IIA Myofibers Research Articles

Resident stem cells are not required for exercise-induced fiber-type switching and angiogenesis but are necessary for activity-dependent muscle growth

Skeletal muscle undergoes active remodeling in response to endurance exercise training, and the underlying mechanisms of this remodeling remain to be defined fully. We have recently obtained evidence that voluntary running induces cell cycle gene expression and cell proliferation in mouse plantaris muscles that undergo fast-to-slow fiber-type switching and angiogenesis after long-term exercise. To ascertain the functional role of cell proliferation in skeletal muscle adaptation, we performed in vivo 5-bromo-2'-deoxyuridine (BrdU) pulse labeling (a single intraperitoneal injection), which demonstrated a phasic increase (5- to 10-fold) in BrdU-positive cells in plantaris muscle between days 3 and 14 during 4 wk of voluntary running. Daily intraperitoneal injection of BrdU for 4 wk labeled 2.0% and 15.4% of the nuclei in plantaris muscle in sedentary and trained mice, respectively, and revealed the myogenic and angiogenic fates of the majority of proliferative cells. Ablation of resident stem cell activity by X-ray irradiation did not prevent voluntary running-induced increases of type IIa myofibers and CD31-positive endothelial cells but completely blocked the increase in muscle mass. These findings suggest that resident stem cell proliferation is not required for exercise-induced type IIb-to-IIa fiber-type switching and angiogenesis but is required for activity-dependent muscle growth. The origin of the angiogenic cells in this physiological exercise model remains to be determined.

Efficacy of 3 days/wk resistance training on myofiber hypertrophy and myogenic mechanisms in young vs. older adults

Resistance training (RT) has shown the most promise in reducing/reversing effects of sarcopenia, although the optimum regime specific for older adults remains unclear. We hypothesized myofiber hypertrophy resulting from frequent (3 days/wk, 16 wk) RT would be impaired in older (O; 60-75 yr; 12 women, 13 men), sarcopenic adults compared with young (Y; 20-35 yr; 11 women, 13 men) due to slowed repair/regeneration processes. Myofiber-type distribution and cross-sectional area (CSA) were determined at 0 and 16 wk. Transcript and protein levels of myogenic regulatory factors (MRFs) were assessed as markers of regeneration at 0 and 24 h postexercise, and after 16 wk. Only Y increased type I CSA 18% (P < 0.001). O showed smaller type IIa (-16%) and type IIx (-24%) myofibers before training (P < 0.05), with differences most notable in women. Both age groups increased type IIa (O, 16%; Y, 25%) and mean type II (O, 23%; Y, 32%) size (P < 0.05). Growth was generally most favorable in young men. Percent change scores on fiber size revealed an age x gender interaction for type I fibers (P < 0.05) as growth among Y (25%) exceeded that of O (4%) men. Myogenin and myogenic differentiation factor D (MyoD) mRNAs increased (P < 0.05) in Y and O, whereas myogenic factor (myf)-5 mRNA increased in Y only (P < 0.05). Myf-6 protein increased (P < 0.05) in both Y and O. The results generally support our hypothesis as 3 days/wk training led to more robust hypertrophy in Y vs. O, particularly among men. However, this differential hypertrophy adaptation was not explained by age variation in MRF expression.

Resting and load-induced levels of myogenic gene transcripts differ between older adults with demonstrable sarcopenia and young men and women

Regenerative capacity appears to be impaired in sarcopenic muscle. As local growth factors and myogenic regulatory factors (MRFs) modulate repair/regeneration responses after overload, we hypothesized that resistance loading (RL)-induced expression of MRFs and muscle IGF-I-related genes would be blunted in older (O) males (M) and females (F) with demonstrable sarcopenia vs. young (Y) adults. Y (20-35 yr, 10 YF, 10 YM) and O (60-75 yr, 9 OF, 9 OM) underwent vastus lateralis biopsy before and 24 h after knee extensor RL. Sarcopenia was assessed by cross-sectional area of type I, IIa, and IIx myofibers. Transcript levels were assessed by relative RT-PCR and analyzed by age x gender x load repeated-measures ANOVA. O were sarcopenic based on type II atrophy with smaller type IIa (P < 0.05) and IIx (P < 0.001) myofibers. Within-gender cross-sectional area differences were more marked in F (OF < YF: IIa 21%, IIx 42%). Load effects (P < 0.05) were seen for four of seven mRNAs as IGF-IEa (34%), myogenin (53%), and MyoD (20%) increased, and myf-6 declined 10%. Increased IGF-IEa was driven by O (48%) and/or M (43%). An age x gender x load interaction was found for MyoD (P < 0.05). An age x load interaction for type 1 IGF receptor (P < 0.05) was driven by a small increase in O (16%, P < 0.05). A gender x load interaction (P < 0.05) was noted for IGF binding protein-4. Age effects (P < 0.05) resulted from higher MyoD (54%), myf-5 (21%), and IGF binding protein-4 (17%) in O and were primarily localized to F at baseline (OF > YF; MyoD 94%, myf-5 47%, P < 0.05). We conclude that RL acutely increases mRNA expression of IGF-IEa and myogenin, which may promote growth/regeneration in both Y and O. Higher resting levels of MRFs in OF vs. YF suggest elevated basal regenerative activity in sarcopenic muscle of OF.

Exercise Stimulates Pgc-1α Transcription in Skeletal Muscle through Activation of the p38 MAPK Pathway

Peroxisome proliferator-activated receptor gamma co-activator 1alpha (PGC-1alpha) promotes mitochondrial biogenesis and slow fiber formation in skeletal muscle. We hypothesized that activation of the p38 mitogen-activated protein kinase (MAPK) pathway in response to increased muscle activity stimulated Pgc-1alpha gene transcription as part of the mechanisms for skeletal muscle adaptation. Here we report that a single bout of voluntary running induced a transient increase of Pgc-1alpha mRNA expression in mouse plantaris muscle, concurrent with an activation of the p38 MAPK pathway. Activation of the p38 MAPK pathway in cultured C2C12 myocytes stimulated Pgc-1alpha promoter activity, which could be blocked by the specific inhibitors of p38, SB203580 and SB202190, or a dominant negative p38. Furthermore, the p38-mediated increase in Pgc-1alpha promoter activity was enhanced by increased expression of the downstream transcription factor ATF2 and completely blocked by ATF2DeltaN, a dominant negative ATF2. Skeletal muscle-specific expression of a constitutively active activator of p38, MKK6E, in transgenic mice resulted in enhanced Pgc-1alpha and cytochrome oxidase IV protein expression in fast-twitch skeletal muscles. These findings suggest that contractile activity-induced activation of the p38 MAPK pathway promotes Pgc-1alpha gene expression and skeletal muscle adaptation.

Mechanical stimulation of the plantar foot surface attenuates soleus muscle atrophy induced by hindlimb unloading in rats

Unloading-induced muscle atrophy occurs in the aging population, bed-ridden patients, and astronauts. This study was designed to determine whether dynamic foot stimulation (DFS) applied to the plantar surface of the rat foot can serve as a countermeasure to soleus muscle atrophy normally observed in hindlimb unloaded (HU) rats. Forty-four mature (6 mo old), male Wistar rats were randomly assigned to ambulatory control, HU alone, HU with active DFS (i.e., plantar contact with active inflation), HU with passive DFS (i.e., plantar contact without active inflation), and HU while wearing a DFS boot with no plantar contact groups. Application of active DFS during HU significantly counteracted the atrophic response by preventing approximately 85% of the reduction in type I myofiber cross-sectional area (CSA) in the soleus while preventing approximately 57% of the reduction in type I myofiber CSA and 43% of the reduction in type IIA myofiber CSA of the medial gastrocnemius muscle. Wearing of a DFS boot without active inflation prevented myofiber atrophy in the soleus of HU animals in a fashion similar to that observed in HU animals that wore an actively inflated DFS boot. However, when a DFS boot without plantar surface contact was worn during HU, no significant protection from HU-induced myofiber atrophy was observed. These results illustrate that the application of mechanical foot stimulation to the plantar surface of the rat foot is an effective countermeasure to muscle atrophy induced by HU.

Voluntary running induces fiber type-specific angiogenesis in mouse skeletal muscle.

Adult skeletal muscle undergoes adaptation in response to endurance exercise, including fast-to-slow fiber type transformation and enhanced angiogenesis. The purpose of this study was to determine the temporal and spatial changes in fiber type composition and capillary density in a mouse model of endurance training. Long-term voluntary running (4 wk) in C57BL/6 mice resulted in an approximately twofold increase in capillary density and capillary-to-fiber ratio in plantaris muscle as measured by indirect immunofluorescence with an antibody against the endothelial cell marker CD31 (466 +/- 16 capillaries/mm2 and 0.95 +/- 0.04 capillaries/fiber in sedentary control mice vs. 909 +/- 55 capillaries/mm2 and 1.70 +/- 0.04 capillaries/fiber in trained mice, respectively; P < 0.001). A significant increase in capillary-to-fiber ratio was present at day 7 with increased concentration of vascular endothelial growth factor (VEGF) in the muscle, before a significant increase in percentage of type IIa myofibers, suggesting that exercise-induced angiogenesis occurs first, followed by fiber type transformation. Further analysis with simultaneous staining of endothelial cells and isoforms of myosin heavy chains (MHCs) showed that the increase in capillary contact manifested transiently in type IIb + IId/x fibers at the time (day 7) of significant increase in total capillary density. These findings suggest that endurance training induces angiogenesis in a subpopulation of type IIb + IId/x fibers before switching to type IIa fibers.

Ste20-like kinase SLK displays myofiber type specificity and is involved in C2C12 myoblast differentiation.

Cell growth and terminal differentiation are controlled by complex signaling cascades that regulate the expression of specific subsets of genes implicated in cell fate and morphogenic processes. We have recently cloned and characterized a novel Ste20-like kinase termed SLK that is associated with adhesion structures during cell adhesion and spreading. However, the specific function of SLK is poorly understood. To gain further insight into the role of SLK, we have characterized its activity, expression, and distribution in skeletal muscle and during the in vitro differentiation of C2C12 myoblasts. Although SLK is expressed ubiquitously in adult tissues, our results show that it is predominantly expressed in muscle masses during development. Furthermore, SLK activity is upregulated during the differentiation of C2C12 myoblasts. In addition, we have found that SLK localizes presynaptically at neuromuscular junctions and that it is preferentially expressed in types I and IIA myofibers at major myofibrillar striations. Supporting a role in myoblast function and differentiation, SLK expression is induced in Myf5- and Pax7-positive activated satellite cells during regeneration and expression of dominant negative SLK in C2C12 cultures impairs myoblast fusion, suggesting a role for SLK in muscle cell differentiation.

Histochemical properties of skeletal muscles in Japanese cattle and their meat production ability

ABSTRACTThe compositional characteristics of the three basic types of myofiber, namely type I (slow‐twitch oxidative), type IIA (fast‐twitch oxidative glycolytic) and type IIB (fast‐twitch glycolytic), are clarified in the skeletal muscles of Japanese Black cattle. The myofiber composition, which is characteristic of the muscles of Japanese Black cattle, markedly changes during their growth, when some type IIA myofibers are transformed into type I or IIB, depending on the different muscles. Independent of these changes with growth, inter‐ and intramuscular variations of myofiber type distribution is evident. The small extensor muscles in deep regions around bone contain a lot of type I myofibers, whereas the large muscles at surface regions have many type II myofibers. Japanese Black cattle have typical white muscles such as the Longissimus thoracis and Semitendinosus, containing half the myofibers as red (type I + IIA). The muscles of Japanese Black cattle show a tendency to contain a higher percentage of type I myofibers than other breeds over an intrabreed variation of the myofiber type composition. In the big muscles such as the Longissimus thoracis and Biceps femoris, a great diversity of myofiber type composition is observed among the different regions. When fattened, heifers produce Longissimus thoracis and Biceps femoris muscles of smaller weight than steers, but in heifers the myofiber size in each type is rather larger. In the Psoas major, Vastus lateralis and Serratus ventralis muscles, heifers contain a higher frequency of red (type I + IIA) myofibers with no differences in myofiber size. Among the several muscles of fattened Japanese Black steers, the percentage distribution of type I myofibers has a positive correlation with the percentage amount of intramuscular fat. From these results, the high potential of Japanese Black cattle to produce marbled beef could be based on the histochemical properties of myofibers in their skeletal muscles.

Gender differences in resistance-training-induced myofiber hypertrophy among older adults.

We tested the hypothesis that older men (n = 9, 69 +/- 2 years) would experience greater resistance-training-induced myofiber hypertrophy than older women (n = 5, 66 +/- 1 years) following knee extensor training 3 days per week at 65-80% of one-repetition maximum for 26 weeks. Vastus lateralis biopsies were analyzed for myofiber areas, myosin heavy chain isoform distribution, and levels of mRNA for insulin-like growth factor 1 (IGF-1), IGFR1, and myogenin. Gender x Training interactions (p <.05) indicate greater myofiber hypertrophy for all three primary fiber types (I, IIa, IIx) and enhanced one-repetition maximum strength gain in men compared with women (p <.05). Covarying for serum IGF-1, dehydroepiandrosterone sulfate, or each muscle mRNA did not negate these interactions. In both genders, type IIx myofiber area distribution and myosin heavy chain type IIx distribution decreased with a concomitant increase in type IIa myofiber area distribution (p <.05). In summary, gender differences in load-induced myofiber hypertrophy among older adults cannot be explained by levels of circulating IGF-1 or dehydroepiandrosterone sulfate, or by expression of the myogenic transcripts examined.

Histology of longissimus muscle from 2-week-old and 8-week-old normal and callipyge lambs

There were no differences in proportion or sizes of myofiber types in longissimus from 2-wk-old lambs of the callipyge and normal genotypes. In 8-wk old lambs, the type I ovine myofibers were oxidative, the majority of type IIA myofibers were glycolytic, and the majority of type IIB myofibers were oxidative. Key words: Callipyge, lamb, muscle, histology

Histochemical Properties of Myofiber Types and Lack of Muscle Spindles in the Caudal Pharyngeal Constrictor Muscles of Sheep

The purpose of the present study was to examine histochemical properties of myofiber types and presence or absence of muscle spindles in the caudal pharyngeal constrictor (thyropharyngeus and cricopharyngeus) muscles of sheep. Unfixed muscle sections were stained with myosin ATPase (preincubation pH4.3 and 10.5), reduced nicotinamido adenine dinucleotide dehydrogenase, and 3-hydroxybutyrate dehydrogenase (3-HBD). Myofiber types were classified by the differences of the histochemical properties. The thyropharyngeus muscle had 12% type I, 88% type IIA, and no type IIB myofibers. The cricopharyngeus muscle possessed 53% type I, 47% type IIA, and no type IIB myofibers. The myofiber types of the thyropharyngeus and cricopharyngeus muscles were small in diameter. Part of type I myofibers showed a strong 3-HBD activity, whereas all type IIA myofibers showed no 3-HBD activity in the two muscles. No muscle spindle was found in the two muscles. The thyropharyngeus and cricopharyngeus muscles composed of types I and IIA myofibers seem to be adapted to close the pharyngeal cavity repeatedly for the rumination.

Effects of fetal spinal cord tissue transplants and cycling exercise on the soleus muscle in spinalized rats.

Studies were carried out to determine if an intraspinal transplant (Trpl) of fetal spinal cord tissue or hind limb exercise (Ex) affected the changes in myosin heavy chain (MyHC) composition or myofiber size that occur following a complete transection (Tx) of the lower thoracic spinal cord of the adult rat. In one group of animals, transplants were made acutely, whereas in a second group, daily cycling exercise was initiated 5 days after injury, with animals in both groups being sacrificed 90 days after injury. The soleus muscle is normally composed of myofibers expressing either type I (90%) or type IIa (10%) MyHC. Following a spinal transection, expression of type I MyHC isoform decreased (18% of myofibers), type IIa MyHC expression increased (65% of myofibers), and the majority of myofibers (80%) expressed type IIx MyHC. Most myofibers coexpressed multiple MyHC isoforms. Compared with Tx only, with Ex or with Trpl, there was a decrease in the number of myofibers expressing type I or IIa isoforms but little change in expression of IIx MyHC. Myofibers expressing the IIb isoform appeared in several transplant recipients but not after exercise. Transection resulted in atrophy of type I myofibers to approximately 50% of normal size, whereas myofibers were significantly larger after exercise (74% of control) and in Trpl recipients (77% of control). Type IIa myofibers also were significantly larger in Trpl recipients compared with the Tx only group. Overall, the mean myofiber size was significantly greater after exercise and in Trpl recipients compared with myofibers in Tx only animals. Thus, although neither strategy shifted the MyHC profile towards the control, both interventions influenced the extent of atrophy observed after spinalization. These data suggest that palliative strategies can be developed to modulate some of the changes in hind limb muscles that occur following a spinal cord injury.

Histochemical Properties of Skeletal Muscles in Different Body Parts of Young Japanese Black Steers

Histochemical examination was carried out on the 66 skeletal muscles in the different body parts of the young Japanese Black steers (11 months of age). Tissues were taken from the central portion of each muscle, cut into serial frozen sections and stained by reactions for myosin ATPase and NADH dehydrogenase activities. Myofibers were divided into Type I, IIA and IIB. Generally the muscles in the young steers contained Type IIA myofibers at higher frequency as compared with the fattened adult. Some Type IIA myofibers seemed to transform into Type I or Type IIB with age. The muscles with much Type I myofibers which are to play a main role in posture maintaining were very small and occupied the deep position, but Mm. serratus ventrales is fairly large. The large muscles important for beef production contained great proportion of Type II myofibers, which are mobilized to do more active motions such as locomotion. These results suggest that every muscle plays its peculiar function at different body parts after obtaining its own histochemical property.

Composition of myofiber types in limb muscles of the house shrew (Suncus murinus): Lack of type I myofibers

Postural muscles have many type I myofibers, which reacted strongly for acid-stable myosin ATPase and were unreactive for alkali-stable myosin ATPase (Ariano et al., J. Histochem. Cytochem., 21:51-55, 1973; Armstrong et al., Am. J. Anat., 163:87-98, 1982; Smith et al., J. Neurophysiol., 40:503-513, 1977). House shrews (Suncus murinus) keep abducting their limbs in locomotion and hardly lift their trunk off the ground. The limb muscles of Suncus were examined by histochemical methods to determine whether the locomotory and postural behavior is related to the proportion of type I myofibers. The observation of whole cross sections from the triceps surae, flexor digitorum superficialis, quadriceps femoris, and caudally situated muscles in the thigh showed that all myofibers of these muscles were unreactive for acid-stable myosin ATPase and strongly reactive for alkali-stable myosin ATPase: Those were classified as type II myofibers. Type II myofibers showed a weak (type IIB), moderate (type IIAB), or strong (type IIA) reaction for NADH tetrazolium reductase. Part of type IIA myofibers reacted weakly to moderately for menadione-linked glycerol-3-phosphate dehydrogenase (m-GPD), which predominated in the soleus muscle. Type IIAB, type IIB, and the remainder of type IIA myofibers reacted strongly for m-GPD. The limb muscles contained subtypes of type II myofibers but no type I myofibers. In Suncus murinus, type I myofibers specialized for a postural maintenance may not be required because all myofibers function exclusively for propulsion.

Distribution of myofiber types in the hip and thigh musculature of sheep.

The composition of muscles by myofiber type is associated with their locomotory or postural functions. In the present study the composition of the hip and thigh musculature of sheep by myofiber types and the differences in their distribution were examined. Myofibers were classified into type I, IIA, and IIB myofibers by differences in myosin ATPase and NADH tetrazolium reductase (NADH-TR) activity. The vastus intermedius muscle consisted only of type I myofibers, which exhibit weak alkali-stable myosin ATPase and strong NADH-TR activity. The gluteus accessorius and profundus muscles had more than 50% type I myofibers. The other muscles had less than 50% type I myofibers as a whole. Type I myofibers were concentrated in the deep portions of the gluteus and quadriceps femoris muscles, which extend the hip and stifle joints, and of the pectineus muscle. They were scattered evenly in the caudally situated locomotory muscles in the thigh. Type IIA myofibers, characterized by strong alkali-stable myosin ATPase and NADH-TR activity, showed little difference in distribution in the hip and thigh muscles. Type IIB myofibers, characterized by strong alkali-stable myosin ATPase and weak NADH-TR activity, were distributed more in the cranial, caudolateral, and caudomedial portions than in the middle portions of the thigh. The distribution of type IIB myofibers is suited to powerful flexion and extension of the thigh and leg. In the hip and thigh musculature, it appears that type I myofibers are effectively distributed to maintain a standing posture without diminishing the propulsive force of the hindlimb.

Distribution of myofiber types in thigh muscles of chickens.

The composition of myofiber types varies within thigh muscles of chickens. The present study was designed to determine whether or not myofiber types were distributed uniformly across the diameter of the thigh muscles of chickens. Cross sections from middle portions of muscles were used histochemically to examine differences in distribution and composition of myofiber types in the muscles. Myofibers that reacted moderately (M) or strongly (S) for myosin adenosine triphosphatase (ATPase) after preincubation at pH 4.3 were classified as type I. Type I myofibers reacted weakly (W), moderately (M), or strongly (S) for ATPase after preincubation at pH 10.6; these type I myofibers were subclassified into four types (ISW , ISM , ISS , and IMM ). Myofibers that reacted negatively for acid-stable ATPase and strongly for alkali-stable ATPase were classified into two types: type IIA, with strong NADH tetrazolium reductase (NADH-TR), and type IIB, with weak NADH-TR activity. The M. pubo-ischio-femoralis pars lateralis had numerous type IIA myofibers and very few type ISM myofibers, whereas the pars medialis had many type IMM myofibers and few type ISS and IIA myofibers. The type I group of myofibers did not exceed about 50% in the other muscles, which had one to three types of type ISW , ISM , and ISS myofibers. The Mm. femorotibiales had more type ISW , and ISM myofibers in the deep regions near the femur than in the superficial regions. The M. iliotibialis cranialis, M. iliofibularis, and M. flexor cruris medialis had more type ISW , ISM , or ISS myofibers in the medial regions than in the lateral regions. A few type ISW myofibers were scattered in the cranial part of M. iliotibialis and in the M. ambiens. The M. flexor cruris lateralis pars pelvica had type IIA and IIB myofibers exclusively. All the muscles had type IIA myofibers. Type IIB myofibers existed in the muscles except the M. puboischio-femoralis. Type IIA and IIB myofibers differed in proportion in different muscles and in their different regions. The type I group of myofibers was generally concentrated more in the deep regions near the femur and in the medial regions than in the superficial and lateral regions of the thigh muscles. The distribution of type IIA myofibers resembled that of type I group. Type IIB myofibers showed a distribution opposite to that of type I group and IIA myofibers. The spatial distribution of myofiber types within individual muscles can account for the various locomotory and postural requirements of the thigh.

Histochmical properties of myofiber types in thigh muscles of the chicken.

The presence of subtypes of dark and light myofibers distinguished by myosin adenosine triphosphatase (ATPase) reaction after alkaline preincubation was studied in thigh muscles of chickens. The dark myofibers were subclassified into type II and SS, and the light myofibers into type I, SM, and MM myofibers by the pH sensitivity of myosin ATPase. Type I myofibers reacted strongly for acid-stable ATPase and weakly for alkali-stable ATPase. Type II myofibers reacted negatively for acid-stable ATPase and strongly for alkali-stable ATPase. Type SM myofibers reacted strongly for acid-stable ATPase and modeiately for alkali-stable ATPase. Type SS myofibers reacted strongly and type MM myofibers reacted moderately for both acid-stable and alkali-stable ATPase. Type II myofibers were subdivided into type IIA with strong activity for NADH: Nitro BT oxidoreductase (NADHOX) and type IIB with weak NADHOX activity. Type IIB myofibers were strong in activity for glycerol-3-phosphate: menadione oxidoreductase (GPOX) whereas many type IIA myofibers were weak. Type IIA myofibers were stained weakly to moderately with 3-hydroxybutyrate: NAD+ oxidoreductase (HBOX) and type IIB myofibers were stained very weakly. Type I, SM, SS, and MM myofibers were stained strongly with NADHOX, weakly to moderately with HBOX, and weakly with GPOX.