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PubMed2001

Caldesmon and smooth-muscle regulation.

Smooth muscles exist in the wall of hollow organs in our body and are responsible for controlling the flow of vital fluids that are essential for the normal function of the cardiovascular, respiratory, digestive, and reproductive systems. Many diseases, such as hypertension, asthma, indigestion, and premature birth, may attribute to malfunction of smooth-muscle contraction. It is therefore important to decipher how smooth-muscle contraction is regulated. This review attempts to give a brief overview of current understanding about the molecular mechanisms of smooth-muscle regulation and, in particular, to discuss possible roles of caldesmon in this regulatory process.

Actins↗
PubMed1993

In vitro studies of determinants of smooth muscle mechanics.

Smooth muscle contraction is dependent upon phosphorylation of the 20,000 Da light chain subunits of myosin. Whereas the kinetics of the hydrolysis of MgATP by smooth muscle myosin suggest a simple phosphorylation-dependent "on-off" mechanism, the contractile response in smooth muscle tissue is complex. Experiments to unravel this complexity have been performed in vitro using a combination of motility assays and kinetic techniques. Some insight into this complexity is obtained, but the mechanism and the regulation of smooth muscle contraction is still not completely known.

Animals↗
PubMed1986

Stoichiometry of contraction and Ca2+ mobilization by inositol 1,4,5-trisphosphate in isolated gastric smooth muscle cells.

Smooth muscle cells were isolated from the circular muscle layer of guinea pig stomach and permeabilized by brief exposure to saponin. Both permeabilized and intact muscle cells contracted in response to cholecystokinin octapeptide (CCK-8) and acetylcholine, but only permeabilized muscle cells contracted in response to inositol 1,4,5-trisphosphate (InsP3). The contractile response to InsP3 was prompt (peak less than 5 s), concentration-dependent (EC50-0.3 microM), and insensitive to antimycin or oligomycin. Contraction induced by either InsP3 or CCK-8 was accompanied by a concentration-dependent increase in free Ca2+ that was directly correlated with the magnitude of contraction. Both InsP3 and CCK-8 caused rapid net efflux of Ca2+ from cells preloaded with 45Ca2+. Contraction, increase in free Ca2+ concentration, and net 45Ca2+ efflux elicited by a combination of maximal concentrations of InsP3 and CCK-8 were not significantly different from those elicited by maximal concentrations of either agent alone. Repeated stimulation of single muscle cells with either InsP3 or CCK-8 in Ca2+-free medium caused eventual loss of the contractile response to all agents. The response to all agents was restored upon re-exposure of the cell to a cytosol-like concentration of Ca2+, implying equal access of InsP3 and receptor-linked agonists to the same intracellular Ca2+ store. The results demonstrate that InsP3 mimics the effects of receptor-linked agonists on contraction and mobilization of intracellular Ca2+ in permeabilized smooth muscle cells that retain the functional properties of intact smooth muscle cells and support a role for InsP3 as membrane-derived messenger responsible for mobilization of intracellular Ca2+ in smooth muscle cells.

Animals↗
PubMed1995

Functional endothelin A receptor on gastric smooth muscle cells.

Smooth muscle cells isolated from the gastric muscle layers of the guinea pig were used to examine the functional endothelin receptor subtype responsible for gastric smooth muscle contraction by endothelin. Endothelin-1 induced the contraction of these cells in a dose-dependent manner, with an ED50 value of 0.2 nM. Endothelin-3-induced contraction was significantly less than that of endothelin-1. Sarafotoxin 6c (10(-11)-10(-7) M), endothelinB highly selective agonist, did not elicit the contraction of gastric smooth muscle cells. BQ-123, endothelinA selective antagonist, inhibited the contractile response produced by 10(-7) M endothelin-1 in a dose-dependent manner, with an IC50 value of 5 nM. These results strongly suggest that endothelin elicited the contraction of gastric smooth muscle cells via functional endothelinA receptor subtype.

Animals↗
PubMed2006

PRISM/PRDM6, a transcriptional repressor that promotes the proliferative gene program in smooth muscle cells.

Smooth muscle cells (SMCs) display remarkable phenotypic diversity and plasticity and can readily switch between proliferative and differentiated states in response to extracellular cues. In an effort to identify novel transcriptional regulators of smooth muscle phenotypes, we compared the gene expression profiles of arterial and venous SMCs by microarray-based transcriptional profiling. Among numerous genes displaying distinct expression patterns in these two SMC types, we discovered an expressed sequence tag encoding a previously uncharacterized zinc finger protein belonging to the PRDM (PRDI-BF1 and RIZ homology domain) family of chromatin-remodeling proteins and named it PRISM (PR domain in smooth muscle). PRISM is expressed in a variety of smooth muscle-containing tissues and displays especially robust expression in the cardiac outflow tract and descending aorta during embryogenesis. PRISM is localized to the nucleus and contains an amino-terminal PR domain and four Krüppel-like zinc fingers at the carboxy terminus. We show that PRISM acts as a transcriptional repressor by interacting with class I histone deacetylases and the G9a histone methyltransferase, thereby identifying PRISM as a novel SMC-restricted epigenetic regulator. Overexpression of PRISM in cultured primary SMCs induces genes associated with the proliferative smooth muscle phenotype while repressing regulators of differentiation, including myocardin and GATA-6. Conversely, small interfering RNA-mediated knockdown of PRISM slows cell growth and induces myocardin, GATA-6, and markers of SMC differentiation. We conclude that PRISM acts as a novel epigenetic regulator of SMC phenotypic plasticity by suppressing differentiation and maintaining the proliferative potential of vascular SMCs.

Amino Acid Sequence↗
PubMed1988

Calcium dependence of myosin light chain phosphorylation in smooth muscle cells.

Smooth muscle cells grown in culture may provide a model system for studying the Ca2+ dependence of myosin light chain phosphorylation. Tracheal smooth muscle cells in culture had 60% of the myosin content of tracheal tissue. Western analysis with appropriate antibodies demonstrated one 20-kDa light chain and the presence of a 150-kDa myosin light chain kinase in both tracheal smooth muscle tissue and cells. Moreover, tracheal cells contained 74% of the myosin light chain kinase activity measured in tissue. Similar types of analyses of nonmuscle cells showed a much lower myosin and myosin light chain kinase content. Carbachol (10 microM) or ionomycin (10 microM) stimulation of fura-2-containing cells resulted in a rapid increase in cytosolic free Ca2+ concentration and in the extent of myosin light chain phosphorylation. Maximal increases in Ca2+ concentrations were greater with ionomycin than with carbachol (4400 versus 492 nM). Light chain phosphorylation increased after the Ca2+ concentration exceeded 200 nM from control values of 165 nM. Half-maximal phosphorylation (33%) occurred at 260 nM Ca2+. There was a similar relationship between free cytosolic Ca2+ concentrations and the extent of myosin light chain phosphorylation in carbachol- and ionomycin-stimulated cells. This relationship had a Hill coefficient of 2.7. These observations indicate that small changes in Ca2+ concentrations stimulate myosin light chain phosphorylation and thus presumably contraction in smooth muscle cells.

Animals↗
PubMed1999

Bronchoalveolar lavage fluid from asthmatic subjects is mitogenic for human airway smooth muscle.

Airway smooth muscle proliferation may contribute to the airway wall remodeling seen in asthma. In this study we tested for the presence of airway smooth muscle mitogenic activity in bronchoalveolar lavage (BAL) fluid obtained from 12 atopic asthmatics before and serially after segmental allergen challenge, and from four normal subjects who did not undergo allergen challenge. Mitogenic effect was assessed by coincubating BAL fluid with human airway smooth muscle cells, and measuring its effect on (3)[H]thymidine incorporation and cell number. Induction of ERK phosphorylation and cyclin D(1) protein abundance were also assessed. Compared with serum-free medium alone, BAL fluid obtained from normal subjects increased thymidine incorporation, cell number, ERK phosphorylation, and cyclin D(1) abundance. BAL fluid from asthmatic subjects prior to allergen challenge induced even greater increases in all measures, except for cell number, which was similar to that observed with normal subjects' BAL fluid. Incubation with lavage fluid obtained 48 h after segmental allergen challenge in atopic asthmatics caused yet further increases in thymidine incorporation, cell number, and cyclin D(1) protein abundance. Molecular sieving of prechallenge BAL fluid from three asthmatic subjects demonstrated that mitogenic activity was present exclusively in the > 10 kD fraction. These results provide the first direct demonstration that fluid lining the airways of asthmatics contains excess mitogenic activity for human airway smooth muscle, and that this activity increases further after allergen challenge.

Adolescent↗
PubMed1963

PHYSIOLOGICAL EVIDENCE FOR MULTIPLE CALCIUM SITES IN SMOOTH MUSCLE.

Inherent smooth muscle tone and acetylcholine-induced contractions of the isolated longitudinal muscle from guinea pig ileum are inhibited by 1.2 M ethanol. The inhibitions are antagonized by high concentrations of calcium ion in the external medium. Previous work has indicated that an acetylcholine-induced increase in potassium efflux from the ileal muscle is also inhibited by ethanol and reactivated by high concentrations of calcium ion. It was found in this study that, in addition to ethanol, a drastic reduction in the calcium ion concentration of the bathing medium appeared to produce a depression of this drug-induced increase in potassium efflux. Preincubating the muscle in a reduced calcium ion concentration also inhibited the increase in potassium efflux initiated by a high potassium medium. Conversely, the exposure of the muscle to 1.2 M ethanol did not depress the potassium-induced increase in potassium efflux. Increases in smooth muscle tone produced by a high potassium medium have been reported to be inhibited both by ethanol and by a depletion of external calcium. These data suggest that the calcium ions which activate or enhance a potassium-induced increase in potassium efflux are not bound to the same loci in the muscle fiber as those involved in an acetylcholine-induced increase in potassium efflux or those involved in a potassium-induced increase in smooth muscle tone.

Acetylcholine↗
PubMed1991

Expression of isoforms of internal Ca2+ pump in cardiac, smooth muscle and non-muscle tissues.

Smooth muscle and several non-muscle tissues contain mRNA for an alternative splice of the mRNA for the cardiac sarcoplasmic reticulum (SR) Ca,Mg-ATPase. Based on amino acid composition deduced from cDNA sequences the cardiac isoform (Ic) is 110 kDa while the smooth muscle and the non-muscle isoform (Is) is 115 kDa. This prediction in their molecular masses was tested at the protein level in rabbit stomach, aorta, uterus and vas deferens smooth muscles; stomach mucosa, brain, liver, kidney and heart. The major species of the acylphosphates formed in the presence of Ca2+ and electrophoresed in acid SDS-acrylamide gels were 5 kDa smaller for the heart (Ic) than those for all the other tissues (Is). The size difference was also confirmed in Western blots using a monoclonal antibody which binds to both Is and Ic. Thus consistent with the mRNA splices for the internal Ca2+ pumps previously reported to be present in these tissues, rabbit heart expresses predominantly the Ca2+ pump protein Ic and the various smooth muscle, mucosa, brain, liver and kidney express mainly the isoform Is.

Animals↗
PubMed1996

Smooth muscle fatty acid binding protein: a regulator of smooth muscle contraction?

A fatty acid binding protein has been isolated from chicken gizzard smooth muscle. The partial amino acid sequence (EMBL P80565) of this protein shows high sequence similarities with other members of the fatty acid binding protein family. This is the first fatty acid binding protein isolated from smooth muscle. It may be involved in the regulation of smooth muscle contraction by transporting polyunsaturated fatty acids (e.g. arachidonic acid).

Amino Acid Sequence↗
PubMed1996

Adenovirus-assisted lipofection: efficient in vitro gene transfer of luciferase and cytosine deaminase to human smooth muscle cells.

Smooth muscle cells (SMC) are a central cell type involved in multiple processes of coronary artery diseases including restenosis and therefore are major target cells for different aspects of gene transfer. Previous attempts to transfect primary arterial cells using different techniques like liposomes, CaPO4 and electroporation resulted in only low transfection efficiency. The development of recombinant adenoviruses dramatically improved the delivery of foreign genes into different cell types including SMC. However, cloning and identification of recombinants remain difficult and time-consuming techniques. The present study demonstrates that a complex consisting of reporter plasmid encoding firefly luciferase (pLUC), polycationic liposomes and replication-deficient adenovirus was able to yield very high in vitro transfection of primary human smooth muscle cells under optimized conditions. The technique of adenovirus-assisted lipofection (AAL) increases transfer and expression of plasmid DNA in human smooth muscle cells in vitro up to 1000-fold compared to lipofection. To verify the applicability of AAL for gene transfer into human smooth muscle cells we studied a gene therapy approach to suppress proliferation of SMC in vitro, using the prokaryotic cytosine deaminase gene (CD) which enables transfected mammalian cells to deaminate 5-fluorocytosine (5-FC) to the highly toxic 5-fluorouracil (5-FU). The effect of a transient CD expression on RNA synthesis was investigated by means of a cotransfection with a RSV-CD expression plasmid and the luciferase reporter plasmid. Western blot analysis demonstrated high expression of CD protein in transfected SMC. Cotransfected SMC demonstrated two-fold less luciferase activity in the presence of 5-FC (5 mmol/l) after 48 h compared to cells transfected with a non-CD coding plasmid. The data demonstrate that a transient expression of CD could be sufficient to reduce the capacity of protein synthesis in human SMC. This simple and effective in vitro transfection method may also be applicable to in vivo delivery of target genes to the vascular wall to inhibit SMC proliferation.

Adenoviruses, Human↗
PubMed1988

Sites phosphorylated in myosin light chain in contracting smooth muscle.

Purified smooth muscle myosin light chain can be phosphorylated at multiple sites by myosin light chain kinase and protein kinase C. We have determined the sites phosphorylated on myosin light chain in intact bovine tracheal smooth muscle. Stimulation with 10 microM carbachol resulted in 66 +/- 5% monophosphorylated and 11 +/- 2% diphosphorylated myosin light chain after 1 min, and 47 +/- 4% monophosphorylated and 5 +/- 2% diphosphorylated myosin light chain after 30 min. Myosin heavy chain contained 0.06 +/- 0.01 mol of phosphate/mol of protein which did not change with carbachol. At both 1 and 30 min the monophosphorylated myosin light chain contained only phosphoserine whereas the diphosphorylated myosin light chain contained both phosphoserine and phosphothreonine. Two-dimensional peptide mapping of tryptic digests of monophosphorylated and diphosphorylated myosin light chain obtained from carbachol-stimulated tissue was similar to the peptide maps of purified light chain monophosphorylated and diphosphorylated, respectively, by myosin light chain kinase; these maps were distinct from the map obtained with tracheal light chain phosphorylated by protein kinase C. Phosphorylation of tracheal smooth muscle myosin light chain by myosin light chain kinase yields the tryptic phosphopeptide ATSNVFAMFDQSQIQEFK with S the phosphoserine in the monophosphorylated myosin light chain and TS the phosphotreonine and phosphoserine in the diphosphorylated myosin light chain. Thus, stimulation of tracheal smooth muscle with a high concentration of carbachol results in formation of both monophosphorylated and diphosphorylated myosin light chain although the amount of diphosphorylated light chain is substantially less than monophosphorylated light chain. In the intact muscle, myosin light chain is phosphorylated at sites corresponding to myosin light chain kinase phosphorylation.

Animals↗
PubMed1995

Effect of guinea-pig purified immunoglobulin G1 on the responsiveness of tracheal, aortic, vas deferens and ileum smooth muscles.

BACKGROUND: Smooth muscles hyperresponsiveness is a common feature in anaphylaxis and allergic diseases. OBJECTIVE: The aim of the present work was to investigate the effect of in vitro passive sensitization with highly purified immunoglobulin G1 (IgG1) on the responsiveness of tracheal, aortic, vas deferens and ileum smooth muscles. METHODS: Firstly, IgG1, obtained from actively sensitized BFA guinea-pigs, was purified by Protein A-Sepharose column and characterized by enzyme-linked immunosorbent assay (ELISA) and immunoelectrophoresis analysis. Concentration-response curves to spasmogens (acetylcholine for trachea and vas deferens, noradrenaline for aorta and histamine for ileum) were established before and after in vitro passive sensitization with IgG1. RESULTS: Contractile responses and maximal contractions were significantly enhanced after passive sensitization for all the organs. Maximal contractions were significantly increased in the trachea (+46.7%), aorta (+51%), vas deferens (+114.2%) and ileum (+117.2%). At the end of the experiments, the application of the sensitizing antigen induced a significant Schultz-Dale reaction of the smooth muscles. CONCLUSION: The present results show that the in vitro application of purified IgG1 can produce non-specific smooth muscle hyperreactivity and hypersensitivity. So, IgG1 can be considered as the main factor involved in the genesis of sensitization-induced hyperresponsiveness, and probably play a great role in hyperreactivity observed during allergic diseases and anaphylaxis.

Animals↗
PubMed2004

Contribution of store-operated Ca2+ entry to pHo-dependent changes in vascular tone of porcine coronary smooth muscle.

Vascular smooth muscle contracts on increases of extracellular pH (pH(o)) and relaxes on pH(o) decreases possibly resulting from changes in transsarcolemmal Ca(2+) influx. Therefore, we studied store-operated Ca(2+) entry (SOCE; i.e. capacitative Ca(2+) entry (CCE)) during acidification (pH(o)=6.5) and alkalinization (pH(o)=8.0) in isolated porcine coronary smooth muscle cells (SMCs) by monitoring cytoplasmic Ca(2+) ([Ca(2+)](i)) and divalent cation entry (Mn(2+) quench) with fura-2/AM-fluorometry. Additionally, we evaluated the contribution of SOCE to pH(o)-dependent changes in isometric tension of porcine coronary smooth muscle strips. SOCE elicited in SMCs by the SERCA inhibitor BHQ was strongly modulated by pH(o) showing a decrease upon acidification and vice versa an increase upon alkalinization. BHQ-mediated tension of smooth muscle strips also revealed strong pH(o) dependence. In contrast, L-VOC-dependent tension ([K(+)](o)=20 and 40 mmol l(-1)) was remarkably less affected by pH(o) changes. Moreover, refilling of depleted Ca(2+) stores after repeated M(3)-cholinergic receptor stimulation could be almost completely inhibited by SKF 96365 and was markedly reduced by acidification and considerably enhanced by alkalinization pointing to a major role of SOCE in refilling. We conclude that vascular tone particularly responds to alterations in pH(o) whenever SOCE substantially contributes to the amount of activator Ca(2+) for contraction.

Acetylcholine↗
PubMed1995

Prostaglandin E2 contracts vascular smooth muscle and inhibits potassium currents in vascular smooth muscle cells of rat tail artery.

There is evidence to suggest that PGE2 plays an important role in the regulation of vascular smooth muscle tone. To determine the cellular basis of this action, we studied the effect of PGE2 on force in helical muscle strips from rat tail artery. PGE2 evoked a sustained contractile response. The contractile response was concentration-dependent, with an EC50 value of 9.6 microM. Patch-clamp studies were conducted to investigate the effects of PGE2 on K channels in isolated vascular smooth muscle cells from rat tail artery. Current-clamp studies showed that PGE2 (1 microM) depolarized the membrane by 15.9 +/- 1.3 mV. Under voltage-clamp conditions, a voltage-dependent, delayed outward rectifier K current was generated by stepwise depolarization from a holding potential of -80 mV. The current, which was activated at -45 to -40 mV and showed almost no inactivation, was inhibited by 45% using 10 mM TEA. PGE2 inhibited the outward K current in a concentration-dependent manner, with EC50 values of 3.5 microM and 4.9 microM in primary and subcultured cells, respectively. The PGE2 receptor antagonist sodium meclofenamate abolished the PGE2-induced K current inhibition. Furthermore, the intracellular application of guanosine 5'-O(-)[2-thiodiphosphate] (GDP beta S), a G protein inhibitor, and pretreatment of the cells with cholera toxin prevented the PGE2-induced inhibition, whereas application of pertussis toxin did not.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1988

Calcium release in smooth muscle.

In smooth muscle, maintenance of the contractile response is due to Ca2+ influx through two types of Ca2+ channel, a voltage-dependent Ca2+ channel and a receptor-linked Ca2+ channel. However, a more transient contraction can be obtained by release of Ca2+ from a cellular store, possibly the sarcoplasmic reticulum. In spike generating smooth muscle (e.g., guinea-pig taenia caeci), spike discharges may trigger the release of cellular Ca2+ by activating a Ca2+-induced Ca2+ release mechanism. Caffeine directly activates this mechanism in the absence of a triggered Ca2+ influx. In contrast to this, maintained depolarization may not only release but also refill the Ca2+ store. Drug-receptor interactions also release Ca2+ from a cellular store. This release may be elicited with inositol trisphosphate produced by receptor-linked phosphoinositide turnover. In non-spike generating smooth muscle (e.g., rabbit thoracic aorta), maintained membrane depolarization does not release but, instead, fills the Ca2+ store. However, caffeine and receptor-agonists release the Ca2+ store - possibly by activating the Ca2+-induced Ca2+ release mechanism and phosphoinositide turnover, respectively. The Ca2+ store in smooth muscle is filled by Ca2+ entry through voltage dependent Ca2+ channels and also by resting Ca2+ influx in the absence of receptor-agonists. The Ca2+ entering the cells through these pathways may be accumulated by the Ca2+ store and may activate the contractile filaments.

Animals↗
PubMed1993

[Drug-receptor interactions in single smooth muscle cells].

Smooth muscle tissues were contracted by excitation of each muscle cell. Single cells prepared from guinea pig taenia caecum and trachea were contracted by extracellular application of acetylcholine and/or carbachol, whose concentrations were the same as those in the tissues. The concentration-response curve was shifted in a parallel fashion by competitive antagonists. The pA2-values of the antagonists were in good agreement with those estimated using the intact tissue. The apparent dissociation constants of cholinergic drugs estimated from inhibition of the specific binding of [3H]QNB (quinuclidinyl benzilate) to the single cells by the cholinergic drugs were also in agreement with the values in other membrane preparations. Similar findings were obtained in the single cells, microsomal fractions and isolated tissues from the guinea pig tracheal smooth muscles. In rabbit aortic single cells, the existence of two pharmacologically distinct alpha 1-adrenoceptor subtypes, alpha 1A and alpha 1B, in vascular smooth muscle cells was supported. Furthermore, the amount of prostaglandin F2 alpha released from guinea pig tracheal single cells was increased through activation by alpha 2-adrenoceptor agonists. The amount of prostaglandin F2 alpha released by norepinephrine decreased with age, while the total amount of alpha 2-adrenoceptors and the dissociation constants of the alpha 2-adrenergic drugs from the receptor did not change. The relaxation induced by beta-adrenoceptors did not alter with age. The total amount of beta-adrenoceptor and the dissociation constants of beta-adrenergic drugs from their receptor did not alter with age. An excellent relationship between the potency of isoprenaline and the maximum binding of [3H]dihydroalprenolol estimated in the single cells from 6- to 40-week-old guinea pigs was found, suggesting that the increase in the potency of isoprenaline is due to the increase in the maximum binding. The value in the single cells from 100-week-old guinea pigs deviated significantly from the regression line. This result suggests that the decrease in potency in the single cells from 100-week-old animals is due to a change in post beta-receptor processes in responsiveness. The smooth muscle single cells are useful for the study of drug-receptor interactions. Furthermore, post-receptor processes in responsiveness were discussed.

Acetylcholine↗
PubMedDate not supplied

Cytodifferentiation and expression of alpha-smooth muscle actin mRNA and protein during primary culture of aortic smooth muscle cells. Correlation with cell density and proliferative state.

To investigate a possible correlation between cytodifferentiation, proliferation, and actin expression, smooth muscle cells from the 9-week-old rabbit aortic media were enzyme-dispersed into single cells and were plated in primary culture at different initial seeding densities. The volume fraction of myofilaments (Vv myo) in cells seeded moderately densely fell from 39.5% +/- 1.2% in the intact aortic media to 11.5% +/- 1.6% on Day 5, one day before the onset of logarithmic growth. The Vv myo remained low over the next 3 days, then began to rise as the density of cells increased, returning almost to the original levels after confluency and 1.84 cumulative population doublings (CPD). The expression of alpha-smooth muscle actin mRNA followed a similar time course of change, falling from 84.7% +/- 1.2% of total actin mRNA in freshly isolated cells to 54.0% +/- 6.5% on Day 5, returning to 87.5% +/- 0.5% after confluency. In these cultures, the alpha-smooth muscle actin protein content was 93.7% +/- 2.9% of total actin in freshly isolated cells, 68.7% +/- 3.1% on Day 5, and 73.3% +/- 2.5% 3 days after confluency. In densely seeded cultures, the Vv myo and expression of alpha-smooth muscle actin mRNA fell only slightly on Day 5 and rose to original levels upon confluency after 0.33 CPD. However, at the protein level, alpha-smooth muscle actin decreased on Day 5 and remained low on Day 12. The Vv myo, alpha-smooth muscle actin mRNA, and actin protein of sparsely seeded cells fell on Day 5 and then remained low throughout the culture period, including 5 days after confluency (Day 24), when the cells had undergone 5.37 CPD. Cells that were maintained subconfluent but quiescent on Day 7 in culture had the same low Vv myo, low alpha-actin mRNA expression, and low alpha-actin protein content as actively proliferating cells. The results show that Vv myo and alpha-smooth muscle actin mRNA undergo parallel changes during primary culture according to seeding density, but not to replication, and that alpha-smooth muscle actin protein decreases in culture then remains low irrespective of culture conditions.

Actins↗