Myosin Heavy Chain mRNA Expression Research Articles

Effects of Varying Intensity Resistance Exercise on Myogenic Regulatory Factors and Myosin Heavy Chain Genes

A single bout of resistance training is capable of activating the expression of many diverse groups of genes including myosin heavy chain (MHC) isoforms and the myogenic regulatory factors (MRF). However, the role of exercise intensity in regards to the effects on transcription and protein synthesis are not well denned. PURPOSE: The purpose of this study was to investigate the effects of resistance exercise intensity on MRF mRNA and protein and MHC mRNA isoforms. METHODS: Thirteen male participants (21.5 ± 2.9 yrs, 86.1 ± 19.5 kg, 69.7 ± 2.7in) completed two bouts [low-intensity/high volume (LI = 65% 1-RM) and high-intensity/ low volume (HI = 85% 1-RM)] of single-legged resistance exercise, each employing four sets of leg press and leg extension. Muscle biopsies were obtained from each leg immediately before exercise (PRE), and at 30 minutes (30MPST), 2 hours (2HRPST), and 6 hours (6HRPST) following each resistance exercise bout. Data were analyzed using RT-PCR for mRNA expression and ELISA for protein expression. Statistical analyses were performed by utilizing a 2 x 4 MANOVA (p ≤ 0.05). RESULTS: Results indicated that MHC I, IIb, and IIx all significantly increased in response to resistance exercise (p < 0.001). Furthermore, exercise intensity played a role with LI exercise stimulating a more positive response in MHC I (p < 0.001) and HI eliciting a greater response in MHC IIb and IIx (p < 0.001). In regards to the MRF, resistance exercise significantly increased the expression of Myogenin, Myf5, and MRF-4 mRNA (p < 0.05). Furthermore, LI exercise stimulated a more positive response in MyoD, Myogenin, and Myf5 mRNA (p < 0.05), as well as MyoD, Myogenin, and MRF-4 protein (p < 0.05). MRF-4 protein also significantly increased at 6HRPST over the baseline values (p < 0.05). Changes in MyoD and Myogenin mRNA were also found to be significantly correlated (p < 0.05) to changes in MHC I, IIa, IIb, and IIx mRNA. CONCLUSIONS: This study suggests that both MHC and MRF mRNA expression is quickly up-regulated in response to resistance exercise, but that LI and high volume may generate a more robust response.

Point:Counterpoint: Satellite cell addition is/is not obligatory for skeletal muscle hypertrophy

Myofiber size is dynamically regulated, increasing and decreasing depending on muscle use. Hypertrophy is defined by increases in myofiber cross-sectional area and mass as well as myofibrillar protein content. Myofibers contain many hundreds of nuclei, each of which has a nuclear domain. A nuclear

Effects of stretching stimulation with different rates on the expression of MyHC mRNA in mouse cultured myoblasts

In vivo studies have shown that changes in the characteristics of skeletal muscle fiber are determined by type of exercise or training. These earlier studies on mechanical stimulation, however, have all employed stimulation applied at a constant intensity, and no studies appear to have investigated change with variation of intensity of stimulation. In this study, we investigated the characteristics and differentiation of myoblasts stretched at different rates. Myoblasts were stimulated at 3 different rates, and the numbers of cells and nuclei on days 1, 3, and 5 were compared. The myosin heavy chain (MyHC) mRNA expression level was also compared. We investigated expression of MyHC-perinatal to determine speed of differentiation of myoblasts, and expression of MyHC-2b, 2d, and 2a to ascertain muscle cell characteristics. Counting cells and nuclei of myoblasts revealed clear promotion of differentiation with stretching. With rapid stretching, expression of MyHC-perinatal was high at first, but then showed a decrease. In terms of effect on muscle fiber characteristics, MyHC-2b, MyHC-2d, and MyHC-2a were high with rapid, medium, and slow stretching, respectively. This indicated that myoblast differentiation was promoted regardless of difference in stretching speed, with the myoblasts acquiring the muscle-fiber characteristics appropriate to each rate of stretching.

Highlights From The Literature

Highlights From The Literature

Mechanisms underlying myosin heavy chain expression during development of the rat diaphragm muscle

During early postnatal development in rat diaphragm muscle (Dia(m)), significant transitions in myosin heavy chain (MHC) isoform expression occur that are associated with fiber growth and increased MHC protein. At present, there is no direct information regarding the transcriptional regulation of MHC isoform expression during postnatal Dia(m) development. We hypothesized postnatal changes in MHC isoform mRNA expression are followed by concomitant changes in MHC protein expression. The Dia(m) was removed at postnatal days 0, 14, 28, and 84 (adult). MHC mRNA expression was determined by real-time RT-PCR. MHC protein expression was determined by SDS-PAGE. There was a significant effect of postnatal age on MHC isoform mRNA and protein expression. At birth, the MHC(Neo) isoform accounted for 28% of MHC mRNA and 54% of total MHC protein. By postnatal day 14, MHC(Neo) mRNA and protein increased significantly, and both decreased significantly by day 28, consistent with transcriptional control of the expression of this developmental isoform. By postnatal day 28, there were minimal changes in mRNA expression for MHC(Slow) and MHC(2X), yet protein expression increased significantly. MHC(2A) mRNA and protein expression did not change during this time. Thus changes in MHC protein expression did not follow (or parallel) changes in MHC mRNA for the adult MHC isoforms. The present findings indicate that changes in MHC expression in the developing rat Dia(m) are not driven solely by changes in mRNA expression. Knowledge of isoform-specific MHC mRNA expression only yields predictive information on MHC protein expression for the MHC(Neo) isoform.

Myosin Heavy Chain Isoforms in Equine Gluteus Medius Muscle: Comparison of mRNA and Protein Expression Profiles

The major structural protein in skeletal muscle, myosin heavy chain (MyHC), is primarily transcriptionally controlled. We compared the expression of MyHC isoforms on the mRNA and protein level in biopsies from the m. gluteus medius from adult untrained horses. In transverse sections, the majority of fibers showed qualitatively identical mRNA and protein expression patterns. However, coexpression of 2a and 2d/x MyHCs was substantially more common at the protein than at the mRNA level, suggesting a fine-tuning of these two genes in normal muscle not subjected to any training protocol. Because transverse sections give a limited sampling of mRNA expression in the case of uneven distribution of transcripts in a muscle fiber, we also analyzed longitudinal sections. We present, for the first time, evidence that expression of MyHC mRNA and protein was equal along the length of the fiber. Hence, mRNA expression is not regulated by differential expression of isoforms by separate myonuclei. It is concluded that the number of protein hybrid fibers in equine gluteus medius muscle is controlled by alteration of the transcription pattern uniformly along the fiber, rather than by simultaneous transcription of genes. The differences with the results in muscle of small animals and humans are discussed.

In vivo inhibition of nitric oxide synthase impairs upregulation of contractile protein mRNA in overloaded plantaris muscle

Inhibition of nitric oxide synthase (NOS) activity in vivo impedes hypertrophy in the overloaded rat plantaris. We investigated the mechanism for this effect by examining early events leading to muscle growth following 5 or 12 days of functional overload. Male Sprague-Dawley rats (approximately 350 g) were randomly divided into three treatment groups: control, N(G)-nitro-L-arginine methyl ester (L-NAME; 90 mg.kg(-1).day(-1)), and 1-(2-trifluoromethyl-phenyl)-imidazole (TRIM; 10 mg.kg(-1).day(-1)). Unilateral removal of synergists induced chronic overload (OL) of the right plantaris. Sham surgery performed on the left hindlimb served as a normally loaded control. L-NAME and TRIM treatments prevented OL-induced skeletal alpha-actin and type I (slow) myosin heavy chain mRNA expression at 5 days. Conversely, neither L-NAME nor TRIM affected hepatocyte growth factor or VEGF mRNA responses to OL at 5 days. However, OL induction of IGF-I and mechanogrowth factor mRNA was greater (P < 0.05) in the TRIM group compared with the controls. Furthermore, the phosphorylated-to-total p70 S6 kinase ratio was higher in OL muscle from NOS-inhibited groups, compared with control OL. At 12 days of OL, the cumulative proliferation of plantaris satellite cells was assessed by subcutaneous implantation of time release 5'-bromo-2'-deoxyuridine pellets during the OL-inducing surgeries. Although OL caused a fivefold increase in the number of mitotically active (5'-bromo-2'-deoxyuridine positive) sublaminar nuclei, this was unaffected by concurrent NOS inhibition. Therefore, NOS activity may provide negative feedback control of IGF-I/p70 S6 kinase signaling during muscle growth. Moreover, NOS activity may be involved in transcriptional regulation of skeletal alpha-actin and type I (slow) myosin heavy chain during functional overload.

Myosin heavy chain mRNA expression correlates higher with muscle protein accretion than growth in Atlantic salmon, Salmo salar

In order to link growth and protein accretion in Atlantic salmon the mRNA expression of muscle myosin heavy chain ( MyHC) was analysed. MyHC gene is expressed throughout muscle development and is consistent with the hypertropic growth in fish. Total RNA was isolated from white muscle tissues ( N = 32) from salmon fed a fish meal based diet with three levels of solubilised protein incorporated as fish protein hydrolysate (FPH) control (0 g kg − 1 ), medium (180 g kg − 1 ) and high (300 g kg − 1 ) FPH inclusion. The salmon were PIT-tagged and fed the experimental diets for a period of 68 days. A high level of dietary FPH resulted in significant up-regulation of MyHC by a factor of 2.4 compared to fish fed the medium FPH inclusion. The fish with highest MyHC expression also contained significant higher levels of muscle protein. MyHC expression correlated higher with protein accretion than individual specific growth rate (SGR) in salmon. These observations indicate that MyHC mRNA expression can be a useful marker for understanding of growth and protein accretion in Atlantic salmon.

Changes in detrusor smooth muscle myosin heavy chain mRNA expression following spinal cord injury in the mouse.

Smooth muscle myosin heavy chain (SMMHC) isoform composition has been shown to be developmentally regulated and to be associated with functional changes in smooth muscle activity. In this study, we sought to determine expression patterns of SMMHC isoforms in a murine model of spinal cord injury (SCI) and to compare these expression patterns to neurologic, cytometric, and morphometric findings. Baseline cystometry was performed on adult, female mice followed by either thoracic spinal cord transection (SCI) or sham operation (Sham). At 1, 3, or 6 weeks postoperatively neurologic evaluation and cystometry were performed, bladders were harvested, and expression patterns of SMMHC isoforms (SM1 vs. SM2 and SMA vs. SMB) were assessed by RT-PCR. Morphometrics utilizing computer-assisted color image analysis was also performed on all bladders. There was a significant increase in bladder weight and capacity 1 week following SCI which normalized over time, however, morphometric analysis did not reveal an alteration in tissue composition amongst the three groups. One week following SCI, SM1 was predominantly expressed over SM2 and began to normalize at 3 weeks. This coincided with the emergence of reflex voiding and detrusor overactivity. SMA was expressed following SCI only, and the number of bladders found to express SMA decreased with increasing duration since SCI. Smooth muscle myosin heavy chain mRNA expression patterns appear to be affected by SCI. We believe the induction of SMA may be a factor in altered bladder function following injury.

Cloning and sequencing of myosin heavy chain isoform cDNAs in golden-mantled ground squirrels: effects of hibernation on mRNA expression.

The golden-mantled ground squirrel is a small rodent hibernator that demonstrates unusual myosin heavy chain (MHC) isoform plasticity during several months of torpor, punctuated by bouts of rewarming and shivering thermogenesis. We measured MHC mRNA levels to determine whether pretranslational control mechanisms were responsible for differences in MHC2x protein expression, as we previously observed between active and hibernating ground squirrels. We first cloned cDNA using the 3' rapid amplification of cDNA ends (3' RACE) technique and identified three sequences corresponding to MHC1, MHC2x, and MHC2b. A DNA control fragment was developed to be used in conjunction with a coupled RT-PCR reaction to simultaneously measure MHC mRNA levels for each isoform in the skeletal muscle of ground squirrels. MHC mRNA and protein expression were strongly correlated, and type IIx and IIb mRNA levels were significantly different between active and hibernating ground squirrels. Pretranslational control of MHC protein is apparently an important process during hibernation, although the exact stimulus is not known. The techniques presented can be used to obtain MHC cDNA sequences and to measure mRNA expression in many vertebrate groups.

Age and Exercise Effects on Myosin Heavy Chain Isoform mRNA Expression in Human Muscle

0389 Aging is associated with changes in the contractile and metabolic properties of human skeletal muscle. Among these changes is a shift in fiber type towards a predominance of Type I fibers. It was reported that mRNA levels of myosin heavy chain (MHC) isoforms, the determinant of contractile phenotype, are altered with age in a manner consistent with fiber type results, although two studies found no changes in MHC mRNA expression. Further, a predominance of Type I fibers has been observed in highly aerobically-trained athletes but the effect of aerobic training on MHC mRNA expression and the potential interaction with age remains to be determined. PURPOSE: To determine the effect of aging and a moderate intensity aerobic exercise training program on mRNA levels of MHC isoforms in human muscle. METHODS: Previously sedentary, healthy men and women aged 22–88 y were studied before and after 16 weeks of bicycle training (30–45 min at 70–80% peak heart rate, 3–4 days/week, N = 62) or control (stretching, N = 37) activity. Muscle biopsies of the vastus lateralis were analyzed for MHC mRNA isoforms I, IIA and IIX using real-time PCR. RESULTS: At baseline MHC-I mRNA was unchanged with age, while levels of IIA (r = −0.45) and IIX (r = −0.34) declined by 14% and 10% per decade, respectively (p < 0.001). After training, the mRNA levels of MHC-I and IIA increased 61 and 99%, respectively, while IIX decreased 50% (all p < 0.001). The increase in MHC-I mRNA was positively associated with age (r = 0.30, p < 0.01). There was no interaction between age and exercise responses for MHC-IIA or IIX, and MHC mRNA did not change in the control group. CONCLUSION: This study demonstrates that MHC isoform expression changes with aging consistent with reported changes in fiber type. Despite baseline differences, older muscle retains the ability to alter expression of MHC isoforms at the mRNA level in response to aerobic training. This agrees with previous data from this study demonstrating that training-induced enhancement of mitochondrial genes and functions are similar among younger and older people. Thus, skeletal muscle plasticity, at least at the transcript level, remains intact in older people. Future work will determine if training-induced changes in MHC mRNA are also reflected at the protein level. Supported by NIH AG-RO109531, MO1-RR00585 and Mayo Foundation.

Transcriptional Control of Myosin Heavy Chain IIB Gene Expression in Normal and Overloaded Rat Plantaris

1038 The rat plantaris (P) muscle predominantly expresses fast-contracting isoforms of myosin heavy chain (MHC) proteins. During hypertrophy induced by chronic mechanical overload, the expression of the fastest type IIb MHC isoform decreases as the muscle shifts phenotype toward expression of slower-contracting MHC isoforms. PURPOSE: To determine the transcriptional control elements that reside downstream of the transcription start site (tss) and regulate MHC IIb expression in normal (Con) and functional overload (FO) rat P. METHODS: Female SD rats (N = 8/group) underwent 7 d FO of the left P. MHC IIb pre-mRNA and mRNA expression were measured by RT-PCR. The transcriptional activities of five MHC IIb promoters were tested by direct gene transfer into the P. Each promoter fragment began −1.431 kb upstream of the tss, and extended into either exon 1 (+0.013 kb), intron 2 (+1.008, +1.568, +2.054 kb), or exon 3 (+2.814 kb). Promoter activity was measured by a firefly luciferase (fLuc) reporter assay 7 d after bilateral intramuscular injection of plasmid constructs containing the fLuc gene controlled by the inserted IIb promoter fragment. To control for plasmid uptake efficiency, the fLuc reporter activity was corrected to the renilla luciferase activity of a co-injected alpha-actin promoter-reporter plasmid. RESULTS: After 7 d FO, the wet weight of the P increased ∼31% (P < 0.05). IIb pre-mRNA and mRNA each decreased by ∼60% after FO (P < 0.05). Activities of the +2.054 and +2.814 kb promoters were similar, and decreased ∼50% after FO (P < 0.05). The three promoters that extended downstream between +0.013 and +1.568 kb were 80–99% less active (P < 0.05) than the +2.814 kb promoter, and did not decrease after FO. CONCLUSION: MHC IIb expression in Con and FO P is controlled by transacting factor(s) located downstream of the tss, and specifically within the intron 2 region between +1.568 and +2.054 kb. Further studies are aimed at identifying the trans-acting factors that bind this region of DNA to control transcription of the IIb gene in the normal rat P and its down-regulation during hypertrophy. Supported by NIH grant AR30346

Systematic analysis of the TGF‐β/Smad signalling pathway in the rhabdomyosarcoma cell line RD

Transforming growth factor-beta (TGF-beta) is a multifunctional regulator of cell growth and differentiation, whose actions are highly cell type specific. To study the role of the TGF-beta1 autocrine loop in regulating growth and myogenic differentiation in the human rhabdomyosarcoma cell line, RD, an attempt was made to establish a framework for the expression of several components of TGF-beta1/Smad signalling pathway at the mRNA and protein levels by reverse transcription-polymerase chain reaction (RT-PCR) and Western blot analysis in RD cells compared with the normal myoblasts. Higher exogenous concentration of TGF-beta1 was necessary to reach a growth-inhibition effect, whereas TGF-beta1 downregulated the expression of myosin heavy-chain mRNA at lower concentrations than that was required for growth inhibition. Treatment with TGF-beta1 significantly decreased the number of sarcomeric actin and myosin-expressing cells. In this study, we have shown that RD cells displayed higher expression of TbetaRI, TbetaRII, Smad2 and Smad4 at both the mRNA and protein levels than myoblasts. Smad3 and Smad7 mRNA were expressed at higher level in RD cells than in myoblasts. The staining patterns of TbetaR and Smads suggest that they may transduce different TGF-beta1 signalling in RD cells than in myoblasts. TGF-beta1 signalling induced a rapid relocation of Smad2 to the nucleus; in contrast, Smad4 remained localized to the cytoplasm unless it was coexpressed with Smad2. These studies suggest that signalling from the cell surface to the nucleus through Smad proteins is a required component of TGF-beta1-induced cell response in RD cells. The RD cell line is a suitable model to study the TGF-beta autocrine loop involved in growth and differentiation of RMS.

Denervation-induced changes in myosin heavy chain expression in the rat diaphragm muscle.

Unilateral denervation (Dnv) of the rat diaphragm muscle (Diam) markedly alters expression of myosin heavy chain (MHC) isoforms. After 2 wk of Diam Dnv, MHC content per half-sarcomere decreases in fibers expressing MHC(2X) and MHC(2B). We hypothesized that changes in MHC protein expression parallel changes in MHC mRNA expression. Relative MHC isoform mRNA levels were determined by Northern analysis after 1, 3, 7, and 14 days of Dnv of the rat Diam. MHC protein expression was determined by SDS-PAGE. Changes in MHC isoform protein and mRNA expression were not concurrent. Expression of MHC(Slow) and MHC(2X) mRNA isoforms decreased dramatically by 3 days of Dnv, whereas that of MHC(2A) and MHC(2B) did not change. Expression of all MHC protein isoforms decreased by 3 days of Dnv. We observed a differential effect of rat Diam Dnv on MHC isoform protein and mRNA expression. The time course of the changes in MHC isoform mRNA and protein expression suggests a predominant effect of altered protein turnover rates on MHC protein expression instead of altered transcription after Dnv.

Myosin heavy chain expression in cranial, pectoral fin, and tail muscle regions of zebrafish embryos.

To investigate whether different myosin heavy chain (MHC) isoforms may constitute myofibrils in the trunk and tail musculature and if their respective expression may be regulated by spadetail (spt) and no tail (brachyury), we identified and characterized mRNA expression patterns of an embryonic- and tail muscle-specific MHC gene (named myhz2) during zebrafish development in wild type, spt, and ntl mutant embryos. The identified myhz2 MHC gene encodes a polypeptide containing 1,935 amino acids. Deduced amino acid comparisons showed that myhz2 MHC shared 92.6% sequence identity with that of carp fast skeletal MHC. Temporal and spatial myhz2 MHC mRNA expression patterns were analyzed by quantitative RT-PCR and whole-mount in situ hybridization using primer pairs and probes designed from the 3'-untranslated region (UTR). Temporally myhz2 MHC mRNA appears in pharyngula embryos and peaks in protruding-mouth larvae. The expression level decreased in 7-day-old hatching larvae, and mRNA expression was not detectable in adult fish. Spatially in pharyngula embryos, mRNA was localized only in the tail somite region, while in long-pec embryos, transcripts were also expressed in the two cranial muscle elements of the adductor mandibulae and medial rectus, as well as in pectoral fin muscles and the tail muscle region. Myhz2 MHC mRNA was expressed in most cranial muscle elements, pectoral fin muscles, and the tail muscle region of 3-day-old hatching larvae. In contrast, no expression of myhz2 MHC mRNA could be observed in spt prim-15 mutant embryos. In spt long-pec mutant embryos, transcripts were expressed in two cranial muscle elements and the tail muscle region, but not in pectoral fin muscles, while only trace amounts of myhz2 MHC mRNA were expressed in the remaining tail muscle region of 38 hpf and long-pec ntl mutant embryos.

Change of chloride ion channel conductance is an early event of slow-to-fast fibre type transition during unloading-induced muscle disuse.

Disuse of postural slow-twitch muscles, as it occurs in hypogravity, induces a slow-to-fast myofibre type transition. Nothing is known about the effects of weightlessness on the resting membrane chloride conductance (gCl), which controls sarcolemma excitability and influences fibre type transition during development and adult life. Using the current-clamp method, we observed that rat hindlimb unloading (HU) for 1-3 weeks increased gCl in fibres of the slow-twitch soleus (Sol) muscle toward values found in fast muscle. Northern blot analysis suggested that this effect resulted from an increased ClC-1 chloride channel mRNA level. In the meantime, a 4-fold increase in fibres expressing fast isoforms of the myosin heavy chain (MHC) was observed by immunostaining of muscle sections. Also, Sol muscle function evolved toward a fast phenotype during HU, as demonstrated by the positive shift of the threshold potential for contraction. After 3-days HU, Sol muscle immunostaining and RT-PCR experiments revealed no change in MHC protein and mRNA expression, whereas the gCl was already maximally increased, due to a pharmacologically probed, increased activity of ClC-1 channels. Thus the increase in gCl is an early event in Sol muscle experiencing unloading, suggesting that gCl may play a role in muscle adaptation to modified use. Pharmacological modulation of ClC-1 channels may help to prevent disuse-induced muscle impairment.

Changes in myosin heavy chain and its localization in rat heart in association with hypobaric hypoxia-induced pulmonary hypertension.

Experimental pulmonary hypertension induced in a hypobaric hypoxic environment (HHE) is characterized by structural remodelling of the heart. In rat cardiac ventricles, pressure and volume overload are well known to be associated with changes in cardiac myosin heavy chain (MHC) isoforms. To study the effects of HHE on the MHC profile in the ventricles, 83 male Wistar rats were housed in a chamber at the equivalent of 5500 m altitude for 1-8 weeks. Pulmonary arterial pressure, right ventricular free wall (RVFW) weight, the ratio of RVFW weight over body weight (BW), the ratio of left ventricular free wall (LVFW) weight over BW, and myocyte diameter in both ventricles showed significant increases after 1 week, 2 weeks, 1 week, 6 weeks, and 4 weeks of HHE, respectively. Semi-quantitative reverse transcriptase-polymerase chain reaction revealed that beta-MHC mRNA expression was increased significantly in both ventricles at 6 and 8 weeks of HHE, whereas alpha-MHC mRNA expression was decreased significantly at 6 and 8 weeks of HHE in the right ventricle (RV) and at 6 weeks of HHE in the left ventricle (LV). The percentage of myosin containing the beta-MHC isoform was increased significantly at 4-8 weeks of HHE in RV and at 6 weeks of HHE in LV. In situ hybridization showed that the area of strong staining for beta-MHC mRNA was increased in both ventricles at 8 weeks of HHE, and showed a decrease from RVFW to cardiac septum, and from cardiac septum to LVFW. These results suggest that HHE has a significant effect on the expression of both MHC mRNA and protein in the heart, particularly in RV. These changes may reflect a role for cardiac MHC in the response to pulmonary hypertension in HHE.

Differential expression of myosin heavy-chain mRNA in muscles of mastication during functional advancement of the mandible in pigs

Surgical and orthodontic treatment of retrognathia aims to improve orofacial function by adaptation and training of muscle capacity, which is connected with a change in muscle fibre-type proportions. The aim here was to analyse the proportion of myosin-heavy chain (MyHC) gene expression in type I (slow twitch/ST) and type IIb (fast twitch/FT) fibres during sagittal advancement of the mandible by reverse transcriptase-polymerase chain reaction (RT-PCR). The experiments were carried out on 10-week-old pigs (six test animals, six controls) over a 28-day period. Six pigs were fitted with acrylic bite blocks for sagittal advancement of the mandible. Tissue was taken from seven different regions of the masseter, temporal, medial pterygoid, and geniohyoid muscles. The 84 samples were used for histological fibre differentiation with ATPase staining and for isolation of total RNA. To measure the two MyHC isoforms, RT-PCR (in a single tube reaction with MyHC I, MyHC IIb, and GAPDH primers) was used. A significant increase was registered in the percentage of ST fibres and in mRNA from MyHC I in the anterior region of the masseter and in the posterior region of the temporal muscle of the treated animals. The proportion of ST fibres to FT fibres was increased by up to 12% after functional advancement of the mandible. The histological findings corresponded with the data for fibre mRNA generated by RT-PCR.

Alpha-blockade downregulates myosin heavy chain gene expression in human benign prostatic hyperplasia

Objectives. Smooth muscle (SM), a major component of prostate stroma, plays an important role in the pathogenesis of benign prostatic hyperplasia. In many muscle systems, steroid hormones and alpha 1-adrenergic neurotransmitters tightly regulate expression of contractile proteins. In this study, SM content and the expression of myosin heavy chain (MHC) in tissues from patients with benign prostatic hyperplasia treated with androgen ablation or alpha-blockade were compared with untreated controls. Methods. Prostatic periurethral tissue specimens from patients receiving luteinizing hormone-releasing hormone analogues (n = 12), alpha-blocking agents (n = 12), and no treatment (n = 13) were examined. The samples were analyzed for SM MHC mRNA expression using competitive reverse transcription-polymerase chain reaction. SM content was measured by morphometric analysis of trichrome-stained sections. Results. Stromal SM constituted 45.4% ± 8.6%, 48.1% ± 18.4%, and 45.9% ± 10.8% of the total tissue in androgen ablated, alpha-blocked, and untreated tissues, respectively. No significant difference was observed among these three groups ( P = 0.84, analysis of variance). However, SM MHC mRNA expression was markedly decreased in the alpha-blockade group (0.15 ± 0.02 attomole/mg tissue) compared with the androgen-ablated (0.58 ± 0.15 attomole/mg tissue) or control (0.44 ± 0.10 attomole/mg tissue) groups. The relationship between SM content and expression of SM MHC significantly differed among the groups ( P = 0.02, analysis of variance). Conclusions. Androgen ablation and alpha-blockade do not appear to alter the histologic characteristics of prostate stroma in men with symptomatic benign prostatic hyperplasia. However, contractile protein gene expression in stromal SM cells is significantly altered after alpha-blockade. These data suggest that, in addition to the simple relaxation of muscle tone, alpha-blocking agents may affect the phenotypic expression of contractile proteins in prostate SM cells.

Plasticity of myosin heavy chain mRNA expression with temperature acclimation is gradually acquired during ontogeny in the common carp

Plasticity of myosin heavy chain mRNA expression with temperature acclimation is gradually acquired during ontogeny in the common carp