Sp1-dependent activation of HDAC7 is required for platelet-derived growth factor-BB-induced smooth muscle cell differentiation from stem cells

We have previously demonstrated that histone deacetylase 7 (HDAC7) expression and splicing play an important role in smooth muscle cell (SMC) differentiation from embryonic stem (ES) cells, but the molecular mechanisms of increased HDAC7 expression during SMC differentiation are currently unknown. In this study, we found that platelet-derived growth factor-BB (PDGF-BB) induced a 3-fold increase in the transcripts of HDAC7 in differentiating ES cells. Importantly, our data also revealed that PDGF-BB regulated HDAC7 expression not through phosphorylation of HDAC7 but through transcriptional activation. By dissecting its promoters with progressive deletion analysis, we identified the sequence between -343 and -292 bp in the 5'-flanking region of the Hdac7 gene promoter as the minimal PDGF-BB-responsive element, which contains one binding site for the transcription factor, specificity protein 1 (Sp1). Mutation of the Sp1 site within this PDGF-BB-responsive element abolished PDGF-BB-induced HDAC7 activity. PDGF-BB treatment enhanced Sp1 binding to the Hdac7 promoter in differentiated SMCs in vivo as demonstrated by the chromatin immunoprecipitation assay. Moreover, we also demonstrated that knockdown of Sp1 abrogated PDGF-BB-induced HDAC7 up-regulation and SMC differentiation gene expression in differentiating ES cells, although enforced expression of Sp1 alone was sufficient to increase the activity of the Hdac7 promoter and expression levels of SMC differentiation genes. Importantly, we further demonstrated that HDAC7 was required for Sp1-induced SMC differentiation of gene expression. Our data suggest that Sp1 plays an important role in the regulation of Hdac7 gene expression in SMC differentiation from ES cells. These findings provide novel molecular insights into the regulation of HDAC7 and enhance our knowledge in SMC differentiation and vessel formation during embryonic development.

Core binding factor beta-smooth muscle myosin heavy chain chimeric protein involved in acute myeloid leukemia forms unusual nuclear rod-like structures in transformed NIH 3T3 cells.

Patients with the M4Eo subtype of acute myeloid leukemia almost invariably are found to have an inversion of chromosome 16 in their leukemic cells, which results in a gene fusion between the transcription factor called core binding factor beta (CBFbeta) on 16q and a smooth muscle myosin heavy chain (SMMHC) gene on 16p. Subcellular localizations of the wild-type CBFbeta and the CBFbeta-SMMHC fusion protein were determined by immunofluorescence of NIH 3T3 cells that overexpress wild-type or fusion protein. Normal CBFbeta showed an unexpected perinuclear pattern consistent with primary localization in the Golgi complex. The CBFbeta-SMMHC fusion protein had a very different pattern. Nuclear staining included rod-like crystalline structures as long as 11 microm. The heterodimeric partner of CBFbeta, CBFalpha, formed part of this complex. Cytoplasmic staining included stress fibers that colocalized with actin, probably as a consequence of the myosin heavy chain component of the fusion protein. Deletion of different regions of the CBFbeta portion of the fusion protein showed that binding to CBFalpha was not required for nuclear translocation. However, deletion of parts of the SMMHC domain of the fusion protein involved in myosin-mediated filament formation resulted in proteins that did not form rod-like structures. These observations confirm previous indirect evidence that the CBFbeta-SMMHC fusion protein is capable of forming macromolecular nuclear aggregates and suggests possible models for the mechanism of leukemic transformation.

Interleukin-1beta induces human proximal tubule cell injury, alpha-smooth muscle actin expression and fibronectin production

BACKGROUND Tubulointerstitial lesions, characterized by tubular injury, interstitial fibrosis and the appearance of myofibroblasts, are the strongest predictors of the degree and progression of chronic renal failure. These lesions are typically preceded by macrophage infiltration of the tubulointerstitium, raising the possibility that these inflammatory cells promote progressive renal disease through fibrogenic actions on resident tubulointerstitial cells. The aim of the present study, therefore, was to investigate the potentially fibrogenic mechanisms of interleukin-1beta (IL-1beta), a macrophage-derived pro-inflammatory cytokine, on human proximal tubule cells (PTC). METHODS Confluent, quiescent, passage 2 PTC were established in primary culture from histologically normal segments of human renal cortex (N = 11) and then incubated in serum- and hormone-free media supplemented with either IL-1beta (0 to 4 ng/mL) or vehicle (control). RESULTS IL-1beta significantly enhanced fibronectin secretion by up to fourfold in a time- and concentration-dependent fashion. This was accompanied by significant (2.5- to 6-fold) increases in alpha-smooth muscle actin (alpha-SMA) expression, transforming growth factor beta (TGF-beta1) secretion, nitric oxide (NO) production, NO synthase 2 (NOS2) mRNA and lactate dehydrogenase (LDH) release. Cell proliferation was dose-dependently suppressed by IL-1beta. NG-methyl-l-arginine (L-NMMA; 1 mmol/L), a specific inhibitor of NOS, blocked NO production but did not alter basal or IL-1beta-stimulated fibronectin secretion. In contrast, a pan-specific TGF-beta neutralizing antibody significantly blocked the effects of IL-1beta on PTC fibronectin secretion (IL-1beta, 268.1 +/- 30.6 vs. IL-1beta+alphaTGF-beta 157.9 +/- 14.4%, of control values, P < 0.001) and DNA synthesis (IL-1beta 81.0 +/- 6.7% vs. IL-1beta+alphaTGF-beta 93.4 +/- 2.1%, of control values, P < 0.01). CONCLUSION IL-1beta acts on human PTC to suppress cell proliferation, enhance fibronectin production and promote alpha-smooth muscle actin expression. These actions appear to be mediated by a TGF-beta1 dependent mechanism and are independent of nitric oxide release.

Beta-arrestins regulate atherosclerosis and neointimal hyperplasia by controlling smooth muscle cell proliferation and migration.

Atherosclerosis and arterial injury-induced neointimal hyperplasia involve medial smooth muscle cell (SMC) proliferation and migration into the arterial intima. Because many 7-transmembrane and growth factor receptors promote atherosclerosis, we hypothesized that the multifunctional adaptor proteins beta-arrestin1 and -2 might regulate this pathological process. Deficiency of beta-arrestin2 in ldlr(-/-) mice reduced aortic atherosclerosis by 40% and decreased the prevalence of atheroma SMCs by 35%, suggesting that beta-arrestin2 promotes atherosclerosis through effects on SMCs. To test this potential atherogenic mechanism more specifically, we performed carotid endothelial denudation in congenic wild-type, beta-arrestin1(-/-), and beta-arrestin2(-/-) mice. Neointimal hyperplasia was enhanced in beta-arrestin1(-/-) mice, and diminished in beta-arrestin2(-/-) mice. Neointimal cells expressed SMC markers and did not derive from bone marrow progenitors, as demonstrated by bone marrow transplantation with green fluorescent protein-transgenic cells. Moreover, the reduction in neointimal hyperplasia seen in beta-arrestin2(-/-) mice was not altered by transplantation with either wild-type or beta-arrestin2(-/-) bone marrow cells. After carotid injury, medial SMC extracellular signal-regulated kinase activation and proliferation were increased in beta-arrestin1(-/-) and decreased in beta-arrestin2(-/-) mice. Concordantly, thymidine incorporation and extracellular signal-regulated kinase activation and migration evoked by 7-transmembrane receptors were greater than wild type in beta-arrestin1(-/-) SMCs and less in beta-arrestin2(-/-) SMCs. Proliferation was less than wild type in beta-arrestin2(-/-) SMCs but not in beta-arrestin2(-/-) endothelial cells. We conclude that beta-arrestin2 aggravates atherosclerosis through mechanisms involving SMC proliferation and migration and that these SMC activities are regulated reciprocally by beta-arrestin2 and beta-arrestin1. These findings identify inhibition of beta-arrestin2 as a novel therapeutic strategy for combating atherosclerosis and arterial restenosis after angioplasty.

Targeting Gbeta gamma signaling in arterial vascular smooth muscle proliferation: a novel strategy to limit restenosis.

Restenosis continues to be a major problem limiting the effectiveness of revascularization procedures. To date, the roles of heterotrimeric G proteins in the triggering of pathological vascular smooth muscle (VSM) cell proliferation have not been elucidated. betagamma subunits of heterotrimeric G proteins (Gbetagamma) are known to activate mitogen-activated protein (MAP) kinases after stimulation of certain G protein-coupled receptors; however, their relevance in VSM mitogenesis in vitro or in vivo is not known. Using adenoviral-mediated transfer of a transgene encoding a peptide inhibitor of Gbetagamma signaling (betaARKct), we evaluated the role of Gbetagamma in MAP kinase activation and proliferation in response to several mitogens, including serum, in cultured rat VSM cells. Our results include the striking finding that serum-induced proliferation of VSM cells in vitro is mediated largely via Gbetagamma. Furthermore, we studied the effects of in vivo adenoviral-mediated betaARKct gene transfer on VSM intimal hyperplasia in a rat carotid artery restenosis model. Our in vivo results demonstrated that the presence of the betaARKct in injured rat carotid arteries significantly reduced VSM intimal hyperplasia by 70%. Thus, Gbetagamma plays a critical role in physiological VSM proliferation, and targeted Gbetagamma inhibition represents a novel approach for the treatment of pathological conditions such as restenosis.

Autonomic modification of intestinal smooth muscle contractility

Intestinal smooth muscle contracts rhythmically in the absence of nerve and hormonal stimulation because of the activity of pacemaker cells between and within the muscle layers. This means that the autonomic nervous system modifies rather than initiates intestinal contractions. The practical described here gives students an opportunity to observe this spontaneous activity and its modification by agents associated with parasympathetic and sympathetic nerve activity. A section of the rabbit small intestine is suspended in an organ bath, and the use of a pressure transducer and data-acquisition software allows the measurement of tension generated by the smooth muscle of intestinal walls. The application of the parasympathetic neurotransmitter ACh at varying concentrations allows students to observe an increase in intestinal smooth muscle tone with increasing concentrations of this muscarinic receptor agonist. Construction of a concentration-effect curve allows students to calculate an EC50 value for ACh and consider some basic concepts surrounding receptor occupancy and activation. Application of the hormone epinephrine to the precontracted intestine allows students to observe the inhibitory effects associated with sympathetic nerve activation. Introduction of the drug atropine to the preparation before a maximal concentration of ACh is applied allows students to observe the inhibitory effect of a competitive antagonist on the physiological response to a receptor agonist. The final experiment involves the observation of the depolarizing effect of K+ on smooth muscle. Students are also invited to consider why the drugs atropine, codeine, loperamide, and botulinum toxin have medicinal uses in the management of gastrointestinal problems.

Non-steroidal anti-inflammatory drugs (NSAIDs) inhibit vascular smooth muscle cell proliferation via differential effects on the cell cycle

Abnormal vascular smooth muscle cell (VSMC) proliferation plays an important role in the pathogenesis of both atherosclerosis and restenosis. Recent studies suggest that high-dose salicylates, in addition to inhibiting cyclooxygenase activity, exert an antiproliferative effect on VSMC growth both in-vitro and in-vivo. However, whether all non-steroidal anti-inflammatory drugs (NSAIDs) exert similar anti proliferative effects on VSMCs, and do so via a common mechanism of action, remains to be shown. In this study, we demonstrate that the NSAIDs aspirin, sodium salicylate, diclofenac, ibuprofen, indometacin and sulindac induce a dose-dependent inhibition of proliferation in rat A10 VSMCs in the absence of significant cytotoxicity. Flow cytometric analyses showed that exposure of A10 cells to diclofenac, indometacin, ibuprofen and sulindac, in the presence of the mitotic inhibitor, nocodazole, led to a significant G0/G1 arrest. In contrast, the salicylates failed to induce a significant G1 arrest since flow cytometry profiles were not significantly different from control cells. Cyclin A levels were elevated, and hyperphosphorylated p107 was present at significant levels, in salicylate-treated A10 cells, consistent with a post-G1/S block, whereas cyclin A levels were low, and hypophosphorylated p107 was the dominant form, in cells treated with other NSAIDs consistent with a G1 arrest. The ubiquitously expressed cyclin-dependent kinase (CDK) inhibitors, p21 and p27, were increased in all NSAID-treated cells. Our results suggest that diclofenac, indometacin, ibuprofen and sulindac inhibit VSMC proliferation by arresting the cell cycle in the G1 phase, whereas the growth inhibitory effect of salicylates probably affects the late S and/or G2/M phases. Irrespective of mechanism, our results suggest that NSAIDs might be of benefit in the treatment of certain vasculoproliferative disorders.

Inhibition of vascular smooth muscle cell proliferation by non-steroidal anti-inflammatory drugs

Abnormal vascular smooth muscle cell (VSMC) proliferation is known to play an important role in the pathogenesis of atherosclerosis, restenosis and instent stenosis. Recent studies suggest that salicylates, in addition to inhibiting cyclooxygenase activity, exert an antiproliferative effect on VSMC growth both in vitro and in vivo. However, whether all non-steroidal anti-inflammatory drugs (NSAID) exert similar antiproliferative effects on VSMCs, and do so via a common mechanism of action, remains unknown. In the present study, we demonstrated that the NSAIDs, aspirin, ibuprofen and sulindac induced a dose-dependent inhibition of proliferation in rat A10 VSMCs (IC50 = 1666 mumol/L, 937 mumol/L and 520 mumol/L, respectively). These drugs did not show significant cytotoxic effects as determined by LDH release assay, even at the highest concentrations tested (aspirin, 5000 mumol/L; ibuprofen, 2500 mumol/L; and sulindac, 1000 mumol/L). Flow cytometric analyses showed that a 48 h exposure of A10 VSMCs to ibuprofen (1000 mumol/L) and sulindac (750 mumol/L) led to a significant G1 arrest (from 68.7 +/- 2.0% of cells in G1 to 76.6 +/- 2.2% and 75.8 +/- 2.2%, respectively, p < 0.05). In contrast, aspirin (2500 mumol/L) failed to induce a significant G1 arrest (68.1 +/- 5.2%). Clearer evidence of a G1 block was obtained by treatment of cells with the mitotic inhibitor, nocodazole (40 ng/ml), for the final 24 h of the experiment. Under these conditions, aspirin still failed to induce a G1 arrest (from 25.9 +/- 10.9% of cells in G1 to 19.6 +/- 2.3%) whereas ibuprofen and sulindac led to a significant accumulation of cells in G1(51.8% +/- 17.2% and 54.1% +/- 10.6%, respectively, p < 0.05). These results indicate that ibuprofen and sulindac inhibit VSMC proliferation by arresting the cell cycle in the G1 phase whereas the effect of aspirin appears to be independent of any special phase of the cell cycle. Irrespective of mechanism, our results suggest that NSAIDs might be of benefit to the treatment of vascular proliferative disorders.

Cell blocks allow reliable evaluation of expression of basal (CK5/6) and luminal (CK8/18) cytokeratins and smooth muscle actin (SMA) in breast carcinoma

Objective:Gene expression studies have revealed several molecular subtypes of breast carcinoma with distinct clinical and biological behaviours. DNA microarray studies correlated with immunohistochemical profiling of breast carcinomas using cytokeratin (CK) markers, Her2/neu, oestrogen receptor (ER), and basal myoepithelial cell markers have identified five breast tumour subtypes: (i) luminal A (ER+; Her2/neu-), (ii) luminal B (ER+; Her2/neu+), (iii) Her2 overexpression (ER-; Her2/neu+), (iv) basal-like (ER-; Her2/neu-, CK5/6 and 14+), and (v) negative for all markers. Luminal carcinomas express cytokeratins in a luminal pattern (CK8/18), and the basal-like type expresses CK5/6 and CK14 or basal epithelial cell markers. CK5/6, CK8/18, and smooth muscle actin (SMA) expression were assessed in cell blocks and compared with expression in surgical specimens.Methods:Sixty-two cases of breast carcinoma diagnosed by fine needle aspiration cytology with cell blocks and available surgical specimens were included. Cell blocks containing at least 10 high-power fields each with at least 10 tumour cells and surgical specimens were immunostained for CK5/6, CK8/18 and SMA.Results:Percentage sensitivity, specificity, positive predictive value, negative predictive value and accuracy were, respectively, 77, 100, 100, 92 and 94 for CK5/6; 98, 66, 96, 80 and 95 for CK8/18; and 92, 96, 85, 98 and 95 for SMA.Conclusion:The identification of CK5/6, CK8/18 and SMA by immunohistochemistry in cell blocks can be a reliable method that yields results close to those obtained in surgical specimens, and can contribute to the classification of breast carcinomas with luminal and basal expression patterns, providing helpful information in the choice of treatment and in the evaluation of prognostic and predictive factors.

Mitochondrial alterations and apoptosis in smooth muscle from aged rats

We studied changes in mitochondrial morphology and function in the smooth muscle of rat colon. Under confocal microscopy, tissues loaded with potentiometric dye displayed rapid and spontaneous depolarization. Cyclosporin A (CsA), inhibitor of the permeability transition pore (PTP), caused an increase in mitochondrial membrane potential (DeltaPsi(m)) in tissues from adult young animals. In aged rats these changes were not observed. This suggests that physiological activation of PTP in aged rats is reduced. Electron microscopy showed alterations of the mitochondrial ultrastructure in tissues from aged rats involving a decreased definition of the cristae and fragmentation of the mitochondrial membranes. We also detected an increase in apoptotic cells in the smooth muscle from aged animals. Our results show that the aging process changes PTP activity, the ability to maintain DeltaPsi(m) and mitochondrial morphology. It is suggested that these can be associated with mitochondrial damage and cell death. (C) 2004 Elsevier B.V. All rights reserved.

Mirabegron relaxes urethral smooth muscle by a dual mechanism involving β3-adrenoceptor activation and α1-adrenoceptor blockade

Mirabegron is the first β3-adrenoceptor (AR) agonist approved for treatment of overactive bladder syndrome (OAB). This study aimed to investigate the effects of β3-adrenoceptor (AR) agonist mirabegron in mouse urethra. The possibility that mirabegron exerts α1-AR antagonism was also tested in rat smooth muscle preparations presenting α1A- (vas deferens and prostate), α1D- (aorta) and α1B-AR (spleen). Functional assays were carried out in mouse and rat isolated tissues. Competition assays for the specific binding of [(3) H]Prazosin to membrane preparations of HEK 293 cells expressing each of the human α1-ARs, as well as β-AR mRNA expression and cyclic AMP measurements in mouse urethra were performed. Mirabegron produced concentration-dependent urethral relaxations that were right shifted by the selective β3-AR antagonist L 748,337, but unaffected by β1- and β2-AR antagonists (atenolol and ICI 118,551, respectively). Mirabegron-induced relaxations were enhanced by the phosphodiesterase-4 inhibitor rolipram, and this agonist stimulated cAMP synthesis. Mirabegron also produced rightward shifts in urethral contractions induced by the α1-AR agonist phenylephrine. Schild regression analysis revealed that mirabegron behaves as a competitive antagonist of α1-AR in urethra, vas deferens and prostate (α1A-AR, pA2  ≅ 5.6) and aorta (α1D-AR, pA2  ≅ 5.4), but not in spleen (α1B-AR). The affinities estimated for mirabegron in functional assays were consistent with those estimated in radioligand binding with human recombinant α1A- and α1D-ARs (pKi ≅ 6.0). The effects of mirabegron in urethral smooth muscle are the result of β3-AR agonism together with α1A / α1D-AR antagonism.

Extracellular matrix in airway smooth muscle is associated with dynamics of airway function in asthma

Background: Altered deposition of extracellular matrix (ECM) in the airway smooth muscle (ASM) layer as observed in asthma may influence ASM mechanical properties. We hypothesized that ECM in ASM is associated with airway function in asthma. First, we investigated the difference in ECM expression in ASM between asthma and controls. Second, we examined whether ECM expression is associated with bronchoconstriction and bronchodilation in vivo. Methods: Our cross-sectional study comprised 19 atopic mild asthma patients, 15 atopic and 12 nonatopic healthy subjects. Spirometry, methacholine responsiveness, deep-breath-induced bronchodilation (Delta R-rs) and bronchoscopy with endobronchial biopsies were performed. Positive staining of elastin, collagen I, III and IV, decorin, versican, fibronectin, laminin and tenascin in ASM was quantified as fractional area and mean density. Data were analysed using Pearson's or Spearman's correlation coefficient. Results: Extracellular matrix expression in ASM was not different between asthma and controls. In asthmatics, fractional area and mean density of collagen I and III were correlated with methacholine dose-response slope and DRrs, respectively (r = 0.71, P < 0.01; r = 0.60, P = 0.02). Furthermore, ASM collagen III and laminin in asthma were correlated with FEV1 reversibility (r = -0.65, P = 0.01; r = -0.54, P = 0.04). Conclusion: In asthma, ECM in ASM is related to the dynamics of airway function in the absence of differences in ECM expression between asthma and controls. This indicates that the ASM layer in its full composition is a major structural component in determining variable airways obstruction in asthma.

Increased fibroblast telomerase expression precedes myofibroblast alpha-smooth muscle actin expression in idiopathic pulmonary fibrosis

OBJECTIVE: This study sought to identify the relationship between fibroblast telomerase expression, myofibroblasts, and telomerase-mediated regulatory signals in idiopathic pulmonary fibrosis. METHODS: Thirty-four surgical lung biopsies, which had been obtained from patients with idiopathic pulmonary fibrosis and histologically classified as usual interstitial pneumonia, were examined. Immunohistochemistry was used to evaluate fibroblast telomerase expression, myofibroblast alpha-smooth muscle actin expression and the tissue expression of interleukin-4, transforming growth factor-beta, and basic fibroblast growth factor. The point-counting technique was used to quantify the expression of these markers in unaffected, collapsed, mural fibrosis, and honeycombing areas. The results were correlated to patient survival. RESULTS: Fibroblast telomerase expression and basic fibroblast growth factor tissue expression were higher in collapsed areas, whereas myofibroblast expression and interleukine-4 tissue expression were higher in areas of mural fibrosis. Transforming growth factor-beta expression was higher in collapsed, mural fibrosis and honeycombing areas in comparison to unaffected areas. Positive correlations were found between basic fibroblast growth factor tissue expression and fibroblast telomerase expression and between interleukin-4 tissue expression and myofibroblast alpha-smooth muscle actin expression. Negative correlations were observed between interleukin-4 expression and basic fibroblast growth factor tissue expression in areas of mural fibrosis. Myofibroblast alpha-smooth muscle actin expression and interleukin-4 tissue expression in areas of mural fibrosis were negatively associated with patient survival. CONCLUSION: Fibroblast telomerase expression is higher in areas of early remodeling in lung tissues demonstrating typical interstitial pneumonia, whereas myofibroblast alpha-smooth muscle actin expression predominates in areas of late remodeling. These events seem to be regulated by basic fibroblast growth factor and interleukin-4 tissue expression, respectively.

Mechanical Alterations in airway smooth muscle after interaction with indoor pollutant – A mathematical model.

The viscoelasticity of mammalian lung is determined by the mechanical properties and structural regulation of the airway smooth muscle (ASM). The exposure to polluted air may deteriorate these properties with harmful consequences to individual health. Formaldehyde (FA) is an important indoor pollutant found among volatile organic compounds. This pollutant permeates through the smooth muscle tissue forming covalent bonds between proteins in the extracellular matrix and intracellular protein structure changing mechanical properties of ASM and inducing asthma symptoms, such as airway hyperresponsiveness, even at low concentrations. In the experimental scenario, the mechanical effect of FA is the stiffening of the tissue, but the mechanism behind this effect is not fully understood. Thus, the aim of this study is to reproduce the mechanical behavior of the ASM, such as contraction and stretching, under FA action or not. For this, it was created a two-dimensional viscoelastic network model based on Voronoi tessellation solved using Runge-Kutta method of fourth order. The equilibrium configuration was reached when the forces in different parts of the network were equal. This model simulates the mechanical behavior of ASM through of a network of dashpots and springs. This dashpot-spring mechanical coupling mimics the composition of the actomyosin machinery of ASM through the contraction of springs to a minimum length. We hypothesized that formation of covalent bonds, due to the FA action, can be represented in the model by a simple change in the elastic constant of the springs, while the action of methacholine (MCh) reduce the equilibrium length of the spring. A sigmoid curve of tension as a function of MCh doses was obtained, showing increased tension when the muscle strip was exposed to FA. Our simulations suggest that FA, at a concentration of 0.1 ppm, can affect the elastic properties of the smooth muscle ¯bers by a factor of 120%. We also analyze the dynamic mechanical properties, observing the viscous and elastic behavior of the network. Finally, the proposed model, although simple, incorporates the phenomenology of both MCh and FA and reproduces experimental results observed with in vitro exposure of smooth muscle to FA. Thus, this new mechanical approach incorporates several well know features of the contractile system of the cells in a tissue level model. The model can also be used in different biological scales.

Mechanical alterations in airway smooth muscle after interaction with indoor pollutant - A mathematical model

The viscoelasticity of mammalian lung is determined by the mechanical properties and structural regulation of the airway smooth muscle (ASM). The exposure to polluted air may deteriorate these properties with harmful consequences to individual health. Formaldehyde (FA) is an important indoor pollutant found among volatile organic compounds. This pollutant permeates through the smooth muscle tissue forming covalent bonds between proteins in the extracellular matrix and intracellular protein structure changing mechanical properties of ASM and inducing asthma symptoms, such as airway hyperresponsiveness, even at low concentrations. In the experimental scenario, the mechanical effect of FA is the stiffening of the tissue, but the mechanism behind this effect is not fully w1derstood. Thus, the aim of this study is to reproduce the mechanical behavior of the ASM, such as contraction and stretching, under FA action or not. For this, it was created a two-dimensional viscoelastic network model based on Voronoi tessellation solved using Runge-Kutta method of fourth order. The equilibrium configuration was reached when the forces in different parts of the network were equal. This model simulates the mechanical behavior of ASM through of a network of dashpots and springs. This dashpot-spring mechanical coupling mimics the composition of the actomyosin machinery of ASM through the contraction of springs to a minimum length. We hypothesized that formation of covalent bonds, due to the FA action, can be represented in the model by a simple change in the elastic constant of the springs, while the action of methacholinc (MCh) reduce the equilibrium length of the spring. A sigmoid curve of tension as a function of MCh doses was obtained, showing increased tension when the muscle strip was exposed to FA. Our simulations suggest that FA, at a concentration of 0.1 ppm, can affect the elastic properties of the smooth muscle fibers by a factor of 120%. We also analyze the dynamic mechanical properties, observing the viscous and elastic behavior of the network. Finally, the proposed model, although simple, ir1corporates the phenomenology of both MCh and FA and reproduces experirnental results observed with ir1 vitro exposure of smooth muscle to .FA. Thus, this new mechanical approach incorporates several well know features of the contractile system of the cells ir1 a tissue level model. The model can also be used in different biological scales.