Muscle unloading potentiates the effects of acetyl-L-carnitine on the slow oxidative muscle phenotype

The effect of acetyl-L-carnitine (ALCAR) supplementation to 3-month-old rats in normal-loading and unloading conditions has been here investigated by a combined morphological, biochemical and transcriptional approach to test whether ALCAR might cause a remodeling of the metabolic/contractile phenotype of soleus muscle. Morphological assessment demonstrated an increase of type I oxidative fiber content and cross-sectional area in ALCAR-treated animals both in normal-loading and in unloading conditions. ALCAR prevented loss of mitochondrial mass in unloaded animals whereas no ALCAR-dependent increase of mitochondrial mass occurred in normal-loaded muscle. Validated microarray analysis delineated an ALCAR-induced maintenance of a slow-oxidative expression program only in unloaded soleus muscle. Indeed, the muscle adjustment of the expression profile of factors underlying mitochondrial oxidative metabolism, protein turnover, fiber type differentiation and an adaptation of voltage-gated ion channel expression was distinguishable with respect to the loading status. This selectivity may suggest a key role of muscle loading status in the manifestation of ALCAR effects. The results extend to a broader level of biological informations the previous notion on ALCAR positive effect in rat soleus muscle during unloading and point to a role of ALCAR for the maintenance of its slow-oxidative fiber character.

Myogenin modulates exercise endurance by altering skeletal muscle metabolism

The function of myogenic regulatory factors (MRFs) during adult life is not well understood. The requirement of one of these MRFs, myogenin (Myog), during embryonic muscle development suggests an equally important role in adult muscle. In this study, we have determined the function of myogenin during adult life using a conditional allele of Myog. In contrast to embryonic development, myogenin is not required for adult viability, and Myog-deleted mice exhibited no remarkable phenotypic changes during sedentary life. Remarkably, sedentary Myog-deleted mice demonstrated enhanced exercise endurance during involuntary treadmill running. Altered blood glucose and lactate levels in sedentary Myog-deleted mice after exhaustion suggest an enhanced glycolytic metabolism and an ability to excessively deplete muscle and liver glycogen stores. Traditional changes associated with enhanced exercise endurance, such as fiber type switching, and increased oxidative potential, were not detected in sedentary Myog-deleted mice. After long-term voluntary exercise, trained Myog-deleted mice demonstrated an enhanced adaptive response to exercise. Trained Myog-deleted mice exhibited superior exercise endurance associated with an increased proportion of slow-twitch fibers and increased oxidative capacity. In a parallel experiment, dystrophin-deficient young adult mice showed attenuated muscle fatigue following the deletion of Myog. These results demonstrate a novel and unexpected role for myogenin in modulating skeletal muscle metabolism.

Protective Effect of Focal Adhesion Kinase against Skeletal Muscle Reperfusion Injury after Acute Limb Ischemia.

OBJECTIVES In cardiac muscle, ischemia reperfusion (IR) injury is attenuated by mitochondrial function, which may be upregulated by focal adhesion kinase (FAK). The aim of this study was to determine whether increased FAK levels reduced rhabdomyolysis in skeletal muscle too. MATERIAL AND METHODS In a translational in vivo experiment, rat lower limbs were subjected to 4 hours of ischemia followed by 24 or 72 hours of reperfusion. FAK expression was stimulated 7 days before (via somatic transfection with pCMV-driven FAK expression plasmid) and outcomes were measured against non-transfected and empty transfected controls. Slow oxidative (i.e., mitochondria-rich) and fast glycolytic (i.e., mitochondria-poor) type muscles were analyzed separately regarding rhabdomyolysis, apoptosis, and inflammation. Severity of IR injury was assessed using paired non-ischemic controls. RESULTS After 24 hours of reperfusion, marked rhabdomyolysis was found in non-transfected and empty plasmid-transfected fast-type glycolytic muscle, tibialis anterior. Prior transfection enhanced FAK concentration significantly (p = 0.01). Concomitantly, levels of BAX, promoting mitochondrial transition pores, were reduced sixfold (p = 0.02) together with a blunted inflammation (p = 0.01) and reduced rhabdomyolysis (p = 0.003). Slow oxidative muscle, m. soleus, reacted differently: although apoptosis was detectable after IR, rhabdomyolysis did not appear before 72 hours of reperfusion; and FAK levels were not enhanced in ischemic muscle despite transfection (p = 0.66). CONCLUSIONS IR-induced skeletal muscle rhabdomyolysis is a fiber type-specific phenomenon that appears to be modulated by mitochondria reserves. Stimulation of FAK may exploit these reserves constituting a potential therapeutic approach to reduce tissue loss following acute limb IR in fast-type muscle.

X-ray microtomography analysis of the damage micromechanisms in 3D woven composites under low-velocity impact

3D woven composites reinforced with either S2 glass, carbon or a hybrid combination of both and containing either polyethylene or carbon z-yarns were tested under low-velocity impact. Different impact energies (in the range of 21–316 J) were used and the mechanical response (in terms of the impact strength and energy dissipated) was compared with that measured in high-performance, albeit standard, 2D laminates. It was found that the impact strength in both 2D and 3D materials was mainly dependent on the in-plane fiber fracture. Conversely, the energy absorption capability was primarily influenced by the presence of z-yarns, having the 3D composites dissipated over twice the energy than the 2D laminates, irrespective of their individual characteristics (fiber type, compaction degree, porosity, etc.). X-ray microtomography revealed that this improvement was due to the z-yarns, which delayed delamination and maintained the structural integrity of the laminate, promoting energy dissipation by tow splitting, intensive fiber breakage under the tup and formation of a plug by out-of-plane shear.

Mutations in the nebulin gene associated with autosomal recessive nemaline myopathy

The congenital nemaline myopathies are rare hereditary muscle disorders characterized by the presence in the muscle fibers of nemaline bodies consisting of proteins derived from the Z disc and thin filament. In a single large Australian family with an autosomal dominant form of nemaline myopathy, the disease is caused by a mutation in the α-tropomyosin gene TPM3. The typical form of nemaline myopathy is inherited as an autosomal recessive trait, the locus of which we previously assigned to chromosome 2q21.2-q22. We show here that mutations in the nebulin gene located within this region are associated with the disease. The nebulin protein is a giant protein found in the thin filaments of striated muscle. A variety of nebulin isoforms are thought to contribute to the molecular diversity of Z discs. We have studied the 3′ end of the 20.8-kb cDNA encoding the Z disc part of the 800-kDa protein and describe six disease-associated mutations in patients from five families of different ethnic origins. In two families with consanguineous parents, the patients were homozygous for point mutations. In one family with nonconsanguineous parents, the affected siblings were compound heterozygotes for two different mutations, and in two further families with one detected mutation each, haplotypes are compatible with compound heterozygosity. Immunofluorescence studies with antibodies specific to the C-terminal region of nebulin indicate that the mutations may cause protein truncation possibly associated with loss of fiber-type diversity, which may be relevant to disease pathogenesis.

Viral mediated expression of insulin-like growth factor I blocks the aging-related loss of skeletal muscle function

During the aging process, mammals lose up to a third of their skeletal muscle mass and strength. Although the mechanisms underlying this loss are not entirely understood, we attempted to moderate the loss by increasing the regenerative capacity of muscle. This involved the injection of a recombinant adeno-associated virus directing overexpression of insulin-like growth factor I (IGF-I) in differentiated muscle fibers. We demonstrate that the IGF-I expression promotes an average increase of 15% in muscle mass and a 14% increase in strength in young adult mice, and remarkably, prevents aging-related muscle changes in old adult mice, resulting in a 27% increase in strength as compared with uninjected old muscles. Muscle mass and fiber type distributions were maintained at levels similar to those in young adults. We propose that these effects are primarily due to stimulation of muscle regeneration via the activation of satellite cells by IGF-I. This supports the hypothesis that the primary cause of aging-related impairment of muscle function is a cumulative failure to repair damage sustained during muscle utilization. Our results suggest that gene transfer of IGF-I into muscle could form the basis of a human gene therapy for preventing the loss of muscle function associated with aging and may be of benefit in diseases where the rate of damage to skeletal muscle is accelerated.

Delineation of a slow-twitch-myofiber-specific transcriptional element by using in vivo somatic gene transfer.

Contractile proteins are encoded by multigene families, most of whose members are differentially expressed in fast- versus slow-twitch myofibers. This fiber-type-specific gene regulation occurs by unknown mechanisms and does not occur within cultured myocytes. We have developed a transient, whole-animal assay using somatic gene transfer to study this phenomenon and have identified a fiber-type-specific regulatory element within the promoter region of a slow myofiber-specific gene. A plasmid-borne luciferase reporter gene fused to various muscle-specific contractile gene promoters was differentially expressed when injected into slow- versus fast-twitch rat muscle: the luciferase gene was preferentially expressed in slow muscle when fused to a slow troponin I promoter, and conversely, was preferentially expressed in fast muscle when fused to a fast troponin C promoter. In contrast, the luciferase gene was equally well expressed by both muscle types when fused to a nonfiber-type-specific skeletal actin promoter. Deletion analysis of the troponin I promoter region revealed that a 157-bp enhancer conferred slow-muscle-preferential activity upon a minimal thymidine kinase promoter. Transgenic analysis confirmed the role of this enhancer in restricting gene expression to slow-twitch myofibers. Hence, somatic gene transfer may be used to rapidly define elements that direct myofiber-type-specific gene expression prior to the generation of transgenic mice.

Locomotor performance of closely related Tropidurus species: relationships with physiological parameters and ecological divergence

Tropidurid lizards have colonized a variety of Brazilian open environments without remarkable morphological variation, despite ecological and structural differences among habitats used. This study focuses on two Tropidurus sister-species that, despite systematic proximity and similar morphology, exhibit great ecological divergence and a third ecologically generalist congeneric species providing an outgroup comparison. We quantified jumping capacity and sprint speed of each species on sand and rock to test whether ecological divergence was also accompanied by differences in locomotor performance. Relevant physiological traits possibly associated with locomotor performance metabolic scopes and fiber type composition, power output and activity of the enzymes citrate synthase, pyruvate kinase and lactate dehydrogenase of the iliofibularis muscle - were also compared among the three Tropidurus species. We found that the two sister-species exhibited remarkable differences in jumping performance, while Tropidurus oreadicus, the more distantly related species, exhibited intermediate values. Tropidurus psamonastes, a species endemic to sand dunes, exhibited high absolute sprint speeds on sand, jumped rarely and possessed a high proportion of glycolytic fibers and low activity of citrate synthase. The sister-species Tropidurus itambere, endemic to rocky outcrops, performed a large number of jumps and achieved lower absolute sprint speed than T. psamonastes. This study provides evidence of rapid divergence of locomotor parameters between sister-species that use different substrates, which is only partially explained by variation in physiological parameters of the iliofibularis muscle.

The MODE-GAP project

The MODE-GAP project was established to identify the optimum fiber type for use beyond the capacity crunch. This paper will review some of the key drivers behind the project and analyze the key limits and potential improvements © 2013 IEEE.

Numerical prediction of the lateral displacement capacity of flexural reinforced concrete shear walls

Previous earthquakes showed that shear wall damage could lead to catastrophic failures of the reinforced concrete building. The lateral load capacity of shear walls needs to be estimated to minimize associated losses during catastrophic events; hence it is necessary to develop and validate reliable and stable numerical methods able to converge to reasonable estimations with minimum computational effort. The beam-column 1-D line element with fiber-type cross-section model is a practical option that yields results in agreement with experimental data. However, shortcomings of using this model to predict the local damage response may come from the fact that the model requires fine calibration of material properties to overcome regularization and size effects. To reduce the mesh-dependency of the numerical model, a regularization method based on the concept of post-yield energy is applied in this work to both the concrete and the steel material constitutive laws to predict the nonlinear cyclic response and failure mechanism of concrete shear walls. Different categories of wall specimens known to produce a different response under in plane cyclic loading for their varied geometric and detailing characteristics are considered in this study, namely: 1) scaled wall specimens designed according to the European seismic design code and 2) unique full-scale wall specimens detailed according to the U.S. design code to develop a ductile behavior under cyclic loading. To test the boundaries of application of the proposed method, two full-scale walls with a mixed shear-flexure response and different values of applied axial load are also considered. The results of this study show that the use of regularized constitutive models considerably enhances the response predictions capabilities of the model with regards to global force-drift response and failure mode. The simulations presented in this thesis demonstrate the proposed model to be a valuable tool for researchers and engineers.

Effect of cement type and water storage time on the push-out bond strength of a glass fiber post

This study investigated the effects of the cement type and the water storage time on the push-out bond strength of a glass fiber post. Glass fiber posts (Fibrekor, Jeneric Pentron) were luted to post spaces using a self-cured resin cement (C&B Cement [CB]), a glass ionomer cement (Ketac Cem [KC]) or a resin-modified glass ionomer cement (GC FujiCEM [FC]) according to the manufacturers’ instructions. For each luting agent, the specimens were exposed to one of the following water storage times (n=5): 1 day (T1), 7 days (T7), 90 days (T90) and 180 days (T180). Push-out tests were performed after the storage times. Control specimens were not exposed to water storage, but subjected to the push-out test 10 min after post cementation. Data (in MPa) were analyzed by Kruskal-Wallis and Dunn`s test (α=0.05). Cement type and water storage time had a significant effect (p<0.05) on the push-out bond strength. CB showed significantly higher values of retention (p<0.05) than KC and FC, irrespective of the water storage time. Water storage increased significantly the push-out bond strength in T7 and T90, regardless of the cement type (p<0.05). The results showed that fiber posts luted to post spaces with the self-cured resin cement exhibited the best bonding performance throughout the 180-day water storage period. All cements exhibited a tendency to increase the bond strength after 7 and 90 days of water storage, decreasing thereafter.

Impairment of mossy fiber long-term potentiation and associative learning in pituitary adenylate cyclase activating polypeptide type I receptor-deficient Mice

The pituitary adenylate cyclase activating polypeptide (PACAP) type I receptor (PAC1) is a G-protein-coupled receptor binding the strongly conserved neuropeptide PACAP with 1000-fold higher affinity than the related peptide vasoactive intestinal peptide. PAC1-mediated signaling has been implicated in neuronal differentiation and synaptic plasticity. To gain further insight into the biological significance of PAC1-mediated signaling in vivo, we generated two different mutant mouse strains, harboring either a complete or a forebrain-specific inactivation of PAC1. Mutants from both strains show a deficit in contextual fear conditioning, a hippocampus-dependent associative learning paradigm. In sharp contrast, amygdala-dependent cued fear conditioning remains intact. Interestingly, no deficits in other hippocampus-dependent tasks modeling declarative learning such as the Morris water maze or the social transmission of food preference are observed. At the cellular level, the deficit in hippocampus-dependent associative learning is accompanied by an impairment of mossy fiber long-term potentiation (LTP). Because the hippocampal expression of PAC1 is restricted to mossy fiber terminals, we conclude that presynaptic PAC1-mediated signaling at the mossy fiber synapse is involved in both LTP and hippocampus-dependent associative learning.

Effect of Cement Type, Relining Procedure, and Length of Cementation on Pull-out Bond Strength of Fiber Posts

Introduction: As opposed to the cementation metal posts, the cementation of fiber posts has several details that can significantly influence the success of post retention. This study evaluated the effect of the relining procedure, the cement type, and the luted length of the post on fiber posts retention. Methods: One hundred eighty bovine incisors were selected to assess post retention; after endodontic treatment, the canals were flared with diamonds burs. Post holes were prepared in lengths of 5, 7.5, and 10 mm; the fiber posts were relined with composite resin and luted with RelyX ARC, RelyX Unicem, or RelyX Luting 2. All cements are manufactured by 3M ESPE (St. Paul, MN). Samples were subjected to a pull-out bond strength test in a universal testing machine; the results (N) were submitted to a three-way analysis of variance and the Tukey post hoc test (alpha = 0.05). Results: The improvement of post retention occurred with the increase of the post length luted into the root canal; the relining procedure improved the pull-out bond strength. RelyX Unicem and RelyX ARC showed similar values of retention, both showing higher values than RelyX Luting 2. Conclusion: Post length, the reining procedure, and the cement type are all important factors for improving the retention of fiber posts. (J Endod 2010;36:1543-1546)

Hollow-fiber liquid-phase microextraction of amphetamine-type stimulants in human hair samples

A fast method was optimized and validated in order to quantify amphetamine-type stimulants (amphetamine, AMP; methamphetamine, MAMP; fenproporex, FPX; 3,4-methylenedioxymethamphetamine, MDMA; and 3,4-methylenedioxyamphetamine, MDA) in human hair samples. The method was based in an initial procedure of decontamination of hair samples (50 mg) with dichloromethane, followed by alkaline hydrolysis and extraction of the amphetamines using hollow-fiber liquid-phase micro extraction (HF-LPME) in the three-phase mode. Gas chromatography-mass spectrometry (GC-MS) was used for identification and quantification of the analytes. The LoQs obtained for all amphetamines (around 0.05 ng/mg) were below the cut-off value (0.2 ng/mg) established by the Society of Hair Testing (SoHT). The method showed to be simple and precise. The intra-day and inter-day precisions were within 10.6% and 11.4%, respectively, with the use of only two deuteratecl internal standards (AMP-d5 and MDMA-d5). By using the weighted least squares linear regression (1/x(2)), the accuracy of the method was satisfied in the lower concentration levels (accuracy values better than 87%). Hair samples collected from six volunteers who reported regular use of amphetamines were submitted to the developed method. Drug detection was observed in all samples of the volunteers. (c) 2012 Elsevier B.V. All rights reserved.