Mechanical properties of rat soleus after long-term spinal cord transection.

The effects of a complete spinal cord transection (ST) on the mechanical properties of the rat soleus were assessed 3 and 6 mo post-ST and compared with age-matched controls. Maximal tetanic force was reduced by approximately 44 and approximately 25% at 3 and 6 mo post-ST, respectively. Similarly, maximum twitch force was reduced by approximately 29% in 3-mo and approximately 17% in 6-mo ST rats. ST resulted in faster twitch properties as evidenced by shorter time to peak tension (approximately 45%) and half-relaxation time (approximately 55%) at both time points. Maximum shortening velocity was significantly increased in ST rats whether measured by extrapolation from the force-velocity curve (approximately twofold at both time points) or by slack-test measurements (over twofold at both time points). A significant reduction in fatigue resistance of the soleus was observed at 3 (approximately 25%) and 6 mo (approximately 45%) post-ST. For the majority of the speed-related properties, no significant differences were detected between 3- and 6-mo ST rats. However, the fatigue resistance of the soleus was significantly lower in 6- vs. 3-mo ST rats. These data suggest that, between 3 and 6 mo post-ST, force-related properties tended to recover, speed-related properties plateaued, and fatigue-related properties continued to decline. Thus some specific functional properties of the rat soleus related to contractile force, speed, and fatigue adapted independently after ST.     

Remarkable heterogeneity in myosin heavy-chain composition of the human young masseter compared with young biceps brachii

Adult human jaw muscles differ from limb and trunk muscles in enzyme-histochemical fibre type composition. Recently, we showed that the human masseter and biceps differ in fibre type pattern already at childhood. The present study explored the myosin heavy-chain (MyHC) expression in the young masseter and biceps muscles by means of gel electrophoresis (GE) and immuno-histochemical (IHC) techniques. Plasticity in MyHC expression during life was evaluated by comparing the results with the previously reported data for adult muscles. In young masseter, GE identified MyHC-I, MyHC-IIa MyHC-IIx and small proportions of MyHC-fetal and MyHC-α cardiac. Western blots confirmed the presence of MyHC-I, MyHC-IIa and MyHC-IIx. IHC revealed in the masseter six isomyosins, MyHC-I, MyHC-IIa, MyHC-IIx, MyHC-fetal, MyHC α-cardiac and a previously not reported isoform, termed MyHC-IIx'. The majority of the masseter fibres co-expressed two to four isoforms. In the young biceps, both GE and IHC identified MyHC-I, MyHC-IIa and MyHC-IIx. MyHC-I predominated in both muscles. Young masseter showed more slow and less-fast and fetal MyHC than the adult and elderly masseter. These results provide evidence that the young masseter muscle is unique in MyHC composition, expressing MyHC-α cardiac and MyHC-fetal isoforms as well as hitherto unrecognized potential spliced isoforms of MyHC-fetal and MyHC-IIx. Differences in masseter MyHC expression between young adult and elderly suggest a shift from childhood to adulthood towards more fast contractile properties. Differences between masseter and biceps are proposed to reflect diverse evolutionary and developmental origins and confirm that the masseter and biceps present separate allotypes of muscle.     

Myosin Heavy Chains IIa and IId Are Functionally Distinct in the Mouse

Myosin in adult murine skeletal muscle is composed primarily of three adult fast myosin heavy chain (MyHC) isoforms. These isoforms, MyHC-IIa, -IId, and -IIb, are >93% identical at the amino acid level and are broadly expressed in numerous muscles, and their genes are tightly linked. Mice with a null mutation in the MyHC-IId gene have phenotypes that include growth inhibition, muscle weakness, histological abnormalities, kyphosis (spinal curvature), and aberrant kinetics of muscle contraction and relaxation. Despite the lack of MyHC-IId, IId null mice have normal amounts of myosin in their muscles because of compensation by the MyHC-IIa gene. In each muscle examined from IId null mice, there was an increase in MyHC-IIa– containing fibers. MyHC-IIb content was unaffected in all muscles except the masseter, where its expression was extinguished in the IId null mice. Cross-sectional fiber areas, total muscle cross-sectional area, and total fiber number were affected in ways particular to each muscle. Developmental expression of adult MyHC genes remained unchanged in IId null mice. Despite this universal compensation of MyHC-IIa expression, IId null mice have severe phenotypes. We conclude that despite the similarity in sequence, MyHC-IIa and -IId have unique roles in the development and function of skeletal muscle.     

猪骨骼肌MyHC-Ⅱb基因上调表达的分子机制

哺乳动物骨骼肌由各种不同类型的肌纤维镶嵌而成,不同类型肌球蛋白重链的表达是造成不同类型肌纤维的主要原因.目前已知的肌球蛋白重链家族包含8种亚型,其中长白猪骨骼肌My HC-Ⅱb的表达量显著高于中国地方猪,然而造成这种差异的分子机制未见报道.本研究用荧光定量PCR证明了长白猪背最长肌中My HC-Ⅱb m RNA的表达量显著高于莱芜猪(P=0.013).删除实验结果表明,从转录起始位点上游-1024 bp删除到-187 bp之后,My HC-Ⅱb表达量显著下降,分析发现,在这段启动子区域内存在3个E-box序列;分别突变这3个E-box序列后,My HC-Ⅱb启动子驱动的荧光素酶活性显著下降(P=0.036).另外,在My HC-Ⅱb上游启动子区?1398 bp处发现一个GT的突变,所检测的64头莱芜猪在该位点全部为GG型,65头长白猪中13头为GG型,16头为TT型,36头为GT型.在C2C12细胞系中的转染实验结果显示,G突变为T之后有增加My HC-Ⅱb表达的趋势.Western blot的结果表明,转录因子Myo D在两猪种间表达差异不显著(P=0.136),而Myf-5在长白猪中的表达量极显著高于其在莱芜猪中的表达量(P=0.0036).这些数据表明,Myf-5是造成猪My HC-Ⅱb基因m RNA上调表达的重要因素之一.     

Reduced lipoprotein lipase activity in postural skeletal muscle during aging.

Lipoprotein lipase (LPL) is a key enzyme for fatty acid and lipoprotein metabolism in muscle. However, the effect of aging on LPL regulation in skeletal muscle is unknown. We report the effect of aging on LPL regulation in the soleus (red oxidative postural) muscle and the tibialis anterior (white glycolytic non-weight-bearing) muscle in 4- and 24-mo-old Fischer 344 rats and 18- and 31-mo-old Fischer 344 x Brown-Norway F1 (F-344 x BN F1) rats. Total and heparin-releasable LPL (HR-LPL) activities were decreased 38% (P < 0.01) and 52% (P < 0.05), respectively, in the soleus muscle of the older Fischer 344 rats. There was a 32% reduction (P < 0.05) of total LPL protein mass in the soleus muscle with aging. The results were confirmed in another strain. A decrease of total LPL activity (-50%, P < 0.05) was also found in the soleus muscle between 18- and 31-mo-old F-344 x BN F1 rats. LPL mRNA concentration in the soleus muscle was not different between ages. Total LPL protein mass was reduced by 46% (P < 0.05) in the soleus muscle of the 31-mo-old F-344 x BN F1 rats. In the tibialis anterior muscle, neither LPL activity nor mRNA concentration was affected by age in either strain. In conclusion, LPL regulation in a non-weight-bearing muscle was not affected by aging. However, there was a pronounced reduction in LPL activity and LPL protein mass in postural muscle with aging.     

Hindlimb unweighting decreases ecNOS gene expression and endothelium-dependent dilation in rat soleus feed arteries.

We tested the hypothesis that hindlimb unweighting (HLU) and the associated reduction in soleus muscle blood flow causes decreased expression of endothelial cell nitric oxide synthase (ecNOS) mRNA and protein and attenuated endothelium-dependent vasodilator responses in rat soleus feed arteries (SFA). Male Sprague-Dawley rats were exposed to HLU (n = 12) or cage control (Con; n = 12) conditions for 14 days. At the end of this period, SFA were isolated, removed, and cannulated with two glass micropipettes for examination of vasodilator responses or frozen for analysis of ecNOS mRNA and protein expression. RT-PCR of RNA from single SFA was used to measure ecNOS mRNA, and immunoblots on single SFAs were used to measure ecNOS protein content. Results revealed that both ecNOS mRNA and ecNOS protein expression were lower in SFA from HLU rats. Dilation to increased intraluminal flow was attenuated in SFA from HLU rats (Con: 88 +/- 8% vs. HLU: 53 +/- 8%) as was maximal vasodilation to acetylcholine (10(-9)-10(-4) M; Con: 88 +/- 5% vs. HLU: 73 +/- 5%). Sensitivity to the endothelium-independent vasodilator sodium nitroprusside (10(-10)-10(-4) M) was enhanced by HLU (EC(50): Con: 4.46 x 10(-7) M vs. HLU: 5.00 x 10(-8) M). Collectively, these data indicate that the chronic reduction in soleus blood flow associated with the reduced physical activity during HLU results in reduced expression of ecNOS mRNA and protein in SFA and attenuated endothelium-dependent vasodilation.     

Muscle oxidative capacity during IL-6-dependent cancer cachexia

Many diseases are associated with catabolic conditions that induce skeletal muscle wasting. These various catabolic states may have similar and distinct mechanisms for inducing muscle protein loss. Mechanisms related to muscle wasting may also be related to muscle metabolism since glycolytic muscle fibers have greater wasting susceptibility with several diseases. The purpose of this study was to determine the relationship between muscle oxidative capacity and muscle mass loss in red and white hindlimb muscles during cancer cachexia development in the Apc(Min/+) mouse. Gastrocnemius and soleus muscles were excised from Apc(Min/+) mice at 20 wk of age. The gastrocnemius muscle was partitioned into red and white portions. Body mass (-20%), gastrocnemius muscle mass (-41%), soleus muscle mass (-34%), and epididymal fat pad (-100%) were significantly reduced in severely cachectic mice (n = 8) compared with mildly cachectic mice (n = 6). Circulating IL-6 was fivefold higher in severely cachectic mice. Cachexia significantly reduced the mitochondrial DNA-to-nuclear DNA ratio in both red and white portions of the gastrocnemius. Cytochrome c and cytochrome-c oxidase complex subunit IV (Cox IV) protein were reduced in all three muscles with severe cachexia. Changes in muscle oxidative capacity were not associated with altered myosin heavy chain expression. PGC-1α expression was suppressed by cachexia in the red and white gastrocnemius and soleus muscles. Cachexia reduced Mfn1 and Mfn2 mRNA expression and markers of oxidative stress, while Fis1 mRNA was increased by cachexia in all muscle types. Muscle oxidative capacity, mitochondria dynamics, and markers of oxidative stress are reduced in both oxidative and glycolytic muscle with severe wasting that is associated with increased circulating IL-6 levels.     

Regulation of lipoprotein lipase in muscle and adipose tissue during exercise.

Lipoprotein lipase (LPL) is regulated in a tissue-specific manner; exercise increases LPL activity in muscle at the same time it is reduced in adipose tissue. The purpose of this study was to determine the relationship between LPL activity and LPL mRNA in muscle and adipose tissue in rats exposed to one bout of exercise. Immediately after a 2-h swim, LPL activity [pmol free fatty acids (FFA).min-1.mg tissue-1] in the exercised animals was reduced 43% in adipose tissue (110 +/- 26 to 63 +/- 17) and increased almost twofold in the soleus muscle (203 +/- 26 to 383 +/- 59) compared with sedentary control animals. At the same time, LPL mRNA was reduced 42% in adipose tissue and increased 50 and 100% in the red vastus and white vastus muscles, respectively. Twenty-four hours after the swim, LPL activity had returned to control levels in adipose tissue and the soleus muscle. At hour 24 of recovery, LPL mRNA was still reduced 23% in the adipose tissue of exercised animals but was not significantly different between exercised and control animals in any of the muscle tissues analyzed. Changes in total RNA concentration could not account for the changes in relative LPL mRNA expression. The relationship between LPL enzyme activity and LPL mRNA in muscle and adipose tissue was +0.86 and +0.93 at 0 and 24 h postexercise, respectively. Thus the tissue-specific changes in enzyme activity induced by exercise could be mediated, in part, through pretranslational control.     

Skeletal muscle IGF-binding protein-3 and -5 expressions are age,muscle, and load dependent

The purpose of the current study was to examine IGFBP-3, -4, and -5 mRNA and protein expression levels as a function of muscle type, age, and regrowth from an immobilization-induced atrophy in Fischer 344 x Brown Norway rats. IGFBP-3 mRNA expression in the 4-mo-old animals was significantly higher in the red and white portions of the gastrocnemius muscle compared with the soleus muscle. However, there were no significant differences in IGFBP-3 mRNA expression among any of the muscle groups in the 30-mo-old animals. There were no significant differences in IGFBP-5 mRNA expression in any of the muscle groups, whereas in the 30-mo-old animals there was significantly less IGFBP-5 mRNA expression in the white gastrocnemius compared with the red gastrocnemius muscles. Although IGFBP-3 and -5 proteins were detected in the type I soleus muscle with Western blot analyses, no detection was observed in the type II red and white portions of the gastrocnemius muscle. Aging from adult (18 mo) to old animals (30 mo) was associated with decreases in IGFBP-3 mRNA and protein and IGFBP-5 protein only in the soleus muscle. After 10 days of recovery from 10 days of hindlimb immobilization, IGFBP-3 mRNA and protein increased in soleus muscles from young (4-mo) rats; however, only IGFBP-3 protein increased in the old (30-mo) rats. Whereas there were no changes in IGFBP-5 mRNA expression during recovery, IGFBP-5 protein in the 10-day-recovery soleus muscle did increase in the young, but not in the old, rats. Because one of the functions of IGFBPs is to modulate IGF-I action on muscle size and phenotype, it is hypothesized that IGFBP-3 and -5 proteins may have potential modulatory roles in type I fiber-dominated muscles, aging, and regrowth from atrophy.     

Hindlimb suspension increases insulin binding and glucose metabolism

After 28 days of hindlimb-suspension, insulin binding, 2-deoxy-D-glucose (2-DG) uptake, and glucose metabolism (glycolysis and glycogenesis) were determined at various insulin concentrations (0.2-30 nM) in soleus muscle of young (18-day-old) and adult (150-day-old) rats. Compared with age-matched controls the young (YS) and adult suspended (AS) rats had lower soleus and body weights and insulin levels (P less than 0.05). Per milligram of protein, insulin binding, 2-DG uptake, and the rate of glycolysis were increased by approximately 200%, and the rate of glycogenesis was increased approximately 100% in the YS group (P less than 0.05). Except for a reduction in glycogenesis (P less than 0.05) all other parameters also increased in the AS rats (P less than 0.05). On the basis of the whole muscle the rate of glucose metabolism (glycogenesis + glycolysis) was reduced in the YS rats (P less than 0.05), but in the AS rats glucose metabolism was similar to the controls. Thus the increased glucose metabolism (i.e., per milligram of protein) in the YS and AS groups may represent a compensatory response by atrophied muscle to attempt to sustain glucose removal from the circulation. Because greater insulin binding occurred in YS muscle [35% slow-twitch (ST)] than in the control group (70% ST), and greater insulin binding occurred in the AS (81% ST) than in the control group (90% ST), higher insulin binding capacities are not always related to a high proportion of ST muscle fibers. In conclusion, after hindlimb suspension, marked increments in insulin binding and glucose metabolism occur in the soleus muscle.     

Regulation of skeletal muscle dihydropyridine receptor gene expression by biomechanical unloading.

Biomechanical unloading of the rat soleus by hindlimb unweighting is known to induce atrophy and a slow- to fast-twitch transition of skeletal muscle contractile properties, particularly in slow-twitch muscles such as the soleus. The purpose of this study was to determine whether the expression of the dihydropyridine (DHP) receptor gene is upregulated in unloaded slow-twitch soleus muscles. A rat DHP receptor cDNA was isolated by screening a random-primed cDNA lambda gt10 library from denervated rat skeletal muscle with oligonucleotide probes complementary to the coding region of the rabbit DHP receptor cDNA. Muscle mass and DHP receptor mRNA expression were assessed 1, 4, 7, 14, and 28 days after hindlimb unweighting in rats by tail suspension. Isometric twitch contraction times of soleus muscles were measured at 28 days of unweighting. Northern blot analysis showed that tissue distribution of DHP receptor mRNA was specific for skeletal muscle and expression was 200% greater in control fast-twitch extensor digitorum longus (EDL) than in control soleus muscles. A significant stimulation (80%) in receptor message of the soleus was induced as early as 24 h of unloading without changes in muscle mass. Unloading for 28 days induced marked atrophy (control = 133 +/- 3 vs. unweighted = 62.4 +/- 1.8 mg), and expression of the DHP receptor mRNA in the soleus was indistinguishable from levels normally expressed in EDL muscles. These changes in mRNA expression are in the same direction as the 37% reduction in time to peak tension and 28% decrease in half-relaxation time 28 days after unweighting. Our results suggest that muscle loading necessary for weight support modulates the expression of the DHP receptor gene in the soleus muscle.     

Co-ordinate expression of cytochrome c oxidase subunit III and VIc mRNAs in rat tissues.

Cytochrome c oxidase (EC 1.9.3.1) is an enzyme which is composed of subunits derived from both the mitochondrial and the nuclear genomes. To determine whether or not the expression of these two genomes is co-ordinated at the mRNA level, we have examined the steady-state levels of mRNAs coding for cytochrome c oxidase subunit III (mitochondrially encoded) and subunit VIc (nuclear-encoded) in rat tissues. This was compared with the tissue concentration of the holoenzyme, which was estimated by measuring cytochrome c oxidase enzyme activity. The tissues (heart, brain, liver, kidney, soleus muscle and superficial white vastus muscle) possessed a 13-fold range of enzyme activity, which was highest in heart and lowest in the superficial vastus muscle. Specific subunit mRNA levels were quantified by using slot-blot hybridization of cDNA probes to total tissue RNA. The highest values for subunit III and Vlc mRNA tissue contents were found in kidney, followed by liver and heart (40-60% of that of kidney). The white vastus muscle contained the lowest subunit mRNA level (15% of that of kidney). Although some variability was apparent within each tissue, a parallel pattern of mRNA expression of the nuclear- and mitochondrially encoded subunits was observed. Differences between muscle (heart, vastus and soleus) and non-muscle tissues were noted in the relationship between mRNA and protein levels of expression. Thus, although this suggests that tissue-specific regulatory processes operate, the steady-state expression of subunit III and subunit Vlc mRNAs appears to be co-ordinately regulated.     

Differential regulation of glucose transporter activity and expression in red and white skeletal muscle.

Insulin-stimulated glucose transport activity and GLUT4 glucose transporter protein expression in rat soleus, red-enriched, and white-enriched skeletal muscle were examined in streptozotocin (STZ)-induced insulin-deficient diabetes. Six days of STZ-diabetes resulted in a nearly complete inhibition of insulin-stimulated glucose transport activity in perfused soleus, red, and white muscle which recovered following insulin therapy. A specific decrease in the GLUT4 glucose transporter protein was observed in soleus (3-fold) and red (2-fold) muscle which also recovered to control values with insulin therapy. Similarly, cardiac muscle displayed a marked STZ-induced decrease in GLUT4 protein that was normalized by insulin therapy. White muscle displayed a small but statistically significant decrease in GLUT4 protein (23%), but this could not account for the marked inhibition of insulin-stimulated glucose transport activity observed in this tissue. In addition, GLUT4 mRNA was found to decrease in red muscle (2-fold) with no significant alteration in white muscle. The effect of STZ-induced diabetes was time-dependent with maximal inhibition of insulin-stimulated glucose transport activity at 24 h in both red and white skeletal muscle and half-maximal inhibition at approximately 8 h. In contrast, GLUT4 protein in red and white muscle remained unchanged until 4 and 7 days following STZ treatment, respectively. These data demonstrate that red skeletal muscle displays a more rapid hormonal/metabolic-dependent regulation of GLUT4 glucose transporter protein and mRNA expression than white skeletal muscle. In addition, the inhibition of insulin-stimulated glucose transport activity in both red and white muscle precedes the decrease in GLUT4 protein and mRNA levels. Thus, STZ treatment initially results in a rapid uncoupling of the insulin-mediated signaling of glucose transport activity which is independent of GLUT4 protein and mRNA levels.     

Up-Regulation of Adiponectin Expression in Antigravitational Soleus Muscle in Response to Unloading Followed by Reloading,and Functional Overloading in Mice

The purpose of this study was to investigate the expression level of adiponectin and its related molecules in hypertrophied and atrophied skeletal muscle in mice. The expression was also evaluated in C2C12 myoblasts and myotubes. Both mRNA and protein expression of adiponectin, mRNA expression of adiponectin receptor (AdipoR) 1 and AdipoR2, and protein expression of adaptor protein containing pleckstrin homology domain, phosphotyrosine binding domain, and leucine zipper motif 1 (APPL1) were observed in C2C12 myoblasts. The expression levels of these molecules in myotubes were higher than those in myoblasts. The expression of adiponectin-related molecules in soleus muscle was observed at mRNA (adiponectin, AdipoR1, AdipoR2) and protein (adiponectin, APPL1) levels. The protein expression levels of adiponectin and APPL1 were up-regulated by 3 weeks of functional overloading. Down-regulation of AdipoR1 mRNA, but not AdipoR2 mRNA, was observed in atrophied soleus muscle. The expression of adiponectin protein, AdipoR1 mRNA, and APPL1 protein was up-regulated during regrowth of unloading-associated atrophied soleus muscle. Mechanical loading, which could increase skeletal muscle mass, might be a useful stimulus for the up-regulations of adiponectin and its related molecules in skeletal muscle.     

Effect of denervation or unweighting on GLUT-4 protein in rat soleus muscle.

The purpose of this study was to test the hypothesis that the decreased capacity for glucose transport in the denervated rat soleus and the increased capacity for glucose transport in the unweighted rat soleus are related to changes in the expression of the regulatable glucose transporter protein in skeletal muscle (GLUT-4). One day after sciatic nerve sectioning, when decreases in the stimulation of soleus 2-deoxyglucose (2-DG) uptake by insulin (-51%, P less than 0.001), contractions (-29%, P less than 0.05), or insulin and contractions in combination (-40%, P less than 0.001) were observed, there was a slight (-18%, NS) decrease in GLUT-4 protein. By day 3 of denervation, stimulation of 2-DG uptake by insulin (-74%, P less than 0.001), contractions (-31%, P less than 0.001), or the two stimuli in combination (-59%, P less than 0.001), as well as GLUT-4 protein (-52%, P less than 0.001), was further reduced. Soleus muscle from hindlimb-suspended rats, which develops an enhanced capacity for insulin-stimulated glucose transport, showed muscle atrophy similar to denervated soleus but, in contrast, displayed substantial increases in GLUT-4 protein after 3 (+35%, P less than 0.05) and 7 days (+107%, P less than 0.001). These results indicate that altered GLUT-4 expression may be a major contributor to the changes in insulin-stimulated glucose transport that are observed with denervation and unweighting. We conclude that muscle activity is an important factor in the regulation of GLUT-4 expression in skeletal muscle.     

Lactate dehydrogenase expression at the onset of altered loading in rat soleus muscle.

Both functional overload and hindlimb disuse induce significant energy-dependent remodeling of skeletal muscle. Lactate dehydrogenase (LDH), an important enzyme involved in anaerobic glycolysis, catalyzes the interconversion of lactate and pyruvate critical for meeting rapid high-energy demands. The purpose of this study was to determine rat soleus LDH-A and -B isoform expression, mRNA abundance, and enzymatic activity at the onset of increased or decreased loading in the rat soleus muscle. The soleus muscles from male Sprague-Dawley rats were functionally overloaded for up to 3 days by a modified synergist ablation or subjected to disuse by hindlimb suspension for 3 days. LDH mRNA concentration was determined by Northern blotting, LDH protein isoenzyme composition was determined by zymogram analysis, and LDH enzymatic activity was determined spectrophotometrically. LDH-A mRNA abundance increased by 372%, and LDH-B mRNA abundance decreased by 43 and 31% after 24 h and 3 days of functional overload, respectively, compared with that in control rats. LDH protein expression demonstrated a shift by decreasing LDH-B isoforms and increasing LDH-A isoforms. LDH-B activity decreased 80% after 3 days of functional overload. Additionally, LDH-A activity increased by 234% following 3 days of hindlimb suspension. However, neither LDH-A or LDH-B mRNA abundance was affected following 3 days of hindlimb suspension. In summary, the onset of altered loading induced a differential expression of LDH-A and -B in the rat soleus muscle, favoring rapid energy production. Long-term altered loading is associated with myofiber conversion; however, the rapid changes in LDH at the onset of altered loading may be involved in other physiological processes.     

Acute effect of fatty acids on metabolism and mitochondrial coupling in skeletal muscle

Acute effects of free fatty acids (FFA) were investigated on: (1) glucose oxidation, and UCP-2 and -3 mRNA and protein levels in 1 h incubated rat soleus and extensor digitorium longus (EDL) muscles, (2) mitochondrial membrane potential in cultured skeletal muscle cells, (3) respiratory activity and transmembrane electrical potential in mitochondria isolated from rat skeletal muscle, and (4) oxygen consumption by anesthetized rats. Long-chain FFA increased both basal and insulin-stimulated glucose oxidation in incubated rat soleus and EDL muscles and reduced mitochondrial membrane potential in C2C12 myotubes and rat skeletal muscle cells. Caprylic, palmitic, oleic, and linoleic acid increased O₂ consumption and decreased electrical membrane potential in isolated mitochondria from rat skeletal muscles. FFA did not alter UCP-2 and -3 mRNA and protein levels in rat soleus and EDL muscles. Palmitic acid increased oxygen consumption by anesthetized rats. These results suggest that long-chain FFA acutely lead to mitochondrial uncoupling in skeletal muscle.     

mRNA levels for alpha-subunit of prolyl 4-hydroxylase and fibrillar collagens in immobilized rat skeletal muscle.

There is evidence that immobilization causes a decrease in total collagen synthesis in skeletal muscle within a few days. In this study, early immobilization effects on the expression of prolyl 4-hydroxylase (PH) and the main fibrillar collagens at mRNA and protein levels were investigated in rat skeletal muscle. The right hindlimb was immobilized in full plantar flexion for 1, 3, and 7 days. Steady-state mRNAs for alpha- and beta-subunits of PH and type I and III procollagen, PH activity, and collagen content were measured in gastrocnemius and plantaris muscles. Type I and III procollagen mRNAs were also measured in soleus and tibialis anterior muscles. The mRNA level for the PH alpha-subunit decreased by 49 and 55% (P < 0.01) in gastrocnemius muscle and by 41 and 39% (P < 0.05) in plantaris muscle after immobilization for 1 and 3 days, respectively. PH activity was decreased (P < 0.05-0.01) in both muscles at days 3 and 7. The mRNA levels for type I and III procollagen were decreased by 26-56% (P < 0.05-0.001) in soleus, tibialis anterior, and plantaris muscles at day 3. The present results thus suggest that pretranslational downregulation plays a key role in fibrillar collagen synthesis in the early phase of immobilization-induced muscle atrophy.