) and diameter of type IIA fibres (median difference -46.8 µm, 95% CI -52.1 to -5.0, P = .03) was observed. Anti-fast MyHC (MY32) cross-immunoreacted with embryonic-MyHC. Whereas MY32-positive fibres were identified as type IIX in controls, in affected horses these fibres were less than 50 µm diameter with internal nuclei and were MYH3-positive for embryonic myosin indicating depletion of type IIX fibres, yet active regeneration and fibre renewal.Small sample size that did not include subclinical cases. Fibre size and appearance rather than staining colour were relied upon to differentiate embryonic from type IIX MyHC.Horses clinically affected with RLN have overall atrophy of fibres, loss of IIX fibres and expression of embryonic myosin indicating regenerative capacity. Despite hypertrophy of some remaining fibres, the overall decline in the bulk of fibres, including those most fatigue-resistant, may be the critical change that results in failure to maintain arytenoid abduction during exercise although direct comparison to subclinical cases is needed to confirm this.

International audience; The changes in secondary structure of proteins with heating were characterised and compared for bovine masseter (fibre type I) and cutaneous trunci (fibre type II) muscles by Differential Scanning Calorimetry (DSC) and Fourier Transform InfraRed (FTIR) microspectroscopy. Heating led to a decrease in alpha helices, and an increase in aggregated strands, random coils and aromatic side chains in the muscle fibres of both muscles. In the intramuscular connective tissue (IMCT) of both muscles, a decrease in alpha helix, turn and unordered structures was complemented with an increase in aggregated strands. At temperatures < 60 degrees C, the greater thermal denaturation of proteins in cutaneous trunci than in masseter (FTIR), supported by a myosin associated peak at 55.8 degrees C for cutaneous trunci and no peak for masseter (DSC), indicates that myosin in type II fibres is more sensitive to thermal denaturation than myosin in type I fibres and this should be considered in thermal meat processing.

This study examines the effects of ageing (1, 14 days), cathepsin inhibition (No or Yes) and temperature (25-90 °C) on the shrinkage of fibre fragments from three bovine muscles (semitendinosus, biceps femoris and psoas major) during heating. Shrinkage was quantified using light microscopy images. Muscle fibres (except in psoas major) had greater transverse shrinkage, and less longitudinal shrinkage in aged than in unaged muscles at temperatures ≥60-75 °C. In addition, cathepsin inhibition during heating at ≥65-90 °C caused greater transverse shrinkage in semitendinosus fibres, and reduced longitudinal shrinkage for all muscles. At temperatures ≥75 °C, the longitudinal and transverse shrinkage of the fibres was correlated for all muscles. Ageing of biceps femoris increases volume shrinkage on a fibre level, and hence potentially cooking loss, while cathepsin activity in the semitendinosus reduces volume shrinkage. In conclusion, cathepsin activity and ageing influence the shrinkage that occurs during heating and these factors should be explored further to enable optimisation of thermal meat processing.

Arising from the ablation of the cytoskeletal protein dystrophin, Duchenne Muscular Dystrophy (DMD) is a debilitating and fatal skeletal muscle wasting disease underpinned by metabolic insufficiency. The inability to facilitate adequate energy production may impede calcium (Ca2+) buffering within, and the regenerative capacity of, dystrophic muscle. Therefore, increasing the metabogenic potential could represent an effective treatment avenue. The aim of our study was to determine the efficacy of adenylosuccinic acid (ASA), a purine nucleotide cycle metabolite, to stimulate metabolism and buffer skeletal muscle damage in the mdx mouse model of DMD. Dystrophin-positive control (C57BL/10) and dystrophin-deficient mdx mice were treated with ASA (3000 µg.mL−1) in drinking water. Following the 8-week treatment period, metabolism, mitochondrial density, viability and superoxide (O2−) production, as well as skeletal muscle histopathology, were assessed. ASA treatment significantly improved the histopathological features of murine DMD by reducing damage area, the number of centronucleated fibres, lipid accumulation, connective tissue infiltration and Ca2+ content of mdx tibialis anterior. These effects were independent of upregulated utrophin expression in the tibialis anterior. ASA treatment also increased mitochondrial viability in mdx flexor digitorum brevis fibres and concomitantly reduced O2− production, an effect that was also observed in cultured immortalised human DMD myoblasts. Our data indicates that ASA has a protective effect on mdx skeletal muscles.

Duchenne muscular dystrophy (DMD) is a lethal muscle wasting disorder caused by mutations in the DMD gene that lead to the absence or severe reduction of dystrophin protein in muscle. The mdx mouse, also dystrophin deficient, is the model most widely used to study the pathology and test potential therapies, but the phenotype is milder than human DMD. This limits the magnitude and range of histological damage parameters and molecular changes that can be measured in pre-clinical drug testing. We used three weeks of voluntary wheel running to exacerbate the mdx phenotype. In mdx mice voluntary exercise increased the amount of damaged necrotic tissue and macrophage infiltration. Global gene expression profiling revealed that exercise induced additional and larger gene expression changes in mdx mice and the pathways most impacted by exercise were all related to immune function or cell-extracellular matrix (ECM) interactions. When we compared the matrisome and inflammation genes that were dysregulated in mdx with those commonly differentially expressed in DMD, we found the exercised mdx molecular signature more closely resembled that of DMD. These gene expression changes in the exercised mdx model thus provide more scope to assess the effects of pre-clinical treatments. Our gene profiling comparisons also highlighted upregulation of extracellular matrix proteins involved in innate immunity pathways, proteases that can release them, and downstream receptors and signalling molecules in exercised mdx and DMD, suggesting that the ECM could be a major source of pro-inflammatory molecules that trigger and maintain the immune response in dystrophic muscle.

A series of experiments was performed to obtain permeability data on FiberForm®, a commercial carbon preform used for manufacturing thermal protection systems. A porous sample was placed in a quartz flow-tube heated by an isothermal furnace. The setup was instrumented to measure mass flow through and pressure drop across the sample. The intrinsic permeability and the Klinkenberg correction, which accounts for rarefied effects, were computed from the experimental data. The role of the gas temperature and pressure on the effective permeability is shown, and it is demonstrated that with proper data reduction, the intrinsic permeability is strictly a function of the micro-structure of the material. A function for the effective permeability of FiberForm, dependent on temperature, pressure, pore geometry, and type of gas is proposed. The intrinsic permeability was evaluated at K 0 = 5.57 × 10 - 11 m2, with a Klinkenberg parameter of 8 c / d p = 2.51 × 10 5 m−1 and a reference porosity of ϕ † = 0.87 .

SummaryReason for performing the study Investigation of toxicosis caused by Malva parviflora was required after 4 horses from the same farm developed severe muscle fasciculations, tachycardia, sweating and periods of recumbency leading to death or euthanasia after ingesting the plant. Objectives To describe historical, clinical, clinicopathological and pathological findings of 4 horses with suspected M. parviflora toxicosis. The role of cyclopropene fatty acids (found in M. parviflora) and mechanism for toxicosis are proposed. Study design Case series. Methods Historical, physical examination, clinicopathological and pathological findings are reported. Due to similarities with atypical myopathy or seasonal pasture myopathy acyl carnitine profiles were performed on sera from 2 cases and equine controls. Presence of cyclopropene fatty acids was also examined in sera of 2 cases. Results M. parviflora had been heavily grazed by the horses with little other feed available. Horse 1 deteriorated rapidly and was subjected to euthanasia. Horse 2 was referred to hospital where severe myocardial disease and generalised myopathy was determined; this horse was subjected to euthanasia 36 h after admission. Horse 3 died rapidly and Horse 4 was subjected to euthanasia at onset of clinical signs. Post-mortem examinations performed on 3 horses revealed acute, multifocal cardiac and skeletal myonecrosis. Myocyte glycogen accumulation was absent when examined in Horse 2. Acyl carnitine profiles revealed increased C14–C18 acyl carnitine concentrations in cases relative to controls. Cyclopropene fatty acids were detected in sera of cases but not controls. Conclusion These findings suggest aetiology different to that of atypical myopathy or seasonal pasture myopathy. We hypothesise that cyclopropene fatty acids in M. parviflora interfere with fatty acid β-oxidation in horses in negative energy balance, causing the clinical signs and abnormal acyl carnitine profiles. These equine cases suggest a pathophysiological course that closely mimics the human genetic condition very long chain acyl CoA dehydrogenase deficiency.

Recent studies have called into question the idea that facial masculinity is a condition-dependent male ornament that indicates immunocompetence in humans. We add to this growing body of research by calculating an objective measure of facial masculinity/femininity using 3D images in a large sample (n = 1,233) of people of European ancestry. We show that facial masculinity is positively correlated with adult height in both males and females. However, facial masculinity scales with growth similarly in males and females, suggesting that facial masculinity is not exclusively a male ornament, as male ornaments are typically more sensitive to growth in males compared with females. Additionally, we measured immunocompetence via heterozygosity at the major histocompatibility complex (MHC), a widely-used genetic marker of immunity. We show that, while height is positively correlated with MHC heterozygosity, facial masculinity is not. Thus, facial masculinity does not reflect immunocompetence measured by MHC heterozygosity in humans. Overall, we find no support for the idea that facial masculinity is a condition-dependent male ornament that has evolved to indicate immunocompetence. ispartof: PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA vol:116 issue:5 pages:1633-1638 ispartof: location:United States status: published

Mouse models have shown that a disintegrin A metalloprotease 12 (ADAM12) is implicated during adipogenesis; the molecular pathways are not well understood. Stealth RNA interference was used to knock down ADAM12 in 3T3-L1 cells. Using gene profiling and metabolic enzymatic markers, we have identified signaling pathways ADAM12 impacts upon during proliferation, differentiation, and maturation of adipocytes. ADAM12 reduced cell numbers in proliferating preadipocytes, delayed differentiation of preadipocytes to adipocytes, and increased lipid accumulation in mature adipocytes. The pathway most affected by ADAM12 knockdown was regulation of insulin-like growth factor (IGF) activity by insulin-like growth factor binding proteins (IGFBPs); ADAM12 is known to cleave IGFBP3 and IGFBP5. The IGF/mTOR signaling pathway was down-regulated, supporting a role for ADAM12 in the IGFBP/IGF/mTOR-growth pathway. PPARγ signaling was also down-regulated by ADAM12 knockdown. Gene ontology (GO) analysis revealed that the extracellular matrix was the cellular compartment most impacted. Filtering for matrisome genes, connective tissue growth factor ( Ctgf) was up-regulated. CTGF and IGBP3 can interact with PPARγ to hinder its regulation. Increased expression of these molecules could have influenced PPARγ signaling reducing differentiation and an imbalance of lipids. We believe ADAM12 regulates cell proliferation of preadipocytes through IGFBP/IGF/mTOR signaling and delays differentiation through altered PPAR signaling to cause an imbalance of lipids within mature adipocytes.

Facial masculinity is thought to be a condition-dependent male ornament, reflecting immunocompetence in humans. To test this hypothesis, we calculated an objective measure of facial masculinity/femininity using three-dimensional images in a large sample (N = 1,233) of people of European ancestry. We show that facial masculinity is positively correlated with adult height in both males and females. This suggests that variation in growth contributes, at least in part, to variation in facial masculinity, which is characteristic of condition-dependent traits. However, facial masculinity scales with growth similarly in males and females, suggesting that facial masculinity is not specifically a male ornament. Additionally, we measured immunocompetence via heterozygosity at the major histocompatibility complex (MHC), a well known genetic marker of immunity. We show that while height is positively correlated with MHC heterozygosity, facial masculinity is not. Thus, facial masculinity does not reflect immunocompetence measured by MHC heterozygosity in humans as thought previously. Overall, we find no support for the idea that facial masculinity is a condition-dependent male ornament that has evolved to indicate immunocompetence.

Duchenne Muscular Dystrophy (DMD) is a progressive and fatal neuromuscular disease which arises from mutations in the dystrophin gene (DMD) that result in the absence or severe reduction of the cytoskeletal protein dystrophin. In addition to the primary dystrophin defect, secondary processes such as inflammation, calcium influx, dysregulated autophagy and fibrosis exacerbate dystrophic pathology and thus increase disease progression. While therapies to restore dystrophin deficiency are being developed, strategies which target these secondary processes could be of benefit to patients. Benfotiamine is a lipid soluble precursor to thiamine that can reduce secondary processes such as inflammation and oxidative stress in diabetic patients. As such we tested it in the mdx mouse model of DMD and found that benfotiamine reduced multiple markers of dystrophic pathology and improved grip strength. In addition, members of the utrophin and dystrophin glycoprotein complexes were significantly increased at the sarcolemma which could improve cell adhesion. We also demonstrated that benfotiamine treatment lowered the expression of macrophage markers and pro-inflammatory cytokines suggesting that benfotiamine is reducing dystrophic pathology by acting on inflammatory processes.

Experimental results demonstrate O2(aΔ1g) (υ=0) formation by catalytic O-atom surface recombination in a room-temperature fused-quartz flow-tube reactor. Resonance-enhanced multiphoton ionization is used to detect O2(aΔ1g) (υ=0) downstream of a nitrogen discharge flow titrated with nitric oxide to introduce oxygen atoms. A calibration procedure based on ozone photodissociation is developed to quantify O2(aΔ1g) (υ=0) resonance-enhanced multiphoton ionization signals. Partial pressures of O2(aΔ1g) (υ=0) in the range of 2.9 to 14 μtorr are measured in the ionization cell for O-atom partial pressures of 1.4–2.9 mtorr atomic oxygen introduced at the titration port. O2(aΔ1g) (υ=1) could not be detected; an upper limit for the O2(aΔ1g) (υ=1) partial pressure is one-fifth of the O2(aΔ1g) (υ=0) partial pressure. A simple chemical kinetics model demonstrates that measured O2(aΔ1g) pressures cannot be explained by gas-phase chemistry alone and must involve O atoms participating in surface reactions. It is found tha...

Grb10 is an adaptor-type signaling protein most highly expressed in tissues involved in insulin action and glucose metabolism, such as muscle, pancreas, and adipose. Germline deletion of Grb10 in mice creates a phenotype with larger muscles and improved glucose homeostasis. However, it has not been determined whether Grb10 ablation specifically in muscle is sufficient to induce hypermuscularity or affect whole body glucose metabolism. In this study we generated muscle-specific Grb10-deficient mice (Grb10-mKO) by crossing Grb10flox/flox mice with mice expressing Cre recombinase under control of the human α-skeletal actin promoter. One-year-old Grb10-mKO mice had enlarged muscles, with greater cross-sectional area of fibers compared with wild-type (WT) mice. This degree of hypermuscularity did not affect whole body glucose homeostasis under basal conditions. However, hyperinsulinemic/euglycemic clamp studies revealed that Grb10-mKO mice had greater glucose uptake into muscles compared with WT mice. Insulin signaling was increased at the level of phospho-Akt in muscle of Grb10-mKO mice compared with WT mice, consistent with a role of Grb10 as a modulator of proximal insulin receptor signaling. We conclude that ablation of Grb10 in muscle is sufficient to affect muscle size and metabolism, supporting an important role for this protein in growth and metabolic pathways.

A disintegrin and metalloproteinase-12 (ADAM12) is involved in the regulation of myogenesis and adipogenesis and is of interest as a potential target to manipulate skeletal muscle development and intramuscular fat (IMF) deposition in cattle to increase beef yield and improve meat quality. The longissimus thoracis muscle (LM) and semitendinosus muscle (STM) from 5 Bos taurus (Angus) and 5 Bos indicus (Brahman) cattle were collected for histological and ADAM12 gene and protein expression analysis. Myofiber typing was used to determine if ADAM12 expression patterns related to differences in muscling and IMF deposition, which are influenced by proportions of the different myofiber types. The STM was found to contain a higher proportion of glycolytic myofibers than the LM, which contained a greater proportion of oxidative myofibers (myofiber ratio of glycolytic to more oxidative types in LM and STM of 1.1 and 3.5, respectively; P < 0.05). ADAM12 gene expression, fluorescent immunohistochemical staining for ADAM12, and image analysis found ADAM12 to be greater in the LM (P < 0.05). Regression analysis found a strong, positive relationship for the distribution of ADAM12 against the proportion of type I myofibers (P < 0.05, r(2) = 0.86). These findings suggest ADAM12 is upregulated in muscles with more slow-oxidative myofibres, such as the LM, and is linked to type I myofibers in cattle. ADAM12 may be important in the regulation and maintenance slow myofibers in the LM muscle.

Observation of the temporal evolution of the fluorescence following laser excitation of boron monoxide molecules to the B 2 Σ + state is used to extract the radiative lifetime and collisional removal rate constant with N 2 as the colliding partner. The two lowest vibrational levels in the 11 BO B 2 Σ + state are examined. The radiative lifetimes are 101 ± 8 ns for υ ′ = 0 and 108 ± 8 ns for υ ′ = 1. The rate constant for collisional removal of B -state υ ′ = 0 11 BO molecules by N 2 is (1.1 ± 0.1) × 10 −11 cm 3 s −1 .

Microbial contamination of carcass surfaces occurs during slaughter and post-slaughter processing steps, therefore interventions are needed to enhance meat safety and quality. Although many studies have been done at the macro-level, little is known about specific processes that influence bacterial attachment to carcass surfaces, particularly the role of extracellular matrix (ECM) proteins. In the present study, the effect of pH and salt (NaCl, KCl and CaCl2) on attachment of Escherichia coli and Salmonella isolates to dominant ECM proteins: collagen I, fibronectin, collagen IV and laminin were assessed. Also, the effects of three chemical rinses commonly used in abattoirs (2% acetic acid, 2% lactic acid and 10% trisodium phosphate (TSP)) were tested. Within a pH range of 5-9, there was no significant effect on attachment to ECM proteins, whereas the effect of salt type and concentration varied depending on combination of strain and ECM protein. A concentration-dependant effect was observed with NaCl and KCl (0.1-0.85%) on attachment of E. coli M23Sr, but only to collagen I. One-tenth percent CaCl2 produced the highest level of attachment to ECM proteins for E. coli M23Sr and EC614. In contrast, higher concentrations of CaCl2 increased attachment of E. coli EC473 to collagen IV. Rinses containing TSP produced >95% reduction in attachment to all ECM proteins. These observations will assist in the design of targeted interventions to prevent or disrupt contamination of meat surfaces, thus improving meat safety and quality.

Exogenous hyaluronan is known to alter muscle precursor cell proliferation, migration, and differentiation, ultimately inhibiting myogenesis in vitro. The aim of the current study was to investigate the role of endogenous hyaluronan synthesis during myogenesis. In quantitative PCR studies, the genes responsible for synthesizing hyaluronan were found to be differentially regulated during muscle growth, repair, and pathology. Although all Has genes (Has1, Has2, and Has3) were differentially regulated in these models, only Has2 gene expression consistently associated with myogenic differentiation. During myogenic differentiation in vitro, Has2 was the most highly expressed of the synthases and increased after induction of differentiation. To test whether this association between Has2 expression and myogenesis relates to a role for Has2 in myoblast differentiation and fusion, C2C12 myoblasts were depleted of Has2 by siRNA and induced to differentiate. Depletion of Has2 inhibited differentiation and caused a loss of cell-associated hyaluronan and the hyaluronan-dependent pericellular matrix. The inhibition of differentiation caused by loss of hyaluronan was confirmed with the hyaluronan synthesis inhibitor 4-methylumbelliferone. In hyaluronan synthesis-blocked cultures, restoration of the pericellular matrix could be achieved through the addition of exogenous hyaluronan and the proteoglycan versican, but this was not sufficient to restore differentiation to control levels. These data indicate that intrinsic hyaluronan synthesis is necessary for myoblasts to differentiate and form syncytial muscle cells, but the hyaluronan-dependent pericellular matrix is not sufficient to support differentiation alone; additional hyaluronan-dependent cell functions that are yet unknown may be required for myogenic differentiation.

Skeletal muscle regeneration in pathology and following injury requires the coordinated actions of inflammatory cells and myogenic cells to remove damaged tissue and rebuild syncytial muscle cells, respectively. Following contusion injury to muscle, the cytokine leukemia inhibitor factor (LIF) is up-regulated and knockout of Lif negatively impacts on morphometric parameters of muscle regeneration. Although it was speculated that LIF regulates muscle regeneration through direct effects on myogenic cells, the inflammatory effects of LIF have not been examined in regenerating skeletal muscle. Therefore, the expression and function of LIF was examined using the antagonist MH35-BD during specific inflammatory and myogenic stages of notexin-induced muscle regeneration in mice. LIF protein and mRNA were up-regulated in two distinct phases following intramuscular injection of notexin into tibialis anterior muscles. The first phase of LIF up-regulation coincided with the increased expression of pro-inflammatory cytokines; the second phase coincided with myogenic differentiation and formation of new myotubes. Administration of the LIF receptor antagonist MH35-BD during the second phase of LIF up-regulation had no significant effects on transcript expression of genes required for myogenic differentiation or associated with inflammation; there were no significant differences in morphometric parameters of the regenerating muscle. Conversely, when MH35-BD was administered during the acute inflammatory phase, increased gene transcripts for the pro-inflammatory cytokines Tnf (Tumor necrosis factor), Il1b (Interleukin-1β) and Il6 (Interleukin-6) alongside an increase in the number of Ly6G positive neutrophils infiltrating the muscle were observed. This was followed by a reduction in Myog (Myogenin) mRNA, which is required for myogenic differentiation, and the subsequent number of myotubes formed was significantly decreased in MH35-BD-treated groups compared to sham. Thus, antagonism of the LIF receptor during the inflammatory phase of skeletal muscle regeneration appeared to induce an inflammatory response that inhibited subsequent myotube formation. We propose that the predominant role of LIF in skeletal muscle regeneration appears to be in regulating the inflammatory response rather than directly effecting myogenic cells.

Leukemia inhibitory factor (LIF) is an important regulator of skeletal muscle regeneration and has been suggested to be mitogenic for myogenic cells because it has been shown to increase the quantity of myoblast cells grown in culture over extended periods of time. Using the established C2C12 murine myoblast cell line, we observed that LIF treatment did not significantly increase the rate at which myoblasts synthesise DNA under conditions which increased cell quantity by 73% above control, whilst the known mitogen fibroblast growth factor-2 significantly increased DNA synthesis under these conditions. Consequently, we examined the capacity of LIF to prevent apoptotic cell death. LIF treatment significantly reduced staurosporine-induced apoptotic DNA fragmentation by 37% compared to control and also reduced the proteolytic activation of caspase-3 by 40% compared to control. This effect of LIF was completely abolished by addition of the phosphatidylinositol 3-kinase inhibitor wortmannin, indicating that the phosphatidylinositol 3-kinase signalling pathway, previously shown to be linked to LIF-dependent increases in cell number, is necessary in mediating the anti-apoptotic effects of LIF. LIF treatment was also associated with increased levels of Bcl-xL and XIAP transcripts compared to control. Therefore, we suggest that the role of LIF in skeletal muscle regeneration and myogenesis is that of a survival factor rather than a mitogen.

Fibrillins 1, 2 and 3 make up a family of genes that encode large, cysteine-rich extracellular matrix glycoproteins found in connective tissues, lung, blood vessels and other extensible tissues. Fibrillins 1 and 2 have both overlapping as well as separate distributions in human embryonic and adult tissues. Fibrillin-containing microfibrils are known to modulate morphogenetic events by proper targeting of growth factors to the extracellular matrix. Mutation of the fibrillin-2 gene causes a genetic disorder, congenital contractural arachnodactyly (CCA), that results in flexion contractures. Previously, we have shown a distinct fibrillin-2 distribution in the pericellular matrix of interior tenocytes and later demonstrated a unique fibrillin-2 containing structure that runs along the tendon cell arrays in the canine flexor tendon. We hypothesized that loss of these fibrillin-2 containing structures might affect normal tendon development. To test our hypothesis, connective tissues from mice null for fibrillin-2 gene expression were studied. Murine flexor digitorum longus tendons were evaluated for total collagen content, and the intermolecular collagen cross-links hydroxylysyl and lysyl pyridinoline. The results show decreased collagen cross-links in fibrillin-2 null mice, however total collagen content remained the same when compared to wild type. Bone morphology was studied using micro computed tomography (CT). Fibrillin-2 null mice display a focal area of decreased bone length in the extremities as compared to wild type mice. Together, these results demonstrate a role for fibrillin-2 in bone and soft connective tissue morphological and biochemical processes.

This book describes the diverse roles that growth factors and cytokines play in skeletal muscle. The extracellular environment has profound effects on the biology of skeletal muscle. The soluble portion of this environment includes a rich milieu of growth factors and cytokines which have been shown to regulate virtually all facets of the response of skeletal muscle to external stimuli, whether it be exercise induced metabolic shifts, remodeling in response to trauma or loading of the ongoing pathology associated with neuromuscular disease. The chapters included in this work illustrate growth factors that directly affect skeletal muscle cells and those which influence non-muscle cells that contribute to the biology of skeletal muscle as a whole tissue. The current state of the art, with the advent of systems biology, allows for the delineation of signaling networks which are regulated by suites of growth factors. This is in stark contrast to early more traditional studies, which only examined the effects of isolated growth factors on the activity of skeletal muscle precursor cells in tissue culture. The work presented in this volume ranges from reviewing and analyzing the roles of individual growth factors in detail, to the complex interplay of multiple soluble factors in the control of muscle functional, and dysfunctional states. The material covered in this volume will particularly suit readers from a range of research fields spanning general muscle biology and physiology, and those working on diseases and conditions affecting skeletal muscle both directly and indirectly

Cytokines are an incredibly diverse group of secreted proteins with equally diverse functions. The actions of cytokines are mediated by the unique and sometimes overlapping receptors to which the soluble ligands bind. Classified within the interleukin-6 family of cytokines are leukemia inhibitory factor (LIF), oncostatin-M (OSM), cardiotrophin-1 (CT-1) and ciliary neurotrophic factor (CNTF). These cytokines all bind to the leukemia inhibitory factor receptor (LIFR) and gp130, and in some cases an additional receptor subunit, leading to activation of downstream kinases and transcriptional activators. LIFR is expressed on a broad range of cell types and can generate pleiotropic effects. In the context of skeletal muscle physiology, these cytokines have been shown to exert effects on motor neurons, inflammatory and muscle cells. From isolated cells through to whole organisms, manipulations of LIFR signaling cytokines have a wide range of outcomes influencing muscle cell growth, myogenic differentiation, response to exercise, metabolism, neural innervation and recruitment of inflammatory cells to sites of muscle injury. This article will discuss the shared and distinct processes that LIFR cytokines regulate in a variety of experimental models with the common theme of skeletal muscle physiology.

Back and hind limb muscles of sheep paternally heterozygous for the callipyge single nucleotide polymorphism undergo extensive hypertrophy shortly after birth. We have established cell cultures from foetal semitendinosus and longissimus dorsi muscles of normal and callipyge animals. Cultures were assessed for rates of proliferation, cell death, myogenicity and DLK1 expression. Myoblasts from callipyge semitendinosus, but not longissimus dorsi muscles, proliferated faster than myoblasts isolated from normal semitendinosus muscle, and cells isolated from either callipyge muscle were more resistant to serum deprivation-induced apoptosis than equivalent cells isolated from normal individuals. Theseobservations indicate that there are intrinsic differences in the behaviour of isolated myoblasts, which are associated with their muscle and genotype of origin. As myoblasts are the cells responsible for hypertrophy of muscle fibres, the observed differences in cell growth may play a role in the hypertrophy of certain muscles in callipyge animals.

An interdisciplinary environmental assessment team from the Howard University Environmental Justice Partnership (HUEJP) conducted a site visit and assessment of the Defense Depot of Memphis, Tennessee in February of 2000. This depot was built in the late 1940’s for storage of numerous chemicals and munitions. As the years progressed, many Memphis citizens have grown to believe that the activities and chemical stockpile located at this site have negatively affected the health environment of their residents. There is anecdotal evidence and documentation of numerous cancers and other illnesses in those local territories, and specifically, at the Memphis Depot site. Currently, this depot is closed and in remediation by the local government. Particularly, citizens of the Rozelle community have started a campaign to investigate any signs of exposure pathways to noted health risks. The HUEJP was contacted and asked to investigate the community concerns. Obliging to the request, we aimed to sample at three drainage sites and a residential site, talk to local citizens, and gain any additional information that would be helpful in relieving anxiety in the Rozelle community. Soil, sediment, and water samples were collected and analyzed for total organic carbon, inorganic anions, and heavy metals. These data show that for the four sites sampled, the highest concentrations of organic compounds and heavy metals were located either within a residential area or in an area with a direct transport pathway to the community. Atomic absorption analysis revealed detectable amounts of cadmium, lead and chromium metals at all sites with direct transport pathways into the residential community, with chromium concentrations being far in excess of the EPA standard limits.

Human IGF-I was over-expressed in skeletal muscles of C57/BL6xCBA mice under the control of the rat skeletal alpha-actin gene promoter. RT-PCR verified expression of the transgene in skeletal muscle but not in the liver of 1- and 21-day old heterozygote transgenic mice. The concentration of endogenous mouse IGF-I, measured by an immunoassay which does not detect human IGF-I, was not significantly different between transgenic mice and wild-type littermates (9.5 +/- 0.8 and 13.3 +/- 1.9 ng/g in muscle; 158.3 +/- 18.6 and 132.9 +/- 33.1 ng/ml in plasma, respectively). In contrast, quantitation with antibodies to human IGF-I showed an increase in IGF-I of about 100 ng/ml in plasma and 150 ng/g in muscle of transgenic mice at 6 months of age. Transgenic males, compared to their age matched wild-type littermates, had a significantly higher body weight (38.6 +/- 0.53 g vs. 35.8 +/- 0.64 g at 6 months of age; P < 0.001), dry fat-free carcass mass (5.51 +/- 0.085 vs. 5.08 +/- 0.092 g; P < 0.001) and myofibrillar protein mass (1.62 +/- 0.045 vs. 1.49 +/- 0.048 g; P < 0.05), although the fractional content of fat in the carcass was lower (167 +/- 7.0 vs. 197 +/- 7.7 g/kg wet weight) in transgenic animals. There was no evidence of muscle hypertrophy and no change in the proportion of slow type I myofibres in the limb muscles of Rskalpha-actin/hIGF-I transgenic mice at 3 or 6 months of age. Phenotypic changes in Rskalpha-actin/hIGF-I mice are likely to be due to systemic as well as autocrine/paracrine effects of overproduction of IGF-I due to expression of the human IGF-I transgene. The effect of muscle specific over-expression of Rskalpha-actin/hIGF-I transgene was tested on: (i) muscle regeneration in auto-transplanted whole muscle grafts; (ii) myofibre atrophy following sciatic nerve transection; and (iii) sarolemmal damage and myofibre necrosis in dystrophic mdx muscle. No beneficial effect of muscle specific over-expression of Rskalpha-actin/hIGF-I transgene was seen in these three experimental models.

Mechanical force is generated within skeletal muscle cells by contraction of specialized myofibrillar proteins. This paper explores how the contractile force generated at the sarcomeres within an individual muscle fiber is transferred through the connective tissue to move the bones. The initial key point for transfer of the contractile force is the muscle cell membrane (sarcolemma) where force is transferred laterally to the basement membrane (specialized extracellular matrix rich in laminins) to be integrated within the connective tissue (rich in collagens) before transmission to the tendons. Connections between (1) key molecules outside the myofiber in the basement membrane to (2) molecules within the sarcolemma of the myofiber and (3) the internal cytoplasmic structures of the cytoskeleton and sarcomeres are evaluated. Disturbances to many components of this complex interactive system adversely affect skeletal muscle strength and integrity, and can result in severe muscle diseases. The mechanical aspects of these crucial linkages are discussed, with particular reference to defects in laminin-alpha2 and integrin-alpha7. Novel interventions to potentially increase muscle strength and reduce myofiber damage are mentioned, and these are also highly relevant to muscle diseases and aging muscle.

We observe electron emission from low work function Cs covered Au surfaces due to the scattering of vibrationally excited NO molecules with 18 quanta of vibration in the ground electronic state. Additional experiments explore the influence of oxygen exposure on the electron emission. These results indicate a nondissociative mechanism for reactivity of vibrationally excited NO on these surfaces and provide evidence against an Auger-like mechanism. We note a remarkable similarity between trends of the vibrationally produced electron emission as the surface composition changes and reports on surface work function.

Loss of the nerve supply to skeletal muscle results in a relentless loss of muscle mass (atrophy) over time. The ability of insulin-like growth factor-1 to reduce atrophy resulting from denervation was examined after transection of the sciatic nerve in transgenic MLC/mIGF-1 mice that over-express mIGF-1 specifically in differentiated myofibres. The cross sectional area (CSA) of all types of myofibres and specifically type IIB myofibres was measured in tibialis anterior muscles from transgenic and wild-type mice at 28 days after denervation. There was a marked myofibre atrophy (w60%) in the muscles of wild-type mice over this time with increased numbers of myofibres with small CSA. In the muscles of MLC/mIGF-1 mice, over-expression of mIGF-1 reduced the rate of denervation induced myofibre atrophy by w30% and preserved myofibres with larger CSA, compared to wild-type muscles. It is proposed that the protective effect of mIGF-1 on denervated myofibres might be due to reduced protein breakdown. q 2004 Elsevier B.V. All rights reserved.

Necrosis of dystrophic myofibers in Duchenne muscular dystrophy and mdx mice results from defects in the subsarcolemmal protein dystrophin that cause membrane fragility and tears in the sarcolemma, and these lead to the destruction of the myofibers. The present study specifically tests whether overexpression of mIGF-1 in mdx/mIGF-1 transgenic mice reduces myofiber breakdown during the acute onset phase of dystrophy (at 21 days). The extent of muscle damage and Evans blue dye (EBD) staining of myofibers was quantitated histologically for mdx/mIGF-1 and their mdx littermates from 15 to 30 days of age. Overexpression of mIGF-1 strikingly reduced the extent of myofiber damage (histology and EBD staining) by up to 97% in tibialis anterior and quadriceps muscles at 21-22 days after birth. In the mdx diaphragm, the onset of muscle breakdown was earlier (by 15 days after birth) but no significant protective effect of IGF-1 was apparent within the first month of age in mdx/IGF-1 mice. These novel observations show that increased mIGF-1 within mdx myofibers specifically reduces the breakdown of dystrophic muscle during the acute onset of muscle degeneration. This mechanism of action can account for the long-term reduced severity of the dystropathology in mdx mice that overexpress mIGF-1 and provides promising opportunities for therapeutic strategies.

We present a detailed report of experiments where hexapole focusing is combined with stimulated emission and Franck–Condon optical pumping techniques on a molecular beam. We show that this approach allows one to achieve a high degree of control over the molecule's ro-vibronic quantum numbers, its laboratory frame velocity, and its transverse divergence. Specifically, new ways of manipulating beams of vibrationally excited molecules emerge, including: (1) transverse refocusing and concomitant improved efficiency for transport of the vibrationally excited molecules, (2) relative enrichment of the concentration of the vibrationally excited molecules with respect to the unexcited portion of the beam, and (3) orientation of vibrationally excited molecules. Laser induced fluorescence, fluorescence depletion, Franck–Condon pumping, stimulated emission pumping, resonance enhanced multi-photon ionization, and hexapole focusing all provide spectroscopic probes into the detailed characteristics of this experiment.

The potential clinical use of stem cells for cell transplantation therapies to replace defective genes in myopathies is an area of intense investigation. Precursor cells derived from non-muscle tissue with myogenic potential have been identified in many tissues, including bone marrow and dermis, although the status of these putative stem cells requires clarification. The incorporation of circulating bone-marrow derived stem cells into regenerating adult skeletal muscle has been demonstrated in mice but the contribution of donor cells is so minimal that it would appear clinically irrelevant at this stage. The possibility of a true stem cell subpopulation within skeletal muscle that replenishes the satellite cells (conventional muscle precursors on the surface of myofibres) is also very attractive as a superior source of myoblasts for muscle construction. A full understanding of the intrinsic factors (i.e. gene expression within the stem cell) and extrinsic factors (i.e. signals from the external environment) which control the commitment of stem cells to the myogenic lineage, and the conditions which favour stem cell expansion in vivo is required before stem cells can be seriously considered for clinical cell therapy.