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A J Halayko

Publications and source records attributed to A J Halayko.

At least 19 recordsLinked to original sources
PubMed2006

Airway smooth muscle phenotype and function: interactions with current asthma therapies.

Asthma incidence has climbed markedly in the past two decades despite an increased use of medications that suppress airway inflammation and repress contraction of smooth muscle that encircles the airways. Asthmatics exhibit episodes of airway inflammation that potentiates reversible airway smooth muscle spasm. A hallmark diagnostic symptom of asthma is airway hyperresponsiveness to inhaled non-allergic stimuli, such as methacholine, that directly induce airway smooth muscle contraction. Inhaled gluccocorticoids are used for first-line prevention of airway inflammation, and are frequently combined with inhaled beta2-adrenoceptor agonists that can effectively relax airway smooth muscle and restore airway conductance. Leukotriene receptor antagonists and anti-cholinergics can also be used in many patients to ensure optimal control of symptoms. With increasing disease duration irreversible airway restriction develops from inflammation-driven fibro-proliferative airway remodeling that includes increased deposition of extracellular matrix, the accumulation of airway smooth muscle, and increased numbers of myofibroblasts. Mature airway smooth muscle cells are phenotypically plastic, enabling them to subserve contractile, proliferative, migratory and secretory functional responses that contribute to airway remodeling and persistent hyperresponsiveness. This review assesses current understanding of acute and chronic effects of common anti-asthma medications on the diverse phenotype and functional characteristics of airway smooth muscle cells. Furthermore, we describe the significance of these effects in the treatment of asthma symptoms and pathogenesis.

Adrenergic beta-Agonists↗
PubMed2005

Hypoxia induces hypersensitivity and hyperreactivity to thromboxane receptor agonist in neonatal pulmonary arterial myocytes.

PPHN, caused by perinatal hypoxia or inflammation, is characterized by an increased thromboxane-prostacyclin ratio and pulmonary vasoconstriction. We examined effects of hypoxia on myocyte thromboxane responsiveness. Myocytes from 3rd-6th generation pulmonary arteries of newborn piglets were grown to confluence and synchronized in contractile phenotype by serum deprivation. On the final 3 days of culture, myocytes were exposed to 10% O2 for 3 days; control myocytes from normoxic piglets were cultured in 21% O2. PPHN was induced in newborn piglets by 3-day hypoxic exposure (Fi(O2) 0.10); pulmonary arterial myocytes from these animals were maintained in normoxia. Ca2+ mobilization to thromboxane mimetic U-46619 and ATP was quantified using fura-2 AM. Three-day hypoxic exposure in vitro results in increased basal [Ca2+]i, faster and heightened peak Ca2+ response, and decreased U-46619 EC50. These functional changes persist in myocytes exposed to hypoxia in vivo but cultured in 21% O2. Blockade of Ca2+ entry and store refilling do not alter peak U-46619 Ca2+ responses in hypoxic or normoxic myocytes. Blockade of ryanodine-sensitive or IP3-gated intracellular Ca2+ channels inhibits hypoxic augmentation of peak U-46619 response. Ca2+ response to ryanodine alone is undetectable; ATP-induced Ca2+ mobilization is unaltered by hypoxia, suggesting no independent increase in ryanodine-sensitive or IP3-linked intracellular Ca2+ pool mobilization. We conclude hypoxia has a priming effect on neonatal pulmonary arterial myocytes, resulting in increased resting Ca2+, thromboxane hypersensitivity, and hyperreactivity. We postulate that hypoxia increases agonist-induced TP-R-linked IP3 pathway activation. Myocyte thromboxane hyperresponsiveness persists in culture after removal from the initiating hypoxic stimulus, suggesting altered gene expression.

15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5↗
PubMed2004

Latrunculin B increases force fluctuation-induced relengthening of ACh-contracted, isotonically shortened canine tracheal smooth muscle.

We hypothesized that differences in actin filament length could influence force fluctuation-induced relengthening (FFIR) of contracted airway smooth muscle and tested this hypothesis as follows. One-hundred micromolar ACh-stimulated canine tracheal smooth muscle (TSM) strips set at optimal reference length (Lref) were allowed to shorten against 32% maximal isometric force (Fmax) steady preload, after which force oscillations of +/-16% Fmax were superimposed. Strips relengthened during force oscillations. We measured hysteresivity and calculated FFIR as the difference between muscle length before and after 20-min imposed force oscillations. Strips were relaxed by ACh removal and treated for 1 h with 30 nM latrunculin B (sequesters G-actin and promotes depolymerization) or 500 nM jasplakinolide (stabilizes actin filaments and opposes depolymerization). A second isotonic contraction protocol was then performed; FFIR and hysteresivity were again measured. Latrunculin B increased FFIR by 92.2 +/- 27.6% Lref and hysteresivity by 31.8 +/- 13.5% vs. pretreatment values. In contrast, jasplakinolide had little influence on relengthening by itself; neither FFIR nor hysteresivity was significantly affected. However, when jasplakinolide-treated tissues were then incubated with latrunculin B in the continued presence of jasplakinolide for 1 more h and a third contraction protocol performed, latrunculin B no longer substantially enhanced TSM relengthening. In TSM treated with latrunculin B + jasplakinolide, FFIR increased by only 3.03 +/- 5.2% Lref and hysteresivity by 4.14 +/- 4.9% compared with its first (pre-jasplakinolide or latrunculin B) value. These results suggest that actin filament length, in part, determines the relengthening of contracted airway smooth muscle.

Acetylcholine↗
PubMed2001

Molecular mechanisms of phenotypic plasticity in smooth muscle cells.

Morphological, functional, molecular and cell biology studies have revealed a striking multifunctional nature of individual smooth muscle cells (SMC). SMCs manifest phenotypic plasticity in response to changes in environment and functional requirements, acquiring a range of structural and functional properties bounded by two extremes, called "synthetic" and "contractile." Each phenotypic state is characterized by expression of a unique set of structural, contractile, and receptor proteins and isoforms that correlate with differing patterns of gene expression. Recent studies have identified signaling pathways and transcription factors (e.g., RhoA GTPase/ROCK, also known as Rho kinase, and serum response factor) that regulate the transcriptional activities of genes encoding proteins associated with the contractile apparatus. Mechanical plasticity of contractile-state smooth muscle further extends SMC functional diversity. This may also be regulated, in part, by the RhoA GTPase/ROCK pathway, via reorganization of cytoskeletal and contractile proteins. Future studies that define transcriptional and posttranscriptional mechanisms of SMC plasticity are necessary to fully understand the role of SMC in the pathogenesis and morbidity of human diseases of the airways, vasculature, and gastrointestinal tract.

Animals↗
PubMed2000

Physiological control of smooth muscle-specific gene expression through regulated nuclear translocation of serum response factor.

Prolonged serum deprivation induces a structurally and functionally contractile phenotype in about 1/6 of cultured airway myocytes, which exhibit morphological elongation and accumulate abundant contractile apparatus-associated proteins. We tested the hypothesis that transcriptional activation of genes encoding these proteins accounts for their accumulation during this phenotypic transition by measuring the transcriptional activities of the murine SM22 and human smooth muscle myosin heavy chain promoters during transient transfection in subconfluent, serum fed or 7 day serum-deprived cultured canine tracheal smooth muscle cells. Contrary to our expectation, SM22 and smooth muscle myosin heavy chain promoter activities (but not viral murine sarcoma virus-long terminal repeat promoter activity) were decreased in long term serum-deprived myocytes by at least 8-fold. Because serum response factor (SRF) is a required transcriptional activator of these and other smooth muscle-specific promoters, we evaluated the expression and function of SRF in subconfluent and long term serum-deprived cells. Whole cell SRF mRNA and protein were maintained at high levels in serum-deprived myocytes, but SRF transcription-promoting activity, nuclear SRF binding to consensus CArG sequences, and nuclear SRF protein were reduced. Furthermore, immunocytochemistry revealed extranuclear redistribution of SRF in serum-deprived myocytes; nuclear localization of SRF was restored after serum refeeding. These results uncover a novel mechanism for physiological control of smooth muscle-specific gene expression through extranuclear redistribution of SRF and consequent down-regulation of its transcription-promoting activity.

Animals↗
PubMed2000

Fas cross-linking induces apoptosis in human airway smooth muscle cells.

Hypertrophy and hyperplasia lead to excess accumulation of smooth muscle in the airways of human asthmatic subjects. However, little is known about mechanisms that might counterbalance these processes, thereby limiting the quantity of smooth muscle in airways. Ligation of Fas on the surface of vascular smooth muscle cells and nonmuscle airway cells can lead to apoptotic cell death. We therefore tested the hypotheses that 1) human airway smooth muscle (HASM) expresses Fas, 2) Fas cross-linking induces apoptosis in these cells, and 3) tumor necrosis factor (TNF)-alpha potentiates Fas-mediated airway myocyte killing. Immunohistochemistry using CH-11 anti-Fas monoclonal IgM antibody revealed Fas expression in normal human bronchial smooth muscle in vivo. Flow cytometry using DX2 anti-Fas monoclonal IgG antibody revealed that passage 4 cultured HASM cells express surface Fas. Surface Fas decreased partially during prolonged serum deprivation of cultured HASM cells and was upregulated by TNF-alpha stimulation. Fas cross-linking with CH-11 antibody induced apoptosis in cultured HASM cells, and this effect was reduced by long-term serum deprivation and synergistically potentiated by concomitant TNF-alpha exposure. TNF-alpha did not induce substantial apoptosis in the absence of Fas cross-linking. These data represent the first demonstration that Fas is expressed on HASM and suggest a mechanism by which Fas-mediated apoptosis could act to oppose excess smooth muscle accumulation during airway remodeling in asthma.

Antibodies, Monoclonal↗
PubMed2000

Selective restoration of calcium coupling to muscarinic M(3) receptors in contractile cultured airway myocytes.

We previously demonstrated that after several days of serum deprivation about one-sixth of confluent cultured canine tracheal myocytes acquire an elongated, structurally and functionally contractile phenotype. These myocytes demonstrated significant shortening on ACh exposure. To evaluate the mechanism by which these myocytes acquire responsiveness to ACh, we assessed receptor-Ca(2+) coupling using fura 2-AM fluorescence imaging and muscarinic receptor expression using Western analysis. Cells were grown to confluence in 10% fetal bovine serum and then maintained for 7-13 days in serum-free medium. A fraction of serum-deprived cells exhibited reproducible intracellular Ca(2+) mobilization in response to ACh that was uniformly absent from airway myocytes before serum deprivation. The Ca(2+) response to 10(-4) M ACh was ablated by inositol 1,4,5-trisphosphate (IP(3)) receptor blockade using 10(-6) M xestospongin C but not by removal of extracellular Ca(2+). Also, 10(-7) M atropine or 10(-7) M 4-diphenylacetoxy-N-methylpiperidine completely blocked the response to ACh, but intracellular Ca(2+) mobilization was not ablated by 10(-6) M pirenzepine or 10(-6) M methoctramine. In contrast, 10(-5) M bradykinin (BK) was without effect in these ACh-responsive myocytes. Interestingly, myocytes that did not respond to ACh demonstrated robust increases in intracellular Ca(2+) on exposure to 10(-5) M BK that were blocked by removal of extracellular Ca(2+) and were only modestly affected by IP(3) receptor blockade. Serum deprivation increased the abundance of M(3) receptor protein and of BK(2) receptor protein by two- to threefold in whole cell lysates within 2 days of serum deprivation, whereas M(2) receptor protein fell by >75%. An increase in M(3) receptor abundance and restoration of M(3) receptor-mediated Ca(2+) mobilization occur concomitant with reacquisition of a contractile phenotype during prolonged serum deprivation. These data demonstrate plasticity in muscarinic surface receptor expression and function in a subpopulation of airway myocytes that show mutually exclusive physiological and pharmacological diversity with other cells in the same culture.

Acetylcholine↗
PubMed2000

Mutagenesis analysis of human SM22: characterization of actin binding.

SM22 is a 201-amino acid actin-binding protein expressed at high levels in smooth muscle cells. It has structural homology to calponin, but how SM22 binds to actin remains unknown. We performed site-directed mutagenesis to generate a series of NH(2)-terminal histidine (His)-tagged mutants of human SM22 in Escherichia coli and used these to analyze the functional importance of potential actin binding domains. Purified full-length recombinant SM22 bound to actin in vitro, as demonstrated by cosedimentation assay. Binding did not vary with calcium concentration. The COOH-terminal domain of SM22 is required for actin affinity, because COOH terminally truncated mutants [SM22-(1-186) and SM22-(1-166)] exhibited markedly reduced cosedimentation with actin, and no actin binding of SM22-(1-151) could be detected. Internal deletion of a putative actin binding site (154-KKAQEHKR-161) partially prevented actin binding, as did point mutation to neutralize either or both pairs of positively charged residues at the ends of this region (KK154LL and/or KR160LL). Internal deletion of amino acids 170-180 or 170-186 also partially or almost completely inhibited actin cosedimentation, respectively. Of the three consensus protein kinase C or casein kinase II phosphorylation sites in SM22, only Ser-181 was readily phosphorylated by protein kinase C in vitro, and such phosphorylation greatly decreased actin binding. Substitution of Ser-181 to aspartic acid (to mimic serine phosphorylation) also reduced actin binding. Immunostains of transiently transfected airway myocytes revealed that full-length NH(2)-terminal FLAG-tagged SM22 colocalizes with actin filaments, whereas FLAG-SM22-(1-151) does not. These data confirm that SM22 binds to actin in vitro and in vivo and, for the first time, demonstrate that multiple regions within the COOH-terminal domain are required for full actin affinity.

Actins↗
PubMed1999

Divergent differentiation paths in airway smooth muscle culture: induction of functionally contractile myocytes.

We tested the hypothesis that prolonged serum deprivation would allow a subset of cultured airway myocytes to reacquire the abundant contractile protein content, marked shortening capacity, and elongated morphology characteristic of contractile cells within intact tissue. Passage 1 or 2 canine tracheal smooth muscle (SM) cells were grown to confluence, then serum deprived for up to 19 days. During serum deprivation, two differentiation pathways emerged. One-sixth of the cells developed an elongated morphology and aligned into bundles. Elongated myocytes contained cables of contractile myofilaments, dense bodies, gap junctions, and membrane caveoli, ultrastructural features of contractile SM in tissue. These cells immunostained intensely for SM alpha-actin, SM myosin heavy chain (MHC), and SM22 (an SM-specific actin-binding protein), and Western analysis of culture lysates disclosed 1.8 (SM alpha-actin)-, 7.7 (SM MHC)-, and 5.8 (SM22)-fold protein increases during serum deprivation. Immunoreactive M3 muscarinic receptors were present in dense foci distributed throughout elongated, SM MHC-positive myocytes. ACh (10(-3) M) induced a marked shortening (59.7 +/- 14.4% of original length) in 62% of elongated myocytes made semiadherent by gentle proteolytic digestion, and membrane bleb formation (a consequence of contraction) occurred in all stimulated cells that remained adherent and so did not shorten. Cultured airway myocytes that did not elongate during serum deprivation instead became short and flattened, lost immunoreactivity for contractile proteins, lacked the M3 muscarinic-receptor expression pattern seen in elongated cells, and exhibited no contractile response to ACh. Thus we demonstrate that prolonged serum deprivation induces distinct differentiation pathways in confluent cultured tracheal myocytes and that one subpopulation acquires an unequivocally functional contractile phenotype in which structure and function resemble contractile myocytes from intact tissue.

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↗
PubMed1998

Expression and cytogenetic localization of the human SM22 gene (TAGLN).

SM22 is a 22-kDa protein identified variously as SM22, transgelin, WS3-10, or mouse p27. Though its precise function is unknown, it is abundant in smooth muscle and so may contribute to the physiology of this widespread tissue. We found that cosmid 16b6 contains the entire 5.4-kb, five-exon human SM22 gene (HGMW-approved symbol, TAGLN), and we cytogenetically localized the gene to chromosome 11q23.2. Northern analysis of human adult tissues showed that SM22 mRNA is most prevalent in smooth muscle-containing tissues, but is also found at lower levels in heart. The human SM22 promoter contains nuclear factor-binding motifs known to regulate transcription in smooth muscle, and human SM22 promoter-luciferase reporter constructs exhibited high transcriptional activity in A7r5 or primary canine aortic smooth muscle cells, but show little activity in nonmuscle COS7 cells. In addition, human SM22 promoter activity increased by two- to threefold upon serum stimulation of nonmuscle cells.

Adult↗
PubMed1998

Airway smooth muscle contractile, regulatory and cytoskeletal protein expression in health and disease.

The major part of research dealing with the biophysical and biochemical properties of airway smooth muscle is based on the assumption that the cells constituting the tissue are homogenous. For striated muscle this has been shown untenable. In recent years almost every property of vascular smooth muscle has been also demonstrated to be heterogeneous. This realization has been late in arriving on the airway smooth muscle research scene. Our own studies have shown that mechanical properties are, in quantitative terms, heterogeneously distributed down the airways and that contractility, for example, in extrapulmonary and intrapulmonary airways differs markedly. Another indication of heterogeneity is derived from studies of the biochemical properties of airway smooth muscle cells (ASMCs) in culture. Dramatic changes in phenotype expression were found with days in culture. Just after isolation from the tissue, the cells were of contractile type and contained mature isoforms of contractile, regulatory and cytoskeletal proteins. After the fourth day in culture the cellular phenotype changed such that contractile filaments diminished rapidly with smooth muscle isoforms being replaced by non-muscle isoforms. The cell assumed secretory or synthetic properties and commenced proliferating rapidly. It is possible that similar changes in phenotype could occur in vivo in cells undergoing hypertrophy or hyperplasia. Thus, a thickened medial layer of the type seen in the walls of airways from asthmatic airways is not necessarily one endowed with increased contractility and, in fact, the latter may be subnormal. Finally, using the so-called motility assay, we studied the velocity of translation of actin filaments by myosin molecules obtained from antigen-sensitized and control airway smooth muscle. We found no change in maximum velocity of actin translation. This was under conditions where the myosin light chain (MLC) was fully phosphorylated. However, in these tissues we found heterogeneity in myosin light chain kinase (MLCK) content which, we inferred, accounted for the difference in shortening velocity between control and sensitized muscle strips in vitro.

Animals↗
PubMed1998

Airway smooth muscle cell proliferation: characterization of subpopulations by sensitivity to heparin inhibition.

Growth and maturation state of airway smooth muscle cells (SMCs) are determinants of asthma pathophysiology. Heparin reduces airway SMC proliferation and arterial SMC replication and phenotypic modulation. Distinct arterial SMC subtypes, differing in heparin sensitivity, have been characterized. We assessed the cellular mechanisms underlying the growth and phenotype of heparin-treated canine tracheal myocytes in primary culture. Heparin reduced replication by 40%. Immunoblot assay of myosin, actin, and myosin light chain kinase revealed heparin had no effect on rapid spontaneous phenotypic modulation after the cells were plated. Heparin increased cellular protein and vimentin contents in confluent cultures, suggesting that it may induce hypertrophic growth. Cell cycle analysis revealed that heparin decreased serum-stimulated replicating myocyte number by 40%. Also, G2-M transit was 20% slower for the set of SMCs that proceeded past G1 in the presence of heparin. These data indicate that heparin does not inhibit airway SMC replication by blocking modulation from the contractile state. Moreover, airway smooth muscle is composed of distinct SMC populations differing in mitogen and antiproliferative mediator responsiveness. Identification of functionally divergent subgroups suggests that distinct sets of SMCs may contribute differentially to airway physiology and pathophysiology.

Animals↗
PubMed1998

Delayed rectifier K+ current of dog bronchial myocytes: effect of pollen sensitization and PKC activation.

The properties of delayed rectifier K+ current [IK(dr)] of canine airway smooth muscle cells isolated from small bronchi and its modulation by protein kinase C (PKC) were studied by whole cell patch clamp. IK(dr) activated positive to -40 mV, with half-maximal activation at -16 +/- 1.2 mV (n = 15) and average current density of 31 +/- 2.6 pA/pF (n = 15) at +30 mV. The capacitive surface area, current density, and voltage dependence of activation of IK(dr) of myocytes of ragweed pollen-sensitized dogs were not different from age-matched control dogs. However, the sensitization reduced the availability of IK(dr) between -40 and -20 mV due to a hyperpolarizing shift in the voltage dependence of steady-state inactivation (-29.9 +/- 1.2 in sensitized versus -26.0 +/- 0.7 mV in control dogs, n = 9 and 11, respectively; P < 0.05). PKC activation with diacylglycerol analog or phorbol ester depressed IK(dr) amplitude, whereas an inactive diacylglycerol analog had no effect. The hyperpolarizing shift in voltage dependence of inactivation and/or modulation of IK(dr) by PKC may be two mechanisms that contribute to the enhanced reactivity of bronchial tissues from ragweed pollen-sensitized dogs.

Animals↗
PubMed1998

Transcriptional regulation of smooth muscle contractile apparatus expression.

The transcriptional regulatory mechanisms that control gene expression during differentiation and contractile protein accumulation are becoming well understood in skeletal and cardiac muscle lineages. Current understanding of smooth muscle-specific gene transcription is much more limited, though recent studies have begun to shed light on this topic. In this review, we summarize some of the themes emerging from these studies and identify transcriptional regulatory elements common to several smooth muscle genes. These include potential binding sites for serum response factor, Sp1, AP2, Mhox, and YY1, as well as a potential transforming growth factor-beta control element. We speculate that it may be possible to manipulate smooth muscle-specific gene expression in asthma or pulmonary arterial hypertension as an eventual therapy.

Asthma↗
PubMed1997

Quantitative densitometry of proteins stained with coomassie blue using a Hewlett Packard scanjet scanner and Scanplot software.

In the present study we evaluated the performance of a software/scanner system that employed the Hewlett Packard (HP) ScanJet Plus and Scanplot Software for densitometric quantification of protein loads stained with Coomassie Brilliant Blue following sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Gels with bovine serum albumin (BSA) standards, ranging from 0.125 to 10 micrograms, were scanned using reflectance densitometry with 127 microns step size in both the x and y directions and a resolution of 200 dots per inch. Densitometric volume was calculated for each protein band from scanner output in the tagged image file format (TIFF) by a customized software package, Scanplot V. 4.05 (Cunningham Engineering). Protein loads between 0.125 and 10.0 micrograms vs. volume were fit by a second-order regression: Volume = -0.58 x protein load2 + 16.82 x protein load + 7.87 (r = 0.991, p < 0.01). The same gels were scanned and quantified using a transmittance laser densitometer; densitometric volumes measured by both systems were highly correlated (r2 = 0.981, p < 0.01). Additional gels of BSA, smooth muscle myosin heavy chain (myosin), and actin displayed linear relationships between protein loads up to 4.0 micrograms and densitometric volume reflecting unique dye binding properties. We conclude that accurate and reproducible quantitative densitometry of SDS-PAGE can be performed using the HP ScanJet Plus scanner and Scanplot software.

Actins↗
PubMed1997

Characterization of molecular determinants of smooth muscle cell heterogeneity.

Broad diversity in contractile and pharmacological properties of different smooth muscles is well recognized. Differences in proliferative capacity, electrophysiology, phenotypic marker protein content, matrix synthesis, and expression of cell-specific transcription factors between individual smooth muscle cells (SMCs) have also been reported. Precise developmental and molecular mechanisms underlying heterogeneity are not known; however, their elucidation is the thrust of much current research involving vascular smooth muscle. In contrast, limited studies of heterogeneity of subtypes of airway SMCs are available. In this report, we review molecular aspects of differentiation that may determine phenotypic heterogeneity of SMCs and also present data from our own studies characterizing heterogeneity in the proliferative capacity and marker protein content of airway SMCs. Using flow cytometry, cell cycle transit was monitored for cultured canine tracheal SMCs. Only 70% of arrested cells responded and traversed the cell cycle when stimulated with 10% fetal bovine serum. Furthermore, heparin inhibited 40% of serum-responsive cells from entering the cell cycle, suggesting that both serum- and heparin-sensitive and -insensitive airway SMCs exist. Flow cytometric analysis of contractile protein and DNA content in freshly dissociated canine tracheal SMCs revealed that diploid (approximately 87%) and tetraploid (approximately 13%) populations exist. Clusters of SMCs having "high" or "low" smooth muscle myosin or alpha-actin content were also discerned, indicating that distinct subtypes of SMCs exist in mature airways. Diversity of SMCs may be a critical factor determining specific responses of smooth muscles to a number of physiological or pathophysiological stimuli that may include, for example, inflammatory mediators in asthmatic airways.

Animals↗