Orchestrating Airway Smooth Muscle Cell Migration: GMFγ Phosphorylation Is the Key.
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ABSTRACT:
SUBMITTER: Ihrie MD
PROVIDER: S-EPMC6670032 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Airway smooth muscle cells require coordinated protrusion and focal adhesion dynamics to migrate properly. However, the signaling cascades that connect these two processes remain incompletely understood. Glia maturation factor (GMF)-γ has been implicated in inducing actin debranching and inhibiting nucleation. In this study, we discovered that GMFγ phosphorylation at Y104 regulates human airway smooth muscle cell migration. Using high-resolution microscopy coupled with three-dimensional object-based quantitative image analysis software, Imaris 9.2.0, phosphomimetic mutant, Y104D-GMFγ, was enriched at nascent adhesions along the leading edge where it recruited activated neural Wiskott-Aldrich syndrome protein (N-WASP; pY256) to promote actin-branch formation, which enhanced lamellipodial dynamics and limited the growth of focal adhesions. Unexpectedly, we found that nonphosphorylated mutant, Y104F-GMFγ, was enriched in growing adhesions where it promoted a linear branch organization and focal adhesion clustering, and recruited zyxin to increase maturation, thus inhibiting lamellipodial dynamics and cell migration. The localization of GMFγ between the leading edge and focal adhesions was dependent upon myosin activity. Furthermore, c-Abl tyrosine kinase regulated the GMFγ phosphorylation-dependent processes. Together, these results unveil the importance of GMFγ phosphorylation in coordinating lamellipodial and focal adhesion dynamics to regulate cell migration.
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Project description:Airway remodelling is an important feature of asthma pathogenesis. A key structural change inherent in airway remodelling is increased airway smooth muscle mass. There is emerging evidence to suggest that the migration of airway smooth muscle cells may contribute to cellular hyperplasia, and thus increased airway smooth muscle mass. The precise source of these cells remains unknown. Increased airway smooth muscle mass may be collectively due to airway infiltration of myofibroblasts, neighbouring airway smooth muscle cells in the bundle, or circulating hemopoietic progenitor cells. However, the relative contribution of each cell type is not well understood. In addition, although many studies have identified pro and anti-migratory agents of airway smooth muscle cells, whether these agents can impact airway remodelling in the context of human asthma, remains to be elucidated. As such, further research is required to determine the exact mechanism behind airway smooth muscle cell migration within the airways, how much this contributes to airway smooth muscle mass in asthma, and whether attenuating this migration may provide a therapeutic avenue for asthma. In this review article, we will discuss the current evidence with respect to the regulation of airway smooth muscle cell migration in asthma.
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Project description:Severe asthma manifests as airway remodeling and irreversible airway obstruction, in part because of the proliferation and migration of human airway smooth muscle (HASM) cells. We previously reported that cyclic adenosine monophosphate-mobilizing agents, including ?(2)-adrenergic receptor (?(2)AR) agonists, which are mainstay of asthma therapy, and prostaglandin E2 (PGE2), inhibit the migration of HASM cells, although the mechanism for this migration remains unknown. Vasodilator-stimulated phosphoprotein (VASP), an anticapping protein, modulates the formation of actin stress fibers during cell motility, and is negatively regulated by protein kinase A (PKA)-specific inhibitory phosphorylation at serine 157 (Ser157). Here, we show that treatment with ?(2)AR agonists and PGE2 induces the PKA-dependent phosphorylation of VASP and inhibits the migration of HASM cells. The stable expression of PKA inhibitory peptide and the small interfering (si) RNA-induced depletion of VASP abolish the inhibitory effects of albuterol and PGE2 on the migration of HASM cells. Importantly, prolonged treatment with albuterol prevents the agonist-induced phosphorylation of VASP at Ser157, and reverses the inhibitory effects of albuterol and formoterol, but not PGE2, on the basal and PDGF-induced migration of HASM cells. Collectively, our data demonstrate that ?(2)AR agonists selectively inhibit the migration of HASM cells via a ?(2)AR/PKA/VASP signaling pathway, and that prolonged treatment with albuterol abolishes the inhibitory effect of ?-agonists on the phosphorylation of VASP and migration of HASM cells because of ?(2)AR desensitization.
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Project description:During organism aging, the process of cellular senescence is triggered by critical stressors such as DNA damage, oncogenes, oxidative stress, and telomere erosion, and vascular cells are not exempted. Senescent cells stop proliferating but remain metabolically active producing pro-inflammatory signals in the environment collectively named senescence-associated secretory phenotype (SASP) that contribute to the amplification of the response to the neighbor and distant cells. Although the shift toward senescence is protective against tumors and needed during wound healing, the accumulation of senescent cells during aging due to an impairment of the immune system deputed to their clearance, can predispose to diseases of the cardiovascular system such as atherosclerosis. In this short review, we describe the main features of senescence of endothelial and smooth muscle cells and focus on the role non-coding RNAs of the microRNAs class in controlling this process. Finally, we discuss the potential of new strategies based on senescence removal in counteracting vascular disease burden.
| S-EPMC7439843 | biostudies-literature
Project description:AimsFocal adhesion kinase (FAK) and its autonomously expressed, C-terminal inhibitor FAK-related non-kinase (FRNK), are important regulators of vascular smooth muscle cell (VSMC) spreading and migration. However, the mechanisms of FRNK-mediated inhibition of FAK-dependent signalling are not fully defined. The aim of this study was to determine the potential role of FRNK tyrosine phosphorylation in regulating these processes.Methods and resultsRat carotid arteries were balloon-injured and FAK and FRNK expression and phosphorylation were examined by immunocytochemistry, immunoprecipitation, and western blotting with total and phosphospecific antibodies. FAK and FRNK expression increased four- and nine-fold, respectively, in alpha-smooth muscle actin-positive VSMCs of injured arteries when compared with contralateral control arteries, and the upregulated FRNK was phosphorylated at residues Y168 and Y232. In A7r5 cells (an embryonic rat VSMC line), endogenously expressed FRNK was also phosphorylated at Y168 and Y232 under basal conditions, and Y168/Y232 phosphorylation increased in response to angiotensin II treatment. When overexpressed in A7r5 cells and adult rat aortic smooth muscle cells (RASM), wild-type (wt) GFP-tagged FRNK was also phosphorylated at residues Y168 and Y232, and GFP-wtFRNK inhibited cell spreading and migration. Mutation of GFP-FRNK at Y168 (GFP-Y168F-FRNK) abrogated FRNK-mediated inhibition of cell spreading and migration, but did not affect its localization in VSMC focal adhesions or its ability to inhibit FAK tyrosine phosphorylation.ConclusionPhosphorylation of Y168 on FRNK may represent a novel mechanism by which FRNK inhibits cell spreading and migration in VSMCs.
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Project description:Airway remodeling due to increased airway smooth muscle (ASM) mass, likely due to enhanced migration and proliferation, has been shown to be highly associated with decline in lung function in asthma. Thymic stromal lymphopoietin (TSLP) is an IL-7-like, pro-allergic cytokine that has been shown to be necessary and sufficient for the development of allergic asthma. Human ASM (HASM) cells express TSLP receptor (TSLPR), the activation of which leads to enhanced release of proinflammatory mediators such as IL-6, CCL11/eotaxin-1, and CXCL8/IL-8. We show here that TSLP induces HASM cell migration through STAT3 activation since lentiviral-shRNA inhibition of STAT3 abrogated the TSLP-induced cell migration. Moreover, TSLP induced multiple cytoskeleton changes in HASM cells such as actin polymerization, cell polarization, and activation of small GTPase Rac1. Collectively, our data suggest a pro-migratory function of TSLP in ASM remodeling and provides better rationale for targeting TSLP/TSLPR pathway for therapeutic approaches in allergic asthma.
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Project description:The chronic phase of pulmonary arterial hypertension (PAH) is associated with vascular remodeling, especially thickening of the smooth muscle layer of large pulmonary arteries and muscularization of small pulmonary vessels, which normally have no associated smooth muscle. Serotonin (5-hydroxytryptamine, 5-HT) has been shown to induce proliferation and hypertrophy of pulmonary artery smooth muscle cells (PASMC), and may be important for in vivo pulmonary vascular remodeling. Here, we show that 5-HT stimulates migration of pulmonary artery PASMC. Treatment with 5-HT for 16h increased migration of PASMC up to four-fold as monitored in a modified Boyden chamber assay. Increased migratory responses were associated with cellular morphological changes and reorganization of the actin cytoskeleton. 5-HT-induced alterations in morphology were previously shown in our laboratory to require cAMP [Lee SL, Fanburg BL. Serotonin produces a configurational change of cultured smooth muscle cells that is associated with elevation of intracellular cAMP. J Cell Phys 1992;150(2):396-405], and the 5-HT4 receptor was pharmacologically determined to be the primary activator of cAMP in bovine PASMC [Becker BN, Gettys TW, Middleton JP, Olsen CL, Albers FJ, Lee SL, et al. 8-Hydroxy-2-(di-n-propylamino)tetralin-responsive 5-hydroxytryptamine4-like receptor expressed in bovine pulmonary artery smooth muscle cells. Mol Pharmacol 1992;42(5):817-25]. We examined the role of the 5-HT4 receptor and cAMP in 5-HT-induced bovine PASMC migration. PASMC express 5-HT4 receptor mRNA, and a 5-HT4 receptor antagonist and a cAMP antagonist completely blocked 5-HT-induced cellular migration. Consistent with our previous report that a cAMP-dependent Cl(-) channel is required for 5-HT-induced morphological changes in PASMC, phenylanthranilic acid, a Cl(-) channel blocker, inhibited actin cytoskeletal reorganization and migration produced by 5-HT. We conclude that 5-HT stimulates PASMC migration and associated cytoskeletal reorganization through the 5-HT4 receptor and cAMP activation of a chloride channel.
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Project description:Airway smooth muscle cell migration plays a role in the progression of airway remodeling, a hallmark of allergic asthma. However, the mechanisms that regulate cell migration are not yet entirely understood. Nestin is a class VI intermediate filament protein that is involved in the proliferation/regeneration of neurons, cancer cells, and skeletal muscle. Its role in cell migration is not fully understood. Here, nestin knockdown (KD) inhibited the migration of human airway smooth muscle cells. Using confocal microscopy and the Imaris software, we found that nestin KD attenuated focal adhesion sizes during cell spreading. Moreover, polo-like kinase 1 (Plk1) and vimentin phosphorylation at Ser-56 have been previously shown to affect focal adhesion assembly. Here, nestin KD reduced Plk1 phosphorylation at Thr-210 (an indication of Plk1 activation), vimentin phosphorylation at Ser-56, the contacts of vimentin filaments to paxillin, and the morphology of focal adhesions. Moreover, the expression of vimentin phosphorylation-mimic mutant S56D (aspartic acid substitution at Ser-56) rescued the migration, vimentin reorganization, and focal adhesion size of nestin KD cells. Together, our results suggest that nestin promotes smooth muscle cell migration. Mechanistically, nestin regulates Plk1 phosphorylation, which mediates vimenitn phosphorylation, the connection of vimentin filaments with paxillin, and focal adhesion assembly.
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Project description:Abnormal migration and proliferation of airway smooth muscle cells (ASMCs) in the airway cause airway wall thickening, which is strongly related with the development of airway remodeling in asthma. Clara cell 10?kDa protein (CC10), which is secreted by the epithelial clara cells of the pulmonary airways, plays an important role in the regulation of immunological and inflammatory processes. Previous studies suggested that CC10 protein had great protective effects against inflammation in asthma. However, the effects of CC10 protein on ASMCs migration and proliferation in airway remodeling were poorly understood. In this study, we constructed the pET-22b-CC10 recombinant plasmid, induced expression and purified the recombinant rat CC10 protein from E. coli by Ni(2+) affinity chromatography and ion exchange chromatography purification. We investigated the effect of recombinant rat CC10 protein on platelet-derived growth factor (PDGF)-BB-induced ASMCs proliferation and migration. Our results demonstrated that the recombinant CC10 protein could inhibit PDGF-BB-induced cell viability, proliferation and migration. Western blot analysis showed that PDGF-BB-induced activation of cyclin D1 was inhibited by CC10. These findings implicated that CC10 could inhibit increased ASMCs proliferation, and migration induced by PDGF-BB, and this suppression effect might be associated with inhibition of cyclin D1 expression, which might offer hope for the future treatment of airway remodeling.
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Project description:Abnormal vascular smooth muscle cell (SMC) proliferation and migration contribute to the development of restenosis after percutaneous transluminal coronary angioplasty and accelerated arteriopathy after cardiac transplantation. Previously, we reported that the macrolide antibiotic rapamycin, but not the related compound FK506, inhibits both human and rat aortic SMC proliferation in vitro by inhibiting cell cycle-dependent kinases and delaying phosphorylation of retinoblastoma protein (Marx, S.O., T. Jayaraman, L.O. Go, and A.R. Marks. 1995. Circ. Res. 362:801). In the present study the effects of rapamycin on SMC migration were assayed in vitro using a modified Boyden chamber and in vivo using a porcine aortic SMC explant model. Pretreatment with rapamycin (2 ng/ml) for 48 h inhibited PDGF-induced migration (PDGF BB homodimer; 20 ng/ml) in cultured rat and human SMC (n = 10; P < 0.0001), whereas FK506 had no significant effect on migration. Rapamycin administered orally (1 mg/kg per d for 7 d) significantly inhibited porcine aortic SMC migration compared with control (n = 15; P < 0.0001). Thus, in addition to being a potent immunosuppressant and antiproliferative, rapamycin also inhibits SMC migration.
| S-EPMC507677 | biostudies-other