Ultrastructure of striated muscle fibers in the middle third of the human esophagus

Striated muscle fibers and their spatial relationship to smooth muscle cells have been studied in the middle third of human esophagus. Biopsies were obtained from 3 patients during surgery. In both the circular and longitudinal layers, the muscle coat of this transition zone was composed of fascicles of uniform dimension (100-200 microns of diameter); some of these bundles were made up of striated muscle fibers, others were pure bundles of smooth muscle cells and some were of the mixed type. Striated muscle fibers represented three different types, which were considered as intermediate, with certain structural features characteristic of the fast fiber type. Of these, the most frequently-found fibers were most similar to the fast fiber type. Satellite cells were numerous; in mixed fascicles they were gradually replaced by smooth muscle cells. The gap between striated muscle fiber and smooth muscle cells was more than 200 nm wide. It contained the respective basal laminae and a delicate layer of amorphous connective tissue. No specialized junctions were formed between consecutive striated muscle fibers, or between striated muscle fibers and smooth muscle cells. Interstitial cells of Cajal were never situated as close to striated muscle fibers as to smooth muscle cells.     

Heterogeneous distribution of isoactins in cultured vascular smooth muscle cells does not reflect segregation of contractile and cytoskeletal domains.

We have previously demonstrated that alpha-smooth muscle (alpha-SM) actin is predominantly distributed in the central region and beta-non-muscle (beta-NM) actin in the periphery of cultured rabbit aortic smooth muscle cells (SMCs). To determine whether this reflects a special form of segregation of contractile and cytoskeletal components in SMCs, this study systematically investigated the distribution relationship of structural proteins using high-resolution confocal laser scanning fluorescent microscopy. Not only isoactins but also smooth muscle myosin heavy chain, alpha-actinin, vinculin, and vimentin were heterogeneously distributed in the cultured SMCs. The predominant distribution of beta-NM actin in the cell periphery was associated with densely distributed vinculin plaques and disrupted or striated myosin and alpha-actinin aggregates, which may reflect a process of stress fiber assembly during cell spreading and focal adhesion formation. The high-level labeling of alpha-SM actin in the central portion of stress fibers was related to continuous myosin and punctate alpha-actinin distribution, which may represent the maturation of the fibrillar structures. The findings also suggest that the stress fibers, in which actin and myosin filaments organize into sarcomere-like units with alpha-actinin-rich dense bodies analogous to Z-lines, are the contractile structures of cultured SMCs that link to the network of vimentin-containing intermediate filaments through the dense bodies and dense plaques.     

Cellular distribution of smooth muscle actins during mammalian embryogenesis: expression of the alpha-vascular but not the gamma- enteric isoform in differentiating striated myocytes

The cellular distribution of the alpha-vascular and gamma-enteric smooth muscle actin isoforms was analyzed in rat embryos from gestational day (gd) 8 through the first neonatal week by in situ antigen localization using isoactin specific monoclonal antibodies. The alpha-vascular actin isoform was first detected on gd 10 in discrete cells lining the embryonic vasculature. By gd 14, this isoform was also present in the inner layers of mesenchymal cells condensing around the developing airways and gut. The gamma-enteric actin, however, was not detected until gd 15 when cells surrounding the developing aorta, airways, and gut labeled with the gamma-enteric-specific probe. There was continued expression of these two actin isoforms in regions of developing smooth muscle through the remainder of gestation and first neonatal week at which time their distribution coincided with that found in the adult. In addition to developing smooth muscle, the alpha- vascular actin isoform was expressed in differentiating striated muscle cells. On gd 10, there was intense labeling with the alpha-vascular specific probe in developing myocardiocytes and, within 24 h, in somitic myotomal cells. Although significant levels of this smooth muscle actin were present in striated myocytes through gd 17, by the end of the first postnatal week, alpha-vascular actin was no longer detectable in either cardiac or skeletal muscle. Thus, the normal developmental sequence of striated muscle cells includes the transient expression of the alpha-vascular smooth muscle actin isoform. In contrast, the gamma-enteric smooth muscle actin was not detected at any time in embryonic striated muscle. The differential timing of appearance and distribution of these two smooth muscle isoforms indicates that their expression is independently regulated during development.     

A monoclonal antibody against alpha-smooth muscle actin: a new probe for smooth muscle differentiation

A monoclonal antibody (anti-alpha sm-1) recognizing exclusively alpha- smooth muscle actin was selected and characterized after immunization of BALB/c mice with the NH2-terminal synthetic decapeptide of alpha- smooth muscle actin coupled to keyhole limpet hemocyanin. Anti-alpha sm- 1 helped in distinguishing smooth muscle cells from fibroblasts in mixed cultures such as rat dermal fibroblasts and chicken embryo fibroblasts. In the aortic media, it recognized a hitherto unknown population of cells negative for alpha-smooth muscle actin and for desmin. In 5-d-old rats, this population is about half of the medial cells and becomes only 8 +/- 5% in 6-wk-old animals. In cultures of rat aortic media SMCs, there is a progressive increase of this cell population together with a progressive decrease in the number of alpha- smooth muscle actin-containing stress fibers per cell. Double immunofluorescent studies carried out with anti-alpha sm-1 and anti- desmin antibodies in several organs revealed a heterogeneity of stromal cells. Desmin-negative, alpha-smooth muscle actin-positive cells were found in the rat intestinal muscularis mucosae and in the dermis around hair follicles. Moreover, desmin-positive, alpha-smooth muscle actin- negative cells were identified in the intestinal submucosa, rat testis interstitium, and uterine stroma. alpha-Smooth muscle actin was also found in myoepithelial cells of mammary and salivary glands, which are known to express cytokeratins. Finally, alpha-smooth muscle actin is present in stromal cells of mammary carcinomas, previously considered fibroblastic in nature. Thus, anti-alpha sm-1 antibody appears to be a powerful probe in the study of smooth muscle differentiation in normal and pathological conditions.     

Action of general and alpha-smooth muscle-specific actin antibody microinjection on stress fibers of cultured smooth muscle cells

Arterial smooth muscle cells express alpha- and gamma-smooth muscle, as well as beta- and gamma-cytoplasmic actins. Two actin antibodies, one recognizing smooth muscle and cytoplasmic actin isoforms, the other recognizing specifically alpha-smooth muscle actin, were microinjected into cultured aortic smooth muscle cells. The effect of these antibodies on stress fiber organization was examined by staining with rhodamine-labeled phalloidin and by immunofluorescence with the same antibodies. Microinjection of the general actin antibody abolished most of the stress fiber staining with all reagents, but did not significantly affect the shape of the injected cells. This suggests that stress fiber integrity is not absolutely necessary for the maintenance of cell shape within the time of observation. Microinjection of the specific alpha-smooth muscle antibody abolished to various extents the staining of stress fibers with this antibody, but left practically intact their staining with rhodamine-labeled phalloidin and with the general actin antibody. This suggests that the incorporation of alpha-smooth muscle actin is not absolutely necessary for the maintenance of stress fiber integrity in cultured smooth muscle cells.     

Acetylcholine receptor degradation measured by density labeling: effects of cholinergic ligands and evidence against recycling.

The methodology of density labeling of proteins by biosynthetic incorporation of 2H, 13C, 15N-amino acids into newly synthesized polypeptide chains allows the direct measurement of the turnover rate of the acetylcholine receptor in cultured chick skeletal muscle. In this study, receptors synthesized in medium containing 2H, 13C, 15N-amino acids were resolved from 1H, 12C, 14N-receptors by velocity sedimentation in sucrose-deuterium oxide gradients, and their proportions were determined by computer analysis of the gradient profiles. The kinetics of turnover of acetylcholine receptors are identical for developing chick muscle fibers grown in medium containing 2H, 13C, 15N-amino acids or 1H, 12C, 14N-amino acids, and the high degree of substitution of normal aminoacyl residues by 2H, 13C, 15N-residues does not affect the turnover rate of the denser receptor. Comparison of the turnover rates in continuous and pulse-labeling experiments gave independent confirmation of these results. The application of a potent, essentially irreversible blocking agent, alpha-bungarotoxin, increases the median lifetime of receptors from 17 hr for the native unbound receptor to 22 hr for the alpha-bungarotoxin-receptor complex. As predicted, the total number of alpha-bungarotoxin binding sites increased in the continued presence of alpha-bungarotoxin due to extension of receptor lifetime. To determine whether other cholinergic agents affect the turnover rate of the receptor, measurements were performed on cultures grown in the presence of 10(-4) M d-tubocurare or 10(-4) M carbachol, a reversible antagonist and a reversible agonist, respectively, of the nicotinic acetylcholine receptor. The receptor degradation rates of the drug-treated cells were identical to control values. The total number of alpha-bungarotoxin binding sites was reduced by 30% in the presence of carbachol, indicating that this agent affects the rate of synthesis of the acetylcholine receptor. Data formerly interpreted as suggesting a cycling of receptor-containing plasma membrane out of and back into the sarcolemma are now understood to reflect the alteration in receptor lifetime upon complexing with alpha-bungarotoxin. The intracellular "hidden" receptor sites were found to remain inside the myotubes and thus do not signify an intracellular pool of recycling plasma membrane.     

Biochemical evidence for interaction between smoothelin and filamentous actin

The two major isoforms of smoothelin (A and B) contain a calponin homology (CH) domain, colocalize with alpha-smooth muscle actin (alpha-SMA) in stress fibers and are only expressed in contractile smooth muscle cells (SMCs). Based on these findings, we hypothesized that smoothelins are involved in smooth muscle cell contraction, presumably via interaction with actin. The interaction between smoothelins and three different actin isoforms (alpha- and gamma-smooth muscle and alpha-skeletal actin [alpha-SKA]) was investigated using several in vitro assays. Smoothelin-B co-immunoprecipitated with alpha-smooth muscle actin from pig aorta extracts. In rat embryonic fibroblasts, transfected smoothelins-A and -B associated with stress fibers. In vitro dot blot assays, in which immobilized actin was overlaid with radio-labeled smoothelin, showed binding of smoothelin-A to actin filaments, but not to monomeric G-actin. A truncated smoothelin, containing the calponin homology domain, associated with stress fibers when transfected and bound to actin filaments in overlay, but to a lesser extent. ELISA results showed that the binding of smoothelin to actin has no significant isoform specificity. Our results indicate an interaction between smoothelin and actin filaments. Moreover, the calponin homology domain and its surrounding sequences appear to be sufficient to accomplish this interaction, although the presence of other domains is apparently necessary to facilitate and/or strengthen the binding to actin.     

Regulation of alpha-smooth muscle actin gene expression in myofibroblast differentiation from rat lung fibroblasts

Myofibroblasts express alpha-smooth muscle actin and have a phenotype intermediate between fibroblasts and smooth muscle cells. Their emergence can be induced by cytokines such as transforming growth factor beta; but the regulatory mechanism for induction of alpha-smooth muscle actin gene expression in myofibroblast differentiation has not been determined. To examine this mechanism at the level of the alpha-smooth muscle actin promoter, rat lung fibroblasts were transfected with varying lengths of the alpha-smooth muscle actin promoter linked to the chloramphenicol acetyl transferase reporter gene and treated with transforming growth factor beta1. The results show that the shortest inducible promoter was 150 base pairs long, suggesting the presence in this region of cis-elements of potential importance in transforming growth factor beta1 induced myofibroblast differentiation. Transfection of "decoy" oligonucleotides corresponding to sequences for four suspected regulatory factors demonstrated that only the transforming growth factor beta control element is involved in the regulation of transforming growth factor beta1-induced alpha-smooth muscle actin expression in myofibroblast differentiation. Consistent with this conclusion is the finding that a mutation in the transforming growth factor beta control element caused a significant reduction in promoter activity. These observations taken together show that alpha-smooth muscle actin promoter regulation during myofibroblast differentiation is uniquely different from that in smooth muscle cells and other cell lines. Since myofibroblasts play a key role in wound contraction and synthesis of extracellular matrix, clarification of this differentiation mechanism should provide new insight into fibrogenesis and suggest future novel strategies for modulation of wound healing and controlling fibrosis.     

Preservation of the myofibroblastic phenotype of human papilloma virus 16 E6/E7 immortalized human bone marrow cells using the lineage limited alpha-smooth muscle actin promoter.

The in vitro study of mammalian hematopoiesis is hindered by the lack of immortalized human stromal cell lines that support hematopoiesis. We have immortalized human stromal vascular smooth muscle cells characterized by the expression of the alpha-smooth muscle (alpha-SM) actin. This marker is usually down-regulated as a result of oncogenic transformation. To correct this dedifferentiation, we placed the expression of human papilloma virus 16 E6/E7 oncogenes under the control of the tissue-specific alpha-SM actin promoter. The immortalization event is rare and requires polyclonal culture, but the corresponding established line retains alpha-SM actin expression. Moreover, when compared with other lines derived from the same cells from vectors made with the same oncogenes but driven by either an internal SV40 promoter or the viral long terminal repeat, this line is less transformed as shown by anchorage-independent growth assay. We show therefore that the use of a physiological promoter allows the production of human cell lines with a conserved phenotype.     

Remodeling of small intestine with metamorphosis in tadpoles (Rhacophorus owstoni)]

The alimentary canal of typical anurans is remodeled during metamorphosis. We examined the tissue structure of small intestine in tadpoles of the treefrog, Rhacophorus owstoni. The cytoskeletal components of the smooth muscle cells were identified by immunohistochemically with alpha-smooth muscle actin antibody. Before metamorphosis, the smooth muscle fibers are arranged in two layers that are circular and longitudinal in orientation, and are stained markedly brown. During metamorphosis, there was concomitant increase in the number of muscle fibers per unit length of tract in both layers. By the end of the metamorphosis, the alimentary tract are transformed morphologically from the larval to adult form, and the smooth muscle cells are rapidly increasing in number. This study suggests that the structural change of musculature are formed in rearrangements of smooth muscle cells, but no proliferation of these cells with the metamorphosis.     

Postnatal changes in vagal control of esophageal muscle contractions in rats

AimsReplacement of smooth muscles by striated muscles occurs in the esophagus during the early postnatal period. The aim of this study was to clarify postnatal changes in vagal control of esophageal muscle contractions in rats.Main methodsAn isolated segment of the neonatal rat esophagus was placed in an organ bath and the contractile responses were recorded using a force transducer.Key findingsElectrical stimulation of the vagus trunk evoked a biphasic contractile response in the neonatal esophageal segment. The first and second components of the contractions were inhibited by α-bungarotoxin and atropine, respectively. Ganglion blockers, hexamethonium and mecamylamine, did not affect vagally mediated contractions. The first component gradually enlarged with age in days, whereas the second component declined during the first week after birth. Application of d-tubocurarine or acetylcholine caused an apparent contraction in the esophageal striated muscle at postnatal day 0, but responses to these drugs were not observed at 1 week after birth. The neonatal esophagus expressed the γ-subunit of nicotinic acetylcholine receptors. In contrast, the ε-subunit was dominantly expressed in the adult esophagus.SignificanceThe vagus nerves directly innervate both the esophageal striated muscles and smooth muscles in the early neonatal period. During the process of muscle rearrangement, the property of the striated muscles is altered substantially. The specific features of striated muscles in the neonatal rat esophagus might compensate for immature formation of neuromuscular junctions. Unsuccessful conversion of the striated muscle property during postnatal muscle rearrangement would be related to disorders of esophageal motility.     

Anatomical and neurochemical features of the extrinsic and intrinsic innervation of the striated muscle in the porcine esophagus: evidence for regional and species differences

Studies of the intrinsic and extrinsic innervation patterns of esophageal motor endplates (MEPs) are mainly confined to small rodents. Therefore, an immunocytochemical, denervation and tracing study was conducted on the pig, an experimental model in which the distribution of the striated esophageal muscle portion more closely resembles the human situation. The purpose of this study was to analyze the origin and neurochemical content of the nerve fibers participating in the myoneural synapse. Fifteen 6-week-old domestic pigs were studied by immunohistochemistry combined with alpha-bungarotoxin labeling to define the co-innervation patterns of nitrergic and peptidergic nerve terminals in MEPs. Some animals were subjected to unilateral infra- or supranodose vagotomy to determine the origin of the nerve terminals in MEPs. Special attention was paid to the interregional differences in terms of co-innervation rates, and these findings were compared with literature data on small mammals. Double stainings revealed that most of the nNOS-immunoreactive (ir) terminals in MEPs co-stained for VIP, GAL and NPY, but not for PACAP and L-ENK. PACAP- and L-ENK-ir terminals were coarser than nNOS-ir terminals, and largely co-localized VAChT. High percentages of MEPs at the cervical level were contacted by PACAP- (approximately 94%) and L-ENK-ir (approximately 78%) terminals, but the proportion of both decreased in the rostrocaudal direction. Vagotomy significantly reduced their presence in MEPs at the thoracic and abdominal levels, while nNOS-ir terminals observed in approximately 30% of the MEPs were unaffected by vagotomy. Immunostainings on brainstem cryosections after retrograde tracing from the cervical esophagus showed that a large number of FB-positive cells in the nucleus ambiguus were PACAP-ir (approximately 72%). C-kit-positive interstitial cells of Cajal were seen adjacent to the striated muscle fibers, apparently without direct relationship to MEPs. Similar to mouse esophagus, intrinsic nitrergic fibers were found to run close to, or even spiral around, these interstitial cells, an association that might point to a role as specialized spindle proprioceptors. In conclusion, the cholinergic terminals-part of which coexpress PACAP and/or L-ENK-that innervate MEPs in the porcine esophagus have a vagal origin, whereas the nNOS/VIP/GAL/NPY-ir fibers co-innervating these MEPs are intrinsic in nature. The regional differences observed along the esophageal length pertain to the neurochemical content of the vagal motor innervation of the MEPs.     

Disruption of Trkb-mediated signaling induces disassembly of postsynaptic receptor clusters at neuromuscular junctions

Neurotrophins and tyrosine receptor kinase (Trk) receptors are expressed in skeletal muscle, but it is unclear what functional role Trk-mediated signaling plays during postnatal life. Full-length TrkB (trkB.FL) as well as truncated TrkB (trkB.t1) were found to be localized primarily to the postsynaptic acetylcholine receptor- (AChR-) rich membrane at neuromuscular junctions. In vivo, dominant-negative manipulation of TrkB signaling using adenovirus to overexpress trkB.t1 in mouse sternomastoid muscle fibers resulted in the disassembly of postsynaptic AChR clusters at neuromuscular junctions, similar to that observed in mutant trkB+/- mice. When TrkB-mediated signaling was disrupted in cultured myotubes in the absence of motor nerve terminals and Schwann cells, agrin-induced AChR clusters were also disassembled. These results demonstrate a novel role for neurotrophin signaling through TrkB receptors on muscle fibers in the ongoing maintenance of postsynaptic AChR regions.     

Changes in expression and organization of smooth-muscle-specific α-actin during fibronectin-mediated modulation of arterial smooth muscle cell phenotype

The spreading of freshly isolated arterial smooth muscle cells on a substrate of fibronectin is mediated by an integrin receptor on the cell surface. It is associated with organization of actin filaments in stress fibers and marked changes in cell morphology and function, collectively referred to as a transition from a contractile to a synthetic phenotype. To study further how extracellular matrix components affect smooth muscle phenotype, we have analyzed the expression and organization of smooth-muscle-specific alpha-actin in freshly isolated rat aortic smooth muscle cells cultured on a substrate of fibronectin under serum-free conditions. Northern-blot analysis showed that the expression of mRNA for smooth muscle alpha-actin, but not for nonmuscle actin, was strongly repressed during primary culture. On the other hand, the cellular content of alpha-actin was only moderately changed during the same period. Indirect immunofluorescence staining revealed that nonmuscle actin was rapidly organized in stress fibers, which did not stain with a monoclonal antibody against smooth muscle alpha-actin. Filament bundles containing alpha-actin were most prominent in the central parts of the cytoplasm and gradually disappeared as the spreading of the cells progressed. In contrast to the situation with nonmuscle actin, there was no apparent overlap in the staining for alpha-actin and the fibronectin receptor (alpha 5 beta 1), indicating that this receptor interacted with nonmuscle actin during the initial spreading process. Taken together, the results show that the expression and organization of smooth muscle alpha-actin are changed during interaction of the cells with fibronectin early in primary culture. They support the notion that integrin-mediated interactions between extracellular matrix components and arterial smooth muscle cells take part in the control of smooth muscle phenotype.     

Phenotype-dependent expression of alpha-smooth muscle actin in visceral smooth muscle cells

Alpha-Smooth muscle actin is one of the molecular markers for a phenotype of vascular smooth muscle cells, because the actin is a major isoform expressed in vascular smooth muscle cells and its expression is upregulated during differentiation. Here, we first demonstrate that the phenotype-dependent expression of this actin in visceral smooth muscles is quite opposite to that in vascular smooth muscles. This actin isoform is not expressed in adult chicken visceral smooth muscles including gizzard, trachea, and intestine except for the inner layer of intestinal muscle layers, whereas its expression is clearly detected in these visceral smooth muscles at early stages of the embryo (10-day-old embryo) and is developmentally downregulated. In cultured gizzard smooth muscle cells maintaining a differentiated phenotype, alpha-smooth muscle actin is not detected while its expression dramatically increases during serum-induced dedifferentiation. Promoter analysis reveals that a sequence (-238 to -219) in the promoter region of this actin gene acts as a novel negative cis-element. In conclusion, the phenotype-dependent expression of alpha-smooth muscle actin would be regulated by the sum of the cooperative contributions of the negative element and well-characterized positive elements, purine-rich motif, and CArG boxes and their respective transacting factors.     

A comparative study of the regeneration of the striated and smooth-muscle tissues of the esophagus]

By means of electron microscopy and observational histological techniques, using a similar experimental model, regeneration of the striated and smooth muscle tissues of the esophagus has been studied in rats. During early periods after lesion in both muscle tissues destructive-necrotic changes develop. Beginning from the 2nd-3d days regeneration processes are observed. The course and periodicity of the regenerative processes are specific for the types of the muscle tissues studied. Each of the muscle tissues of the esophagus has its own source of regeneration. For the smooth muscles those are myoblasts, that convert into smooth myocytes, for the striated ones--myosatellites, which after activation get out of the muscle fiber. During the restorative process of the muscular membrane no tissue interconnections are observed. This also proves certain specificity of the striated and smooth muscle tissues of the esophagus.     

Structure of neuromuscular junctions and differentiation of striated muscle fibers in mdx mice after bone-marrow stem cell therapy

Mdx mice are an experimental model of Duchenne muscular dystrophy caused by mutations in the dystrophin gene. Repeated cycles of muscle degeneration-regeneration are common for mdx mice. Disrupted neuromuscular junctions also characterize mdx mice. The structure of mdx mice neuromuscular junctions and the differentiation of striated muscle fibers were investigated 4, 8, 16, and 24 weeks after transplantation of C57BL/6 Lin(−) bone-marrow stem cells. We found that the death of striated muscle fibers decreased 4 weeks after the transplantation of bone-marrow stem cells. Accumulation of muscle fibers without centrally located nuclei began in 8 weeks and dystrophin synthesis increased in 16–24 weeks after the bone-marrow stem cells transplantation. On the longitudinal sections of quadriceps muscle of mdx mice 4 weeks after transplantation, we observed a reduced quantity of acetylcholine receptor clusters and an increase in their area in neuromuscular junctions. Sixteen weeks after the transplantation, the total area of neuromuscular junctions increased due to an enlarged number of acethylcholine receptors and their extended area. The single intramuscular transplantation of C57BL/6 Lin(−) bone-marrow stem cells induces the differentiation of mdx mice striated muscle fibers and improves the structure of neuromuscular junctions.     

Cytoskeletal remodeling in differentiated vascular smooth muscle is actin isoform dependent and stimulus dependent

Dynamic remodeling of the actin cytoskeleton plays an essential role in the migration and proliferation of vascular smooth muscle cells. It has been suggested that actin remodeling may also play an important functional role in nonmigrating, nonproliferating differentiated vascular smooth muscle (dVSM). In the present study, we show that contractile agonists increase the net polymerization of actin in dVSM, as measured by the differential ultracentrifugation of vascular smooth muscle tissue and the costaining of single freshly dissociated cells with fluorescent probes specific for globular and filamentous actin. Furthermore, induced alterations of the actin polymerization state, as well as actin decoy peptides, inhibit contractility in a stimulus-dependent manner. Latrunculin pretreatment or actin decoy peptides significantly inhibit contractility induced by a phorbol ester or an alpha-agonist, but these procedures have no effect on contractions induced by KCl. Aorta dVSM expresses alpha-smooth muscle actin, beta-actin, nonmuscle gamma-actin, and smooth muscle gamma-actin. The incorporation of isoform-specific cell-permeant synthetic actin decoy peptides, as well as isoform-specific probing of cell fractions and two-dimensional gels, demonstrates that actin remodeling during alpha-agonist contractions involves the remodeling of primarily gamma-actin and, to a lesser extent, beta-actin. Taken together, these results show that net isoform- and agonist-dependent increases in actin polymerization regulate vascular contractility.     

Transient production of alpha-smooth muscle actin by skeletal myoblasts during differentiation in culture and following intramuscular implantation

alpha-smooth muscle actin (SMA) is typically not present in post-embryonic skeletal muscle myoblasts or skeletal muscle fibers. However, both primary myoblasts isolated from neonatal mouse muscle tissue, and C2C12, an established myoblast cell line, produced SMA in culture within hours of exposure to differentiation medium. The SMA appeared during the cells' initial elongation, persisted through differentiation and fusion into myotubes, remained abundant in early myotubes, and was occasionally observed in a striated pattern. SMA continued to be present during the initial appearance of sarcomeric actin, but disappeared shortly thereafter leaving only sarcomeric actin in contractile myotubes derived from primary myoblasts. Within one day after implantation of primary myoblasts into mouse skeletal muscle, SMA was observed in the myoblasts; but by 9 days post-implantation, no SMA was detectable in myoblasts or muscle fibers. Thus, both neonatal primary myoblasts and an established myoblast cell line appear to similarly reprise an embryonic developmental program during differentiation in culture as well as differentiation within adult mouse muscles.