Muscle Of Wild-type Mice Research Articles

β2-Integrins contribute to skeletal muscle hypertrophy in mice

We tested the contribution of beta(2)-integrins, which are important for normal function of neutrophils and macrophages, to skeletal muscle hypertrophy after mechanical loading. Using the synergist ablation model of hypertrophy and mice deficient in the common beta-subunit of beta(2)-integrins (CD18(-/-)), we found that overloaded muscles of wild-type mice had greater myofiber size, dry muscle mass, and total protein content compared with CD18(-/-) mice. The hypertrophy in wild-type mice was preceded by elevations in neutrophils, macrophages, satellite cell/myoblast proliferation (5'-bromo-2'-deoxyuridine- and desmin-positive cells), markers of muscle differentiation (MyoD1 and myogenin gene expression and formation and size of regenerating myofibers), signaling for protein synthesis [phosphorylation of Akt and 70-kDa ribosomal protein S6 kinase (p70S6k)], and reduced signaling for protein degradation (decreased gene expression of muscle atrophy F box/atrogin-1). The deficiency in beta(2)-integrins, however, altered the accumulation profile of neutrophils and macrophages, disrupted the temporal profile of satellite cell/myoblast proliferation, reduced the markers of muscle differentiation, and impaired the p70S6k signaling, all of which could serve as mechanisms for the impaired hypertrophy in overloaded CD18(-/-) mice. In conclusion, our findings indicate that beta(2)-integrins contribute to the hypertrophic response to muscle overload by temporally regulating satellite cells/myoblast proliferation and by enhancing muscle differentiation and p70S6k signaling.

Effects of UCP3 genotype, temperature and muscle type on energy turnover of resting mouse skeletal muscle

Uncoupling protein 3 (UCP3) is a mitochondrial transporter protein which, when over-expressed in mice, is associated with increased metabolic rate, increased feeding and low body weight. This phenotype probably reflects the increased levels of UCP3 partially uncoupling mitochondrial respiration from cellular ATP demands. Consistent with that, mitochondria isolated from muscles of mice that over-express UCP3 are less tightly coupled than those from wild-type mice but the degree of uncoupling is not modulated by likely physiological regulatory factors. To determine whether this also applies to intact muscle fibres, we tested the hypothesis that UCP3 constitutively (i.e. in an unregulated fashion) uncouples mitochondria in muscles from mice that over-expressed human UCP3 (OE mice). The rate of heat production of resting muscles was measured in vitro using bundles of fibres from soleus and extensor digitorum longus muscles of OE, wild-type (WT) and UCP3 knock-out mice. At 20 degrees C, the only significant effect of genotype was that the rate of heat production of OE soleus (3.04+/-0.16 mW g(-1)) was greater than for WT soleus (2.31+/-0.05 mW g(-1)). At physiological temperature (35 degrees C), the rate of heat production was independent of genotype and equal to the expected in vivo rate for skeletal muscles of WT mice. We conclude that at 35 degrees C, the transgenic UCP3 was not constitutively active, but at 20 degrees C in slow-twitch muscle, it was partially activated by unknown factors. The physiological factor(s) that activate mitochondrial uncoupling by UCP3 in vivo was either not present or inactive in resting isolated muscles.

Sarcalumenin plays a critical role in age‐dependent reduction in cardiac function and SERCA2a activity

Sarcalumenin (SAR), a Ca2+‐binding protein located in the longitudinal sarcoplasmic reticulum (SR), interacts with cardiac SR Ca2+‐ATPase(SERCA2a) to modulate its activity and expression. We have demonstrated that SAR deficiency resulted in mild cardiac dysfunction in young mice (JBC 2005). Since it is well known that SERCA2a activity is decreased with aging, we examined the effects of aging on cardiac function in SAR knockout mice (SAR‐KO). Cardiac function assessed by in vivo hemodynamic study and the expression levels of SERCA2a protein were progressively decreased in senescent SARKO (n=13) when compared with senescent wild‐type (WT) (n=12) or young SARKO (4month old, n=12). LV dp/dtmax (mmHg/s) was 3897 ± 134, 7091 ± 657, and 6836 ± 298 in senescent SARKO, senescent WT, and young SARKO, respectively. The expression levels of SERCA2a protein (% of young WT) were 28 ± 8, 84 ± 17, and 44 ± 17 in senescent SARKO, senescent WT, and young SARKO, respectively. Interestingly, downregulation of SAR preceded downregulation of SERCA in aging cardiac muscles of wild‐type mice. These findings indicated that SAR plays a critical role in age‐dependent reduction in cardiac function and SR Ca2+ reuptake.

Microarray analysis of mdx mice expressing high levels of utrophin: Therapeutic implications for dystrophin deficiency

Duchenne Muscular Dystrophy (DMD) is a fatal muscle wasting disorder caused by dystrophin deficiency. Previous work suggested that increased expression of the dystrophin-related protein utrophin in the mdx mouse can reduce the dystrophic pathophysiology. Physiological tests showed that the transgenic mouse muscle functioned in a way similar to normal muscle. More recently, it has become possible to analyse disease pathways using microarrays, a sensitive method to evaluate the efficacy of a therapeutic approach. We thus examined the gene expression profile of mdx mouse muscle compared to wild-type mouse muscle and compared the data with that obtained from the transgenic line overexpressing utrophin. The data confirm that the expression of utrophin in the mdx mouse muscle results in a global gene expression profile more similar to that seen for the wild-type mouse. This study confirms that a strategy to up-regulate utrophin is likely to be beneficial in dystrophin deficiency.

Insulin Action in the Double Incretin Receptor Knockout Mouse

The incretins glucagon-like peptide 1 and glucose-dependent insulinotropic polypeptide have been postulated to play a role in regulating insulin action, although the mechanisms behind this relationship remain obscure. We used the hyperinsulinemic-euglycemic clamp to determine sites where insulin action may be modulated in double incretin receptor knockout (DIRKO) mice, which lack endogenous incretin action. DIRKO and wild-type mice were fed regular chow or high-fat diet for 4 months. Clamps were performed on 5-h-fasted, conscious, unrestrained mice using an arterial catheter for sampling. Compared with wild-type mice, chow and high fat-fed DIRKO mice exhibited decreased fat and muscle mass associated with increased energy expenditure and ambulatory activity. Clamp rates of glucose infusion (GIR), endogenous glucose production (endoR(a)), and disappearance (R(d)) were not different in chow-fed wild-type and DIRKO mice, although insulin levels were lower in DIRKO mice. Liver Akt expression was decreased but Akt activation was increased in chow-fed DIRKO compared with wild-type mice. High-fat feeding resulted in fasting hyperinsulinemia and hyperglycemia in wild-type but not in DIRKO mice. GIR, suppression of endoR(a), and stimulation of R(d) were inhibited in high fat-fed wild-type mice but not in DIRKO mice. High-fat feeding resulted in impaired tissue glucose uptake (R(g)) in skeletal muscle of wild-type mice but not of DIRKO mice. Liver and muscle Akt activation was enhanced in high fat-fed DIRKO compared with wild-type mice. In summary, DIRKO mice exhibit enhanced insulin action compared with wild-type mice when fed a regular chow diet and are protected from high-fat diet-induced obesity and insulin resistance.

Absence of keratin 19 in mice causes skeletal myopathy with mitochondrial and sarcolemmal reorganization

Intermediate filaments, composed of desmin and of keratins, play important roles in linking contractile elements to each other and to the sarcolemma in striated muscle. We examined the contractile properties and morphology of fast-twitch skeletal muscle from mice lacking keratin 19. Tibialis anterior muscles of keratin-19-null mice showed a small but significant decrease in mean fiber diameter and in the specific force of tetanic contraction, as well as increased plasma creatine kinase levels. Costameres at the sarcolemma of keratin-19-null muscle, visualized with antibodies against spectrin or dystrophin, were disrupted and the sarcolemma was separated from adjacent myofibrils by a large gap in which mitochondria accumulated. The costameric dystrophin-dystroglycan complex, which co-purified with gamma-actin, keratin 8 and keratin 19 from striated muscles of wild-type mice, co-purified with gamma-actin but not keratin 8 in the mutant. Our results suggest that keratin 19 in fast-twitch skeletal muscle helps organize costameres and links them to the contractile apparatus, and that the absence of keratin 19 disrupts these structures, resulting in loss of contractile force, altered distribution of mitochondria and mild myopathy. This is the first demonstration of a mammalian phenotype associated with a genetic perturbation of keratin 19.

Urokinase-type plasminogen activator and macrophages are required for skeletal muscle hypertrophy in mice

Adult skeletal muscle possesses remarkable potential for growth in response to mechanical loading; however, many of the cellular and molecular mechanisms involved remain undefined. The hypothesis of this study was that the extracellular serine protease, urokinase-type plasminogen activator (uPA), is required for muscle hypertrophy, in part by promoting macrophage accumulation in muscle subjected to increased mechanical loading. Compensatory muscle hypertrophy was induced in mouse plantaris (PLT) muscles by surgical ablation of synergist muscles. Following synergist ablation, PLT muscles in wild-type mice demonstrated edema and infiltration of neutrophils and macrophages but an absence of overt muscle fiber damage. Sham procedures resulted in no edema or accumulation of inflammatory cells. In addition, synergist ablation was associated with a large increase in activity of uPA in the PLT muscle. uPA-null mice demonstrated complete abrogation of compensatory hypertrophy associated with reduced macrophage accumulation, indicating that uPA is required for hypertrophy. Macrophages isolated from wild-type PLT muscle during compensatory hypertrophy expressed uPA and IGF-I, both of which may contribute to hypertrophy. To determine whether macrophages are required for muscle hypertrophy, clodronate liposomes were administered to deplete macrophages in wild-type mice; this resulted in reduced muscle hypertrophy. Decreased macrophage accumulation was associated with reduced cell proliferation but did not alter signaling through the mammalian target of rapamycin pathway. These data indicate that uPA and macrophages are required for muscle hypertrophy following synergist ablation.

Skeletal Muscle Adaptation to Exercise Training

Regular endurance exercise has profound benefits on overall health, including the prevention of obesity, cardiovascular disease, and diabetes. The objective of this study was to determine whether AMP-activated protein kinase (AMPK) mediates commonly observed adaptive responses to exercise training in skeletal muscle. Six weeks of voluntary wheel running induced a significant (P < 0.05) fiber type IIb to IIa/x shift in triceps muscle of wild-type mice. Despite similar wheel running capacities, this training-induced shift was reduced by approximately 40% in transgenic mice expressing a muscle-specific AMPKalpha2 inactive subunit. Sedentary mice carrying an AMPK-activating mutation (gamma1TG) showed a 2.6-fold increase in type IIa/x fibers but no further increase with training. To determine whether AMPK is involved in concomitant metabolic adaptations to training, we measured markers of mitochondria (citrate synthase and succinate dehydrogenase) and glucose uptake capacity (GLUT4 and hexokinase II). Mitochondrial markers increased similarly in wild-type and AMPKalpha2-inactive mice. Sedentary gamma1TG mice showed a approximately 25% increase in citrate synthase activity but no further increase with training. GLUT4 protein expression was not different in either line of transgenic mice compared with wild-type mice and tended to increase with training, although this increase was not statistically significant. Training induced a approximately 65% increase in hexokinase II protein in wild-type mice but not in AMPKalpha2-inactive mice. Hexokinase II was significantly elevated in sedentary gamma1TG mice, without an additional increase with training. AMPK is not necessary for exercise training-induced increases in mitochondrial markers, but it is essential for fiber type IIb to IIa/x transformation and increases in hexokinase II protein.

Estrogen receptor-α mediates estrogen-inducible abnormalities in the developing penis

Previously, we reported an association between estrogen receptor-alpha (ERalpha) upregulation and detrimental effects of neonatal diethylstilbestrol (DES) exposure in the rat penis. The objective of this study was to employ the ERalpha knockout (ERalphaKO) mouse model to test the hypothesis that ERalpha mediates DES effects in the developing penis. ERalphaKO and wild-type C57BL/6 mice received oil or DES at a dose of 0.2 microg/pup per day (0.1 mg/kg) on alternate days from postnatal days 2 to 12. Fertility was tested at 80-240 days of age and tissues were examined at 96-255 days of age. DES caused malformation of the os penis, significant reductions in penile length, diameter, and weight, accumulation of fat cells in the corpora cavernosa penis, and significant reductions in weight of the bulbospongiosus and levator ani muscles in wild-type mice. Conversely, ERalphaKO mice treated with DES developed none of the above abnormalities. While nine out of ten male mice sired pups in the wild-type/control group, none did in the wild-type/DES group. ERalphaKO mice, despite normal penile development, are inherently infertile. Both plasma and intratesticular testosterone levels were unaltered in the DES-treated wild-type or DES-treated ERalphaKO mice when compared with controls, although testosterone concentration was much higher in the ERalphaKO mice. Hence, the resistance of ERalphaKO mice to developing penile abnormalities provides unequivocal evidence of an obligatory role for ERalpha in mediating the harmful effects of neonatal DES exposure in the developing penis.

Role of AMP-activated protein kinase in exercise capacity, whole body glucose homeostasis, and glucose transport in skeletal muscle: –Insight from analysis of a transgenic mouse model–

To examine the role of muscle AMP-activated protein kinase (AMPK) in maximal exercise capacity, whole body glucose homeostasis, and glucose transport in skeletal muscle, we generated muscle-specific transgenic mice carrying cDNAs of inactive AMPK α2 (α2i TG). Fed blood glucose was slightly higher in α2i TG mice compared to wild type littermates, however, the difference was not statistically significant. In α2i TG mice, glucose tolerance was slightly impaired in male, but not in female mice, compared to wild type littermates. Maximal exercise capacity was dramatically reduced in α2i TG mice, suggesting that AMPK α2 has a critical role in skeletal muscle during exercise. We confirmed that known insulin-independent stimuli of glucose transport including mitochondrial respiration inhibition, hyperosmolarity, and muscle contraction increased both AMPK α1 and α2 activities in isolated EDL muscle in wild type mice. While, α2 activation was severely blunted and α1 activation was only slightly reduced in α2i TG mice by these insulin independent stimuli compared to wild type mice. Mitochondrial respiration inhibition-induced glucose transport was fully inhibited in isolated EDL muscles in α2i TG mice. However, contraction- or hyperosmolarity-induced glucose transport was nearly normal. These results suggest that AMPK α2 activation is essential for some, but not all insulin-independent glucose transport.

Oxidative Phenotype Protects Myofibers from Pathological Insults Induced by Chronic Heart Failure in Mice

The fiber specificity of skeletal muscle abnormalities in chronic heart failure (CHF) has not been defined. We show here that transgenic mice (8 weeks old) with cardiac-specific overexpression of calsequestrin developed CHF (50.9% decrease in fractional shortening and 56.4% increase in lung weight, P<0.001), cachexia (37.8% decrease in body weight, P<0.001), and exercise intolerance (69.3% decrease in running distance to exhaustion, P<0.001) without a significant change in muscle fiber-type composition. Slow oxidative soleus muscle maintained muscle mass, whereas fast glycolytic tibialis anterior and plantaris muscles underwent atrophy (11.6 and 13.3%, respectively; P<0.05). In plantaris muscle, glycolytic type IId/x and IIb, but not oxidative type I and IIa, fibers displayed significant decreases in cross-sectional area (20.3%, P<0.05). Fast glycolytic white vastus lateralis muscle showed sarcomere degeneration and decreased cytochrome c oxidase IV (39.5%, P<0.01) and peroxisome proliferator-activated receptor gamma co-activator 1alpha protein expression (30.3%, P<0.01) along with a dramatic induction of the MAFbx/Atrogin-1 mRNA. These findings suggest that exercise intolerance can occur in CHF without fiber type switching in skeletal muscle and that oxidative phenotype renders myofibers resistant to pathological insults induced by CHF.

Fetal muscle-derived cells can repair dystrophic muscles in mdx mice

We have previously reported that CD34 + cells purified from mouse fetal muscles can differentiate into skeletal muscle in vitro and in vivo when injected into muscle tissue of dystrophic mdx mice. In this study, we investigate the ability of such donor cells to restore dystrophin expression, and to improve the functional muscle capacity of the extensor digitorum longus muscle (EDL) of mdx mice. For this purpose green fluorescent-positive fetal GFP +/CD34 + cells or desmin +/ −LacZ/CD34 + cells were transplanted into irradiated or non-irradiated mdx EDL muscle. Donor fetal muscle-derived cells predominantly fused with existing fibers. Indeed more than 50% of the myofibers of the host EDL contained donor nuclei delivering dystrophin along 80–90% of the length of their sarcolemma. The presence of significant amounts of dystrophin (about 60–70% of that found in a control wild-type mouse muscle) was confirmed by Western blot analyses. Dystrophin expression also outcompeted that of utrophin, as revealed by a spatial shift in the distribution of utrophin. At 1 month post-transplant, the recipient muscle appeared to have greater resistance to fatigue than control mdx EDL muscle during repeated maximal contractions.

Overexpression of osteoactivin protects skeletal muscle from severe degeneration caused by long-term denervation in mice

We have previously shown that osteoactivin, a type I membrane glycoprotein expressed in myofibers, upregulated expression of matrix metalloprotease (MMP)-3 and MMP-9 in fibroblasts infiltrated denervated skeletal muscle in mice. To address whether osteoactivin-mediated increase in MMPs in skeletal muscle is useful for regeneration of denervated skeletal muscle, we subjected osteoactivin-transgenic mice to long-term denervation for 70 or 90 days. Long-term denervation caused severe degeneration of myofibers and fibrosis in skeletal muscle of wild-type mice. However, overexpression of osteoactivin protected skeletal muscle from such changes. Infiltration of fibroblast-like cells and collagen deposition were sustained at low levels after long-term denervation in skeletal muscle of osteoactivin-transgenic mice. This cytoprotective effect of osteoactivin was supported by the expression of regeneration/degeneration-associated genes in the gastrocnemius muscle during denervation. Denervation significantly upregulated the expression of anti-fibrotic genes, such as glypican-1 and decorin-1, in the gastrocnemius muscle of osteoactivin-transgenic mice, compared with wild-type mice. In contrast, overexpression of osteoactivin caused a significant reduction in denervation-induced expression of elongation factor 1A-1, an indicator for the persistence of degenerated cells. Our results suggest that an osteoactivin-mediated increase in MMPs in skeletal muscle might be useful for protecting injured muscle from fibrosis, leading to full regeneration after denervation.

Genetic modification of the manganese superoxide dismutase/glutathione peroxidase 1 pathway influences intracellular ROS generation in quiescent, but not contracting, skeletal muscle cells

Increased amounts of reactive oxygen species (ROS) are generated by skeletal muscle during contractile activity, but their intracellular source is unclear. The oxidation of 2′,7′-dichlorodihydrofluorescein (DCFH) was examined as an intracellular probe for reactive oxygen species in skeletal muscle myotubes derived from muscles of wild-type mice and mice that were heterozygous knockout for manganese superoxide dismutase ( Sod2 +/− ), homozygous knockout for glutathione peroxidase 1 ( GPx1 −/− ), or MnSOD transgenic overexpressors ( Sod2-Tg). Myoblasts were stimulated to fuse and loaded with DCFH 5–7 days later. Intracellular DCF epifluorescence was measured and myotubes were electrically stimulated to contract for 15 min. Quiescent myotubes with decreased MnSOD or GPx1 showed a significant increase in the rate of DCFH oxidation whereas those with increased MnSOD did not differ from wild type. Following contractions, myotubes from all groups showed an equivalent increase in DCF fluorescence. Thus the oxidation of DCFH in quiescent skeletal muscle myotubes is influenced by the content of enzymes that regulate mitochondrial superoxide and hydrogen peroxide content. In contrast, the increase in DCFH oxidation following contractions was unaffected by reduced or enhanced MnSOD or absent GPx1, indicating that reactive oxygen species produced by contractions were predominantly generated by nonmitochondrial sources.

Raising the antioxidant levels within mouse muscle fibres does not affect contraction‐induced injury

A protocol of 75 lengthening contractions (LCP) administered to skeletal muscles of mice causes an initial force deficit owing to the mechanical disruption of sarcomeres and a reduction in calcium release from the sarcoplasmic reticulum. During the 3 days following the LCP, a 'sealing off process' and inflammatory response occurs. The reactive oxygen species (ROS) released by invading inflammatory cells produce a secondary force deficit that is more severe than the initial deficit. The timing of the infiltration of inflammatory cells and increase in force deficit relative to the sealing off process is not well documented. We tested the null hypothesis that following a lifetime of overexpression of the genes for the intracellular antioxidants manganese superoxide dismutase, copper zinc superoxide dismutase or catalase in transgenic mice, the force deficits 3 days following the administration of a 75 LCP to in situ extensor digitorum longus muscles are not different from those of wild-type mice. Following the LCP, the force deficits ranged from 39 to 59% for the muscles of transgenic mice that overexpressed the genes for intracellular antioxidants and were not different from the force deficit of 44% observed for muscles of wild-type mice. The results provide evidence that the ROS damage does not occur within the cytosol of the injured fibres. Apparently, the hypercontraction of sarcomeres and accumulation of vesicles seal off and protect the intact portions of damaged fibres, such that the ROS damage and repair occurs in the milieu of the necrotic segments that are continuous with the extracellular matrix.

Leukemia inhibitory factor restores the hypertrophic response to increased loading in the LIF(−/−) mouse

Cytokines and growth factors are thought to contribute to skeletal muscle hypertrophy. Leukemia inhibitory factor (LIF), a cytokine, enhances skeletal muscle regeneration; however the role of LIF in skeletal muscle hypertrophy remains uncertain. We examined the hypertrophic ability of the plantaris and soleus muscles in wild-type mice (WT) and LIF knock-out mice [LIF(−/−)] in response to increased mechanical load. Using the functional overload model to induce increases in mechanical load on the plantaris and soleus muscle, WT mice demonstrated increases in plantaris and soleus mass after 7, 21, and 42 days of loading. However, the LIF(−/−) mice had no significant increases in plantaris muscle mass at any time point, while the soleus muscle exhibited a delayed hypertrophic response. Systemic delivery of LIF to the LIF(−/−) mice returned the hypertrophic response to the same levels as the WT mice after 21 days of functional overload. These data demonstrate for the first time that LIF expression in loaded skeletal muscle is critical for the development of skeletal muscle hypertrophy in the functional overload model.

Targeted disruption of iNOS prevents LPS-induced S-nitrosation of IRβ/IRS-1 and Akt and insulin resistance in muscle of mice

We have previously demonstrated that the insulin resistance associated with inducible nitric oxide synthase (iNOS) induction in two different models of obesity, diet-induced obesity and the ob/ob mice, is mediated by S-nitrosation of proteins involved in insulin signal transduction: insulin receptor beta-subunit (IRbeta), insulin receptor substrate 1(IRS-1), and Akt. S-nitrosation of IRbeta and Akt impairs their kinase activities, and S-nitrosation of IRS-1 reduces its tissue expression. In this study, we observed that LPS-induced insulin resistance in the muscle of wild-type mice, as demonstrated by reduced insulin-induced tyrosine phosphorylation of IRbeta and IRS-1, reduced IRS-1 expression and reduced insulin-induced serine phosphorylation of Akt. This resistance occurred in parallel with enhanced iNOS expression, which was accompanied by S-nitrosation of IRbeta/IRS-1 and Akt. In the muscle of iNOS(-/-) mice, we did not observe enhanced iNOS expression or any S-nitrosation of IRbeta/IRS-1 and Akt after LPS treatment. Moreover, insulin resistance was not present. The preservation of insulin-induced tyrosine phosphorylation of IRbeta and IRS-1, of IRS-1 protein expression, and of insulin-induced serine phosphorylation of Akt observed in LPS-treated iNOS(-/-) mice strongly suggests that the insulin resistance induced by LPS is iNOS mediated, probably through S-nitrosation of proteins of early steps of insulin signaling.

Urokinase‐type plasminogen activator and hepatocyte growth factor in muscle regeneration

Muscle healing has been shown to be severely impaired in urokinase plasminogen activator-deficient (uPA-/-) mice, and accelerated in plasminogen activator inhibitor-1 (PAI1-/-) mice. The purpose of this study was to test the hypothesis that one mechanism by which uPA regulates muscle repair is through activation of hepatocyte growth factor (HGF). Extensor digitorum longus muscles were injured by cardiotoxin injection and harvested at 1, 3, 5 and 10 days post-injury. As expected, muscles from wild-type mice demonstrated increased expression of HGF following injury. Muscles from uPA-/- mice showed decreased levels of active HGF compared to muscle from wild-type mice, and muscles from PAI1-/- mice showed increased levels of active HGF. Muscles from wild-type mice showed uptake of bromodeoxyuridine into proliferating cell nuclei following injury, with a peak at 5 days post-injury. Muscles from uPA-/- mice showed an impaired proliferative response following injury, and muscles from PAI1-/- mice showed an accelerated proliferative response. The number of MyoD positive cells showed a similar trend, with a peak at 5 days post-injury in muscle of wild-type mice, reduced numbers of MyoD positive cells in muscle of uPA-/- mice compared with muscle of wild-type mice and increased numbers in muscle of PAI1-/- mice. These data indicate that uPA activation of HGF may be a key step in the activation and proliferation of satellite cells and the regulation of muscle regeneration. Supported by the Department of Defense Peer Reviewed Medical Research Program.

The differential gene expression profiles of proximal and distal muscle groups are altered in pre-pathological dysferlin-deficient mice

The selective pattern of muscle involvement is a key feature of muscular dystrophies. Dysferlinopathy is a good model for studying this process since it shows variable muscle involvement that can be highly selective even in individual patients. The transcriptomes of proximal and distal muscles from wildtype C57BL/10 and dysferlin deficient C57BL/10.SJL- Dysf mice at a prepathological stage were assessed using the Affymetrix oligonucleotide-microarray system. We detected significant variation in gene expression between proximal and distal muscle in wildtype mice. Dysferlin defiency, even in the absence of pathological changes, altered this proximal distal difference but with little specific overlap with previous microarray analyses of dysferlinopathy. In conclusion, proximal and distal muscle groups show distinct patterns of gene expression and respond differently to dysferlin deficiency. This has implications for the selection of muscles for future microarray analyses, and also offers new routes for investigating the selectivity of muscle involvement in muscular dystrophies.

Inverse relationship between PGC-1α protein expression and triacylglycerol accumulation in rodent skeletal muscle

PGC-1alpha is a key regulator of tissue metabolism, including skeletal muscle. Because it has been shown that PGC-1alpha alters the capacity for lipid metabolism, it is possible that PGC-1alpha expression is regulated by the intramuscular lipid milieu. Therefore, we have examined the relationship between PGC-1alpha protein expression and the intramuscular fatty acid accumulation in hindlimb muscles of animals in which the capacity for fatty acid accumulation in muscle is increased (Zucker obese rat) or reduced [FAT/CD36 null (KO) mice]. Rates of palmitate incorporation into triacylglycerols were determined in perfused red (RG) and white gastrocnemius (WG) muscles of lean and obese Zucker rats and in perfused RG and WG muscles of FAT/CD36 KO and wild-type (WT) mice. In obese Zucker rats, the rate of palmitate incorporation into triacylglycerol depots in RG and WG muscles were 28 and 24% greater than in lean rats (P < 0.05). In FAT/CD36 KO mice, the rates of palmitate incorporation into triacylglycerol depots were lower in RG (-50%) and WG muscle (-24%) compared with the respective muscles in WT mice (P < 0.05). In the obese animals, PGC-1alpha protein content was reduced in both RG (-13%) and WG muscles (-15%) (P < 0.05). In FAT/CD36 KO mice, PGC-1alpha protein content was upregulated in both RG (+32%, P < 0.05) and WG muscles (+50%, P < 0.05). In conclusion, from studies in these two animal models, it appears that PGC-1alpha protein expression is inversely related to components of intramuscular lipid metabolism, because 1) PGC-1alpha protein expression is downregulated when triacylglycerol synthesis rates, an index of intramuscular lipid metabolism, are increased, and 2) PGC-1alpha protein expression is upregulated when triacylglycerol synthesis rates are reduced. Therefore, we speculate that the intramuscular lipid sensing may be involved in regulating the protein expression of PGC-1alpha in skeletal muscle.