scholarly journals FFAR1 activation attenuates histamine-induced myosin light chain phosphorylation and cortical tension development in human airway smooth muscle cells

2020 ◽  
Vol 21 (1) ◽  
Author(s):  
Shengjie Xu ◽  
Anthony Schwab ◽  
Nikhil Karmacharya ◽  
Gaoyuan Cao ◽  
Joanna Woo ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Airway Smooth Muscle ◽  
Light Chain ◽  
Myosin Light Chain ◽  
Airway Hyperreactivity ◽  
Human Airway Smooth Muscle ◽  
Tension Development ◽  
Cortical Tension ◽  
Human Airway ◽  
Mlc Phosphorylation

Abstract Background Activation of free fatty acid receptors (FFAR1 and FFAR4) which are G protein-coupled receptors (GPCRs) with established (patho)physiological roles in a variety of obesity-related disorders, induce human airway smooth muscle (HASM) cell proliferation and shortening. We reported amplified agonist-induced cell shortening in HASM cells obtained from obese lung donors. We hypothesized that FFAR1 modulate excitation–contraction (EC) coupling in HASM cells and play a role in obesity-associated airway hyperresponsiveness. Methods In HASM cells pre-treated (30 min) with FFAR1 agonists TAK875 and GW9508, we measured histamine-induced Ca2+ mobilization, myosin light chain (MLC) phosphorylation, and cortical tension development with magnetic twisting cytometry (MTC). Phosphorylation of MLC phosphatase and Akt also were determined in the presence of the FFAR1 agonists or vehicle. In addition, the effects of TAK875 on MLC phosphorylation were measured in HASM cells desensitized to β2AR agonists by overnight salmeterol treatment. The inhibitory effect of TAK875 on MLC phosphorylation was compared between HASM cells from age and sex-matched non-obese and obese human lung donors. The mean measurements were compared using One-Way ANOVA with Dunnett’s test for multiple group comparisons or Student’s t-test two-group comparison. For cortical tension measurements by magnetic twisted cytometry, mixed effect model using SAS V.9.2 was applied. Means were considered significant when p ≤ 0.05. Results Unexpectedly, we found that TAK875, a synthetic FFAR1 agonist, attenuated histamine-induced MLC phosphorylation and cortical tension development in HASM cells. These physiological outcomes were unassociated with changes in histamine-evoked Ca2+ flux, protein kinase B (AKT) activation, or MLC phosphatase inhibition. Of note, TAK875-mediated inhibition of MLC phosphorylation was maintained in β2AR-desensitized HASM cells and across obese and non-obese donor-derived HASM cells. Conclusions Taken together, our findings identified the FFAR1 agonist TAK875 as a novel bronchoprotective agent that warrants further investigation to treat difficult-to-control asthma and/or airway hyperreactivity in obesity.

scholarly journals Obesity increases airway smooth muscle responses to contractile agonists

2018 ◽  
Vol 315 (5) ◽  
pp. L673-L681 ◽  
Author(s):  
Sarah Orfanos ◽  
Joseph Jude ◽  
Brian T. Deeney ◽  
Gaoyuan Cao ◽  
Deepa Rastogi ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Airway Smooth Muscle ◽  
Calcium Mobilization ◽  
Health Concern ◽  
Airway Smooth Muscle Cells ◽  
Human Airway Smooth Muscle ◽  
Cell Shortening ◽  
Human Airway ◽  
Obese Subjects ◽  
Mlc Phosphorylation

The asthma-obesity syndrome represents a major public health concern that disproportionately contributes to asthma severity and induces insensitivity to therapy. To date, no study has shown an intrinsic difference between human airway smooth muscle (HASM) cells derived from nonobese subjects and those derived from obese subjects. The objective of this study was to address whether there is a greater response to agonist-induced calcium mobilization, phosphorylation of myosin light chain (MLC), and greater shortening in HASM cells derived from obese subjects. HASM cells derived from nonobese and obese subjects were age and sex matched. Phosphorylation of MLC was measured after having been stimulated by carbachol. Carbachol- or histamine-induced mobilization of calcium and cell shortening were assessed in HASM cells derived from nonobese and obese donors. Agonist-induced MLC phosphorylation, mobilization of calcium, and cell shortening were greater in obese compared with non-obese-derived HASM cells. The MLC response was comparable in HASM cells derived from obese nonasthma and nonobese fatal asthma subjects. HASM cells derived from obese female subjects were more responsive to carbachol than HASM cells derived from obese male subjects. Insulin pretreatment had little effect on these responses. Our results show an increase in agonist-induced calcium mobilization associated with an increase in MLC phosphorylation and an increase in ASM cell shortening in favor of agonist-induced hyperresponsiveness in HASM cells derived from obese subjects. Our studies suggest that obesity induces a retained phenotype of hyperresponsiveness in cultured human airway smooth muscle cells.

NMDA receptors mediate contractile responses in human airway smooth muscle cells

2015 ◽  
Vol 308 (12) ◽  
pp. L1253-L1264 ◽  
Author(s):  
Vidyanand Anaparti ◽  
Ramses Ilarraza ◽  
Kanami Orihara ◽  
Gerald L. Stelmack ◽  
Oluwaseun O. Ojo ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Airway Inflammation ◽  
Airway Smooth Muscle ◽  
Airway Hyperreactivity ◽  
Glutamate Toxicity ◽  
Airway Smooth Muscle Cells ◽  
Collagen Gels ◽  
Human Airway Smooth Muscle ◽  
Contractile Responses ◽  
Human Airway

Human airway smooth muscle (HASM) exhibits enhanced contractility in asthma. Inflammation is associated with airway hypercontractility, but factors that underpin these features are not fully elucidated. Glutamate toxicity associated with increased plasma glutamate concentrations was observed in airway inflammation, suggesting that multisubunit glutamate receptors, N-methyl- d-aspartate receptors (NMDA-R) contribute to airway hyperreactivity. We tested the hypothesis that HASM expresses NMDA-R subunits that can form functional receptors to mediate contractile responses to specific extracellular ligands. In cultured HASM cells, we measured NMDA-R subunit mRNA and protein abundance by quantitative PCR, immunoblotting, flow cytometry, and epifluorescence immunocytochemistry. We measured mRNA for a number of NMDA-R subunits, including the obligatory NR1 subunit, which we confirmed to be present as a protein. In vitro and ex vivo functional NMDA-R activation in HASM cells was measured using intracellular calcium flux (fura-2 AM), collagen gel contraction assays, and murine thin-cut lung slices (TCLS). NMDA, a pharmacological glutamate analog, induced cytosolic calcium mobilization in cultured HASM cells. We detected three different temporal patterns of calcium response, suggesting the presence of heterogeneous myocyte subpopulations. NMDA-R activation also induced airway contraction in murine TCLS and soft collagen gels seeded with HASM cells. Responses in cells, lung slices, and collagen gels were mediated by NMDA-R, as they could be blocked by (2R)-amino-5-phosphonopentanoate, a specific NMDA-R inhibitor. In summary, we reveal the presence of NMDA-R in HASM that mediate contractile responses via glutamatergic mechanisms. These findings suggest that accumulation of glutamate-like ligands for NMDA-R associated with airway inflammation contributes directly to airway hyperreactivity.

scholarly journals T-bet is induced by interferon-γ to mediate chemokine secretion and migration in human airway smooth muscle cells

2011 ◽  
Vol 300 (4) ◽  
pp. L633-L641 ◽  
Author(s):  
Cherie A. Singer
Keyword(s):  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Airway Smooth Muscle ◽  
Muscle Cells ◽  
Airway Disease ◽  
Airway Hyperreactivity ◽  
Airway Smooth Muscle Cells ◽  
Human Airway Smooth Muscle ◽  
Human Airway ◽  
Inflammatory Airway Disease

An inappropriate balance between T-helper (Th)1 and Th2 cytokine production underlies inflammatory changes that result in airway disease. Expression of the T-box transcription factor T-bet regulates differentiation of Th cells and production of Th1 cytokines, particularly IFNγ. T-bet-deficient mice develop airway hyperreactivity, undergo airway remodeling, and exhibit defects in IFNγ production while overproducing Th2 cytokines. T-bet is also reduced in the airways of asthmatic patients, suggesting loss of T-bet expression or activity promotes development of inflammatory airway disease. We present novel data demonstrating T-bet expression is induced in human airway smooth muscle cells (ASMC) by IFNγ. This IFNγ-stimulated expression of T-bet is dependent on signaling through JAK2 and signal transducers and activators of transcription 1 (STAT1) and activates T-bet-dependent DNA binding activity. Expression of T-bet stimulates IFNγ-stimulated IFNγ expression, secretion, and promoter activity, while inhibiting IFNγ-stimulated release of chemokines including monocyte chemoattractant protein (MCP)-1/CCL2, regulated on activation normal T-expressed and secreted (RANTES)/CCL5, and eotaxin/CCL11. This is accompanied by changes in expression of the chemokine receptors CCR3 and IL12Rβ2 and TNFα. T-bet expression also reduces chemotactic migration of ASMC in response to serum and PDGF, which contributes to airway hyperplasia. These results are the first to identify T-bet expression and activity in a structural cell of the lung and may provide new insights into therapeutic targets for inflammatory airway disease.

PGE2 release by bradykinin in human airway smooth muscle cells: involvement of cyclooxygenase-2 induction

1997 ◽  
Vol 273 (6) ◽  
pp. L1132-L1140 ◽  
Author(s):  
Linhua Pang ◽  
Alan J. Knox
Keyword(s):  
Arachidonic Acid ◽  
Smooth Muscle ◽  
Airway Smooth Muscle ◽  
Arachidonic Acid Release ◽  
Human Airway Smooth Muscle ◽  
Short Term ◽  
Human Airway ◽  
Acid Release ◽  
Cox 2

Prostanoids may be involved in bradykinin (BK)-induced bronchoconstriction in asthma. We investigated whether cyclooxygenase (COX)-2 induction was involved in prostaglandin (PG) E2 release by BK in cultured human airway smooth muscle (ASM) cells and analyzed the BK receptor subtypes responsible. BK stimulated PGE2release, COX activity, and COX-2 induction in a concentration- and time-dependent manner. It also time dependently enhanced arachidonic acid release. In short-term (15-min) experiments, BK stimulated PGE2 generation but did not increase COX activity or induce COX-2. In long-term (4-h) experiments, BK enhanced PGE2 release and COX activity and induced COX-2. The long-term responses were inhibited by the protein synthesis inhibitors cycloheximide and actinomycin D and the steroid dexamethasone. The effects of BK were mimicked by the B2-receptor agonist [Tyr(Me)8]BK, whereas the B1 agonist des-Arg9-BK was weakly effective at high concentrations. The B2antagonist HOE-140 potently inhibited all the effects, but the B1 antagonist des-Arg9,(Leu8)-BK was inactive. This study is the first to demonstrate that BK can induce COX-2. Conversion of increased arachidonic acid release to PGE2 by COX-1 is mainly involved in the short-term effect, whereas B2 receptor-related COX-2 induction is important in the long-term PGE2 release.

A human airway smooth muscle cell line that retains physiological responsiveness

1989 ◽  
Vol 256 (2) ◽  
pp. C329-C335 ◽  
Author(s):  
R. A. Panettieri ◽  
R. K. Murray ◽  
L. R. DePalo ◽  
P. A. Yadvish ◽  
M. I. Kotlikoff
Keyword(s):  
Smooth Muscle ◽  
Cell Line ◽  
Airway Smooth Muscle ◽  
Cytosolic Calcium ◽  
Airway Smooth Muscle Cell ◽  
Airway Smooth Muscle Cells ◽  
Functional Coupling ◽  
Functional Responses ◽  
Human Airway Smooth Muscle ◽  
Human Airway

We report the development of a nontransformed line of human airway smooth muscle cells retaining smooth muscle-specific contractile protein expression and physiological responsiveness to agonists implicated in inflammatory airway diseases. Specific responses to histamine, leukotrienes, bradykinin, platelet-activating factor, substance P, and thromboxane analogues are demonstrated as well as functional coupling to beta-adrenergic receptors. The cell line was characterized using indirect immunofluorescence, as well as electrophoretic separation and immunoblot analysis of smooth muscle-specific actin. Functional responses were assessed by measurements of cytosolic calcium and stimulation of adenosine 3',5'-cyclic monophosphate production. The cells retain their responsiveness over many population doublings and should be a useful model to examine specific receptor-effector mechanisms, as well as the effects of neurohumoral agents on the regulation of airway smooth muscle growth and differentiation.

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