Slow Myosin Light Chains Research Articles

Stretch-induced transformations in myosin expression of quail anterior latissimus dorsi muscle

We have previously found alterations in expression of native myosin isoforms during adaptations to 30 d of stretch-induced hypertrophy in the quail anterior latissimus dorsi muscle (ALD). However, myosin heavy and light chain expression have not been studied during adaptations to stretch. In the current study, expression of myosin heavy and light chains and native myosin isoforms were characterized after placing a weight on one wing of Japanese quail for 0, 7, 14, or 21 d (N = 8 birds.d-1). ALD muscle mass was significantly greater at days 14 (67.0 +/- 4.4%) and 21 (70.2 +/- 4.0%) of stretch compared to day 7 (53.6 +/- 2.9%) of stretch. Native slow myosin-2 (SM2) expression increased from 43.1 +/- 17% in control muscles to 55 +/- 1.2% in stretch muscles, relative to the total myosin pool. The increase in native SM2 expression (approximately 12%) was observed at all three time points of stretch. In contrast, native slow myosin-1 (SM1) expression significantly decreased from 34.1% +/- 1.7% in the control muscles to 24.6 +/- 1.2% in the stretch muscles, relative to the total myosin pool. There was an increased expression of slowly heavy chain-two (SHC2) concurrent with a downregulation of the slow heavy chain-1 (shc1). There was no change in the relative percentages of either slow or fast myosin light chains in overloaded muscles at any of the time points following stretch. These results demonstrate that changes in myosin heavy chain expression, but not myosin light chain expression accompany stretch-induced hypertrophy in the quail ALD.

低頻度持続電気刺激による雄家兎外尿道括約筋の表現型の変化について

The present study investigates the effects of chronic low-frequency electrical stimulation on the external urethral sphincter (EUS) of rabbits through the biochemical analysis of the isoforms of myosin light and heavy chains. Twenty-eight adult male rabbits were used in the test. Of these, the EUS of 14 rabbits were continuously stimulated directly at 1.8 Hz, 0.8 msec, 5.0 V with an implantable electrical cardiac pacemaker for more than 10 weeks. The other 14 rabbits were used as the control group and were nurtured under the same conditions as the stimulated group but without the electrical stimulation. Upon conclusion of the stimulation program, the urethra was removed from all 28 rabbits. The portion of the urethra containing the EUS (from the middle of the prostate to the pelvic diaphragm) was cut transversely into thin serial sections and glycerinated. The glycerinated muscle fibers were then isolated under a stereomicroscope and samples for electrophoretic analysis were prepared. Two-dimensional electrophoresis according to the procedure of O'Farrell for myosin light chains and SDS-PAGE containing 40% glycerol for myosin heavy chains were carried out. The molar ratio of myosin subunits was determined by quantification through the dye elution process. The average percentages of slow myosin light chain molecules in 8 unstimulated and 8 stimulated EUS were 33.4 +/- 8.9% (mean +/- SD) and 70.1 +/- 12.8%, respectively. The average percentages of slow myosin heavy chain molecules in 6 unstimulated and 6 stimulated EUS were 17.6 +/- 5.7% and 40.2 +/- 7.1%, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

Effects of thyroid hormone on fast- and slow-twitch skeletal muscles in young and old rats.

1. The effects of 4 weeks of thyroid hormone treatment on contractile, enzyme-histochemical and morphometric properties and on the myosin isoform composition were compared in the slow-twitch soleus and the fast-twitch extensor digitorum longus (EDL) muscle in young (3-6 months) and old (20-24 months) male rats. 2. In the soleus of untreated controls, contraction and half-relaxation times of the isometric twitch increased by 19-32% with age. The change in contractile properties was paralleled by an age-related increase in the proportions of type I fibres and type I myosin heavy chain (MHC) and slow myosin light chain (MLC) isoforms. 3. In the EDL of controls, contraction and half-relaxation times were significantly prolonged (21-38%) in the post-tetanus twitch in the old animals. No significant age-related changes were observed in enzyme-histochemical fibre-type proportions, although the number of fibres expressing both type IIA and IIB MHCs and of fibres expressing slow MLC isoforms was increased in the old animals. 4. Serum 3,5,3',5'-tetraiodothyronine (T4) levels were lower (34%) in the old animals, but the primary byproduct of T4, 3,5,3'-triiodothyronine (T3), did not differ between young and old animals. 5. The effects of 4 weeks of thyroid hormone treatment were highly muscle specific, and were more pronounced in soleus than in EDL, irrespective of animal age. In the soleus, this treatment shortened the contraction and half-relaxation times by 35-57% and decreased the number of type I fibres by 66-77% in both young and old animals. In EDL, thyroid hormone treatment significantly shortened the contraction time by 24%, but the change was restricted to the old animals. 6. In conclusion, the ability of skeletal muscle to respond to thyroid hormone treatment was not impaired in old age and the age-related changes in speed of contraction and enzyme-histochemical properties and myosin isoform compositions were diminished after thyroid hormone treatment in both the soleus and EDL.

Clones of Human Satellite Cells Can Express in Vitro both Fast and Slow Myosin Heavy Chains

In order to test the diversification among satellite cells in man, satellite cells were isolated from human quadriceps and masseter muscles. The growth kinetics and morphological features of these cells were determined in vitro and the expression of the different myosin heavy (embryonic, fetal, fast, and slow) and light chain isoforms was analyzed. In all satellite cell cultures, only the four fast-type light chains (MLClemb, MLC1F, MLC2F, and MLC3F) were synthesized and no slow myosin light chains were ever detected. However, we found that fused cultures of human satellite cells express both adult fast and slow myosin heavy chains (MHCs), in addition to embryonic and fetal isoforms. In order to determine if distinct fast and slow cell lineages could be detected among the satellite cells, a clonal analysis was carried out on both cell populations. This analysis was first carried out on clonal populations and was confirmed by the analysis of isolated clones. Double-labeling experiments confirmed that all myogenic clones which expressed fast MHC also coexpressed slow MHC. Therefore, we found no evidence for the existence of different fast and slow satellite cell lineages in human postnatal skeletal muscle.

Stimulation-induced expression of slow muscle myosin in a fast muscle of the rat: Evidence of an unrestricted adaptive capacity

Fast muscles of the rat hind limb were stimulated continuously at 10 or 20 Hz for periods of 55–61 days by means of an implantable neuromuscular stimulator. Gel electrophoresis clearly demonstrated the presence in stimulated muscles of slow myosin light and heavy chains, although fast isoforms were still present in all cases. Thus, contrary to previous reports, induction of slow myosin isoforms does occur in this, as in other, mammalian species. The time course of the response to stimulation appears to be more extended than that seen in the rabbit.

Developmental modulation of myosin expression by thyroid hormone in avian skeletal muscle.

It is well established that a rise in circulating thyroid hormone during the second half of chick embryo development significantly influences muscle weight gain and bone growth. We studied thyroid influence on differentiation in slow anterior latissimus dorsi (ALD) and fast posterior latissimus dorsi (PLD) muscles of embryos rendered hypothyroid by hypophysectomy or administration of an anti-thyroid drug. The expression of native myosins and myosin light chains (MLCs) was studied by electrophoretic analysis, and the myosin heavy chain (MHC) was characterized by immunohistochemistry. The first effects of hypothyroid status were observed at day 21 of embryonic development (stage 46 according to Hamburger and Hamilton). Analysis of myosin isoform expression in PLD muscles of hypothyroid embryos showed persistence of slow migrating native myosins and slow MLCs as well as inhibition of neonatal fast MHC expression, indicating retarded differentiation of this muscle. In ALD muscle, hypothyroidism maintained fast embryonic MHC and induced noticeable amounts of fast MLCs, thus delaying slow muscle differentiation. Our results suggest that thyroid hormones play a role in modulating the appearance of neonatal fast MHC and the disappearance of isomyosins transiently present during embryogenesis. However, T3 supplemental treatment would seem to compensate in part for the effects of hypothyroidism induced by hypophysectomy, suggesting that thyroid hormone might interfere with other factors also accounting for the observed effects.

Muscle sarcomere lesions and thrombosis after spaceflight and suspension unloading.

Spaceflight (flight) and tail suspension-hindlimb unloading (unloaded) produced significant decreases in fiber cross-sectional areas of the adductor longus (AL), a slow-twitch antigravity muscle. However, the mean wet weight of the flight AL muscles was near normal, whereas that of the suspension unloaded AL muscles was significantly reduced. Interstitial edema within the flight AL, but not in the unloaded AL, appeared to account for this apparent disagreement. In both experimental conditions, the slow-twitch oxidative fibers atrophied more than the fast-twitch oxidative-glycolytic and fast-twitch glycolytic fibers. Immunostaining showed that slow-twitch oxidative fibers expressed fast myosin, producing hybrid fibers containing slow and fast myosin isoforms. Two-dimensional gel electrophoresis of flight AL muscles revealed increased content of fast myosin light chains and decreased amounts of slow myosin light chains and fatty acid-binding protein. In the flight AL, absolute mitochondrial content decreased, but the relatively greater breakdown of myofibrillar proteins maintained mitochondrial concentration near normal in the central intermyofibrillar regions of fibers. Subsarcolemmal mitochondria were preferentially lost and reduced below normal concentration. Elevated fiber immunostaining for ubiquitin conjugates was suggestive of ubiquitin-mediated breakdown of myofibrillar proteins. On return to weight bearing for 8-11 h, the weakened atrophic muscles exhibited eccentric contraction-like lesions (hyperextension of sarcomeres with A-band filaments pulled apart and fragmented), tearing of the supporting connective tissue, and thrombosis of the microcirculation. Segmental necrosis of muscle fibers, denervation of neuromuscular junctions, and extravasation of red blood cells were minimal. Lymphocyte antibody markers did not indicate a significant immune reaction. The flight AL exhibited threefold more eccentric-like lesions than the unloaded AL; the high reentry G forces experienced by the flight animals, but not the unloaded group, possibly accounted for this difference. Muscle atrophy appears to increase the susceptibility to form eccentric contraction-like lesions after reloading; this may reflect weakening of the myofibrils and extracellular matrix. Microcirculation was also compromised by spaceflight, such that there was increased formation of thrombi in the post-capillary venules and capillaries. This blockage led to edema by 8-11 h after resumption of weight bearing by the COSMOS 2044 rats. The present findings indicate that defective microcirculation most likely accounted for the extensive tissue necrosis and microhemorrhages observed for COSMOS 1887 rats killed 2 days after landing.

Age effects on myosin subunit and biochemical alterations with skeletal muscle hypertrophy.

The purpose of this study was to determine whether skeletal muscle mass, myofibrillar adenosinetriphosphatase activity, and the expression of myosin heavy (MHC) and light chain subunits are differentially affected in juvenile (4 wk) and young adult (12 wk) rats by a hypertrophic growth stimulus. Hypertrophy of the plantaris or soleus was studied 4 wk after ablation of either two [gastrocnemius (GTN) and soleus or plantaris] or one (GTN) synergistic muscle(s). There was no difference in the relative magnitude of hypertrophy because of age. Plantaris myofibrillar adenosinetriphosphatase activity was decreased 21 and 12% in juvenile and adult rats, respectively, as a result of ablation of both the GTN and soleus. Slow myosin light chain isoforms (1s and 2s) were expressed to a greater extent in hypertrophied plantaris muscles of both ages, but the increase in 1s was greater in juvenile rats. The relative expression of slow beta-MHC in hypertrophied plantaris muscles increased by 470 and 350%, whereas MHC IIb decreased by 70 and 33% in juvenile and adult rats, respectively. The relative expression of MHC IIa increased (56%) in the plantaris after ablation in juvenile rats only. These shifts in myosin subunit expression and the increases in mass were generally about one-half the magnitude when only the GTN was removed. There were no detectable myosin shifts in hypertrophied soleus muscles. Although the extent of muscle hypertrophy is similar, the shifts in myosin subunits were greater in juvenile than in young adult rats.

Biphasic expression of slow myosin light chains and slow tropomyosin isoforms during the development of the human quadriceps muscle

Using a two-dimensional electrophoresis technique coupled with sensitive silver staining, we have investigated the chronology of appearance of the myosin light chain and tropomyosin isoforms during early stages of human quadriceps development. Our results show that slow myosin light chains and the slow tropomyosin isoform are not detected at 6 weeks of gestation. These isoforms transiently appear between 12.5 weeks and 15 weeks of gestation and then disappear. The slow myosin light chains are re-expressed at 31 weeks of gestation and the slow tropomyosin isoform later at 36 weeks of gestation, and normally remained expressed into the adulthood. Our study thus reveals a biphasic expression of the slow myosin light chains and the slow tropomyosin isoform in developing human quadriceps muscle.

Temperature Acclimation in the Common Carp: Force-Velocity Characteristics and Myosin Subunit Composition of Slow Muscle Fibres

ABSTRACT Live slow fibre bundles were isolated from the superficial region of the pectoral fin abductor superficialis muscle of common carp (Cyprinus carpio L.) acclimated to either 8 or 20°C. The maximum tetanic tension (P0) of fibre bundles was similar when measured at the acclimation temperature of each group. However, at 8°C, P0 was significantly higher in 8 °C-than in 20°C-acclimated fish (202 ±8 versus 153±4kNm−2, respectively). For isometric tetani at 8°C, the times to 50% peak force and from peak force to 50 % relaxation were 15–20 % faster in preparations from cold-than from warm-acclimated carp. Force-velocity (P–V) curves were fitted using a hyperbolic-linear equation. The curvature of the P–V relationship was found to be independent of acclimation temperature. Unloaded contraction velocity (Vmax) was 17 % higher at 8°C in fibres from fish acclimated to 8°C than in fish acclimated to 20°C (1.18±0.04 and 0.98±0.04 muscle lengths s−1, respectively). Calculated values for maximum power output at 8°C were 26.5 W kg−1 for cold-acclimated and 18.0 W kg−1 for warm-acclimated fish. Native myosin was purified from isolated fibre bundles using sodium pyrophosphate gel electrophoresis. The mobility of myosin heavy chains on 8% SDS– PAGE gels was similar for both acclimation groups. Myosin light chain subunits were separated on 15% SDS–PAGE gels. Fibre bundles from warm-acclimated fish contained almost exclusively slow myosin light chains (LC1S and LC2S). Preparations from cold-acclimated fish contained a significant proportion of fast myosin light chains (LClf and LC2f) in addition to LC1S and LC2S. Histochemical studies revealed no differences in the fibre composition of preparations from warm- and cold-acclimated fish: both contained an average of 3% fast oxidative fibres in addition to slow fibres. It is concluded that cold-acclimation results in modest improvements in the contractile performance of slow muscle fibres at low temperatures. The mechanism may involve the expression of myosin light chain isoforms normally associated with faster-contracting fibre types.

The proportions of fiber types in human external urethral sphincter: electrophoretic analysis of myosin

The proportions of fast and slow myosin molecules in external urethral sphincter specimens from ten urodynamically normal male bladder carcinoma patients were estimated from the contents of fast and slow myosin light chains in two-dimensional electrophoretic gels. The percentages of fast and slow myosin molecules ranged from 5.0% to 61.4% with a mean of 35.5% and from 38.6% to 95.0% with a mean of 65.5% respectively. It is therefore concluded that the human external urethral sphincter is composed of both fast and slow muscle fibers as well as other voluntary muscles. The human external urethral sphincter is considered to be a highly fatigue-resistant muscle with a very high proportion of slow muscle fibers. In the cases studied so far, there is a great diversity in the proportions of fast and slow myosin molecules; the reason for this remains unknown.

Myosin heavy and light chain expression during human skeletal muscle development and precocious muscle maturation induced by thyroid hormone

It has now been well established that during mammalian muscle development there is a sequential transition of the myosin isoforms, with the developmental isoforms being replaced just before or just after birth by the adult isozymes. In a previous study of human fetal muscle, we demonstrated the differentiation of two fiber populations as early as 15 weeks: one population of large diameter fibers containing predominantly slow myosin heavy and light chains, and another population which stained homogeneously for fetal myosin heavy chain and corresponded to histochemical type IIC fibers. We have carried out an immunocytochemical and biochemical study of human fetal quadriceps between 7 and 40 weeks. A chronology of the changes which occur in the expression of the myosin heavy and light chains is correlated with the results obtained by enzyme histochemistry. Evidence is also presented that in man excessive amounts of thyroid hormone act directly on the muscle, and result in a precocious accumulation of the adult myosin heavy chains and a precocious maturation of the muscle.

Comparative effects of hindlimb suspension and exercise on skeletal muscle myosin isozymes in rats

The purpose of this study was to ascertain the time course of changes, whilst suspending the hindlimb and physical exercise training, of myosin light chain (LC) isoform expression in rat soleus and vastus lateralis muscles. Two groups of six rats were suspended by their tails for 1 or 2 weeks, two other groups of ten rats each were subjected to exercise training on a treadmill for 9 weeks, one to an endurance training programme (1-h running at 20 m.min-1 5 days.week-1), and the other to a sprint programme (30-s bouts of running at 60 m.min-1 with rest periods of 5 min). At the end of these experimental procedures, soleus and vastus lateralis superficialis muscles were removed for myosin LC isoform determination by two-dimensional gel electrophoresis. Hindlimb suspension for 2 weeks significantly increased the proportion of fast myosin LC and decreased slow myosin LC expression in the soleus muscle. The pattern of myosin LC was unchanged in the vastus lateralis muscle. Sprint training or endurance training for 9 weeks increased the percentage of slow myosin LC in vastus lateralis muscle, whereas soleus muscle myosin LC was not modified. These data indicate that hindlimb suspension influences myosin LC expression in postural muscle, whereas physical training acts essentially on phasic muscle. There were no differences in myosin LC observed under the influence of sprint- or endurance-training programme.

Human skeletal muscle myosin light chains analyzed by immobilized pH gradients during ontogenesis: Identification of new phosphorylatable isoforms of light chain 2

Previous studies using two-dimensional gel electrophoresis have described adult and fetal isoforms of skeletal muscle myosin light chains (MLC). They have also revealed an embryo-specific light chain (LC1emb), apparently absent in most adult skeletal muscles. In order to characterize more thoroughly the MLC family, we have analyzed the MLCs from human skeletal muscle at different developmental stages using a two-dimensional electrophoresis technique with an immobilized pH gradient in the first dimension. The high resolution of this novel technique, resolving components which in isoelectric points are less than or equal to 0.01 pH, combined with sensitive silver staining, has allowed us to identify four phosphorylatable isoforms of MLC2: two slow-myosin light chains (MLC2Sa and b), two fast myosin light chains (MLC2Fa and b), and their phosphorylated counterparts: MLC2SaP and bP, MLC2FaP and bP. The following major modifications during development were observed: (i) The embryonic LC (LC1emb) persists up to at least 26 weeks of fetal life. (ii) The polymorphism of LC2 is already evident at 10 weeks of development but only the nonphosphorylated forms of LC2S and LC2F seem to be present. The LC2Fa form is predominant. As early as 26 weeks of fetal life, the 4 phosphorylated forms are detected. In the adult, LC2Fb is a minor component. (iii) LC3F (fast) is already expressed at an early embryonic stage (10 weeks).

Is the myosin type altered in the ageing platysma?

The myosin light chains of platysma samples originating from 2 to 86 year old patients of the maxillo-facial surgery clinic were investigated. The platysma contained both fast and slow myosin. No change in the proportion of myosin type in relation to age was found. Female patients often displayed less fast myosin than male patients. Each time when the platysma muscle activity has been reduced a decrease in the amount of slow myosin light chains was observed. In the reported study, variations of the fast and slow myosin light chains resulted mainly from the platysma activity level, i.e. from the pathology requiring the surgery, rather than from the patients' age.

Electrostimulation-induced fast-to-slow transitions of myosin light and heavy chains in rabbit fast-twitch muscle at the mRNA level

Chronic low-frequency stimulation of rabbit fast-twitch muscle induces progressive increases in slow myosin light chain mRNAs followed by an increase in the slow myosin heavy chain HCI mRNA. Therefore, the effects of chronic stimulation are more pronounced in rabbit than in rat fast-twitch muscle. The latter responds mainly with a rearrangement of its fast isomyosin pattern.

A comparative study of chicken ventricular and slow skeletal myosin light chains

1. 1_ Comparison of ventricular and slow skeletal myosin light chains was performed using high performance liquid chromatography (HPLC), peptide mapping of electrophoretically purified proteins and immunoblots. 2. 2_ Both in rabbit and in chicken ventricular and slow skeletal myosin light chain composition was identical.

Effects of chronic sympathectomy on muscle fibre composition ISO-myosin patterns, protein synthesis and calcium content in canine gracilis muscle.

We have previously reported functional and histological studies in five beagle dogs with unilateral lumbar sympathectomy. Three months later, fatiguability in the gracilis muscles was increased on the denervated sides, and this was associated with an increase in the relative distribution of FT fibres. Biochemical studies now show that these changes were associated with an increase in cytosolic protein without change in DNA content; this is consistent with an increase in cell size. There was a reduction in the proportion of slow myosin light chain isoforms from 50 +/- 7 to 34 +/- 6%. Noradrenaline levels were increased on the denervated sides but this may reflect greater vascularity. Calcium content did not correlate with fibre type but there was a positive relation with both noradrenaline content (r = 0.73; P less than 0.05) and DNA content (r = 0.84; P less than 0.05). It is concluded that sympathectomy induces several biochemical changes in skeletal muscle which constitute a change and increase in fast myosin light chain synthesis and a corresponding fibre type transformation.

Innervation, rythme d'activité et accumulation des chaînes légères de myosine d'un muscle rapide de poulet

The fast posterior latissimus dorsi muscle (PLD) of the chick ceases to accumulate slow myosin light chains (MLC) during neonatal development. On day 18 of embryonic life slow MLC represented only 2% of total MLC, and LC3F was first detected. In chick embryo, spinal cord stimulation at a slow rhythm modifies PLD differentiation toward the slow type: LC3F did not accumulate and slow MLC increased. In contrast, stimulation at a fast rhythm accelerated LC3F accumulation. PLD denervation on day 2 after hatching inhibited the synthesis of LC3F. Direct stimulation at a fast rhythm led to post-hatching development into normal fast type while a slow rhythm influenced the development of denervated PLD towards the slow type. In innervated PLD, the effect of stimulation at a slow rhythm was less important than in denervated PLD. These results suggest that the rhythm of the neural and/or contractile activity plays an important role in the MLC expression during embryonic and post-natal development of the chicken fast muscle.

高強度・持久トレーニングによるラット骨格筋タンパクの変化

Effects of long-term with high intensity endurance training upon histochemically assessed myofibrillar actomyosin ATPase, myosin composition and creatine kinase content were analyzed in muscles from rats which were trained to run up to 240min/day at 40m/min. Following 16 weeks of the training, in M. soleus, proportions of slow-twitch oxidative (S0) fibers and slow type myosin light chains were increased; no changes in myosin light chain were significantly observed. Furthermore, a decrease in creatine kinase was found in this muscle, indicating that SO fibers might have acquired slower properties. On the other hand, in M. extensor digitorum longus, there was only a transformation between fast-twitch fiber subgroups, with a decrease in LC3f by which fast myosin constructed was characterized by the highest myofibrillar ATPase activities. These data suggest that fiber type transformation may be brought about not only from fast-twitch glycolytic (FG) fibers to fast-twitch oxdative glycolytic (FOG) fibers but also from FOG fibers to SO fibers by long-term with high intensity endurance training.