scholarly journals TLR3 activation increases chemokine expression in human fetal airway smooth muscle cells

2016 ◽  
Vol 310 (2) ◽  
pp. L202-L211 ◽  
Author(s):  
Arij Faksh ◽  
Rodney D. Britt ◽  
Elizabeth R. Vogel ◽  
Michael A. Thompson ◽  
Hitesh C. Pandya ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Respiratory Syncytial Virus ◽  
Signaling Pathways ◽  
Airway Smooth Muscle ◽  
Poly I:C ◽  
Airway Smooth Muscle Cells ◽  
Tlr Agonists ◽  
Polycytidylic Acid ◽  
Syncytial Virus

Viral infections, such as respiratory syncytial virus and rhinovirus, adversely affect neonatal and pediatric populations, resulting in significant lung morbidity, including acute asthma exacerbation. Studies in adults have demonstrated that human airway smooth muscle (ASM) cells modulate inflammation through their ability to secrete inflammatory cytokines and chemokines. The role of ASM in the developing airway during infection remains undefined. In our study, we used human fetal ASM cells as an in vitro model to examine the effect of Toll-like receptor (TLR) agonists on chemokine secretion. We found that fetal ASM express multiple TLRs, including TLR3 and TLR4, which are implicated in the pathogenesis of respiratory syncytial virus and rhinovirus infection. Cells were treated with TLR agonists, polyinosinic-polycytidylic acid [poly(I:C)] (TLR3 agonist), lipopolysaccharide (TLR4 agonist), or R848 (TLR7/8 agonist), and IL-8 and chemokine (C-C motif) ligand 5 (CCL5) secretion were evaluated. Interestingly, poly(I:C), but neither lipopolysaccharide nor R848, increased IL-8 and chemokine (C-C motif) ligand 5 secretion. Examination of signaling pathways suggested that the poly(I:C) effects in fetal ASM involve TLR and ERK signaling, in addition to another major inflammatory pathway, NF-κB. Moreover, there are variations between fetal and adult ASM with respect to poly(I:C) effects on signaling pathways. Pharmacological inhibition suggested that ERK pathways mediate poly(I:C) effects. Overall, our data show that poly(I:C) initiates activation of proinflammatory pathways in developing ASM, which may contribute to immune responses to infection and exacerbation of asthma.

scholarly journals Respiratory syncytial virus induces β2-adrenergic receptor dysfunction in human airway smooth muscle cells

2021 ◽  
Vol 14 (685) ◽  
pp. eabc1983
Author(s):  
Terri J. Harford ◽  
Fariba Rezaee ◽  
Manveen K. Gupta ◽  
Vladimir Bokun ◽  
Sathyamangla V. Naga Prasad ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Respiratory Syncytial Virus ◽  
Airway Obstruction ◽  
Airway Smooth Muscle ◽  
Adrenergic Receptor ◽  
Young Patients ◽  
Airway Smooth Muscle Cells ◽  
Human Airway Smooth Muscle ◽  
Rsv Infection ◽  
Syncytial Virus

Pharmacologic agonism of the β2-adrenergic receptor (β2AR) induces bronchodilation by activating the enzyme adenylyl cyclase to generate cyclic adenosine monophosphate (cAMP). β2AR agonists are generally the most effective strategy to relieve acute airway obstruction in asthmatic patients, but they are much less effective when airway obstruction in young patients is triggered by infection with respiratory syncytial virus (RSV). Here, we investigated the effects of RSV infection on the abundance and function of β2AR in primary human airway smooth muscle cells (HASMCs) derived from pediatric lung tissue. We showed that RSV infection of HASMCs resulted in proteolytic cleavage of β2AR mediated by the proteasome. RSV infection also resulted in β2AR ligand–independent activation of adenylyl cyclase, leading to reduced cAMP synthesis compared to that in uninfected control cells. Last, RSV infection caused stronger airway smooth muscle cell contraction in vitro due to increased cytosolic Ca2+ concentrations. Thus, our results suggest that RSV infection simultaneously induces loss of functional β2ARs and activation of multiple pathways favoring airway obstruction in young patients, with the net effect of counteracting β2AR agonist–induced bronchodilation. These findings not only provide a potential mechanism for the reported lack of clinical efficacy of β2AR agonists for treating virus-induced wheezing but also open the path to developing more precise therapeutic strategies.

scholarly journals Basolateral activation with TLR agonists induces polarized cytokine release and reduces barrier function in RPE in vitro

2020 ◽  
Author(s):  
Laura Terheyden ◽  
Johann Roider ◽  
Alexa Klettner
Keyword(s):  
Cell Viability ◽  
Barrier Function ◽  
Neuronal Cell ◽  
Poly I:C ◽  
Rpe Cells ◽  
Ussing Chambers ◽  
Tlr Agonists ◽  
Danger Signals ◽  
Polycytidylic Acid

Abstract Purpose Systemic inflammation may be of importance in the development of AMD. RPE cells can recognize danger signals with toll-like receptors (TLR) and may react in a pro-inflammatory manner. In this study, we evaluated the basal and apical secretions of TNFα, IL-6, and IL-1β in primary RPE cells and RPE/choroid explant cells under basolateral stimulation of TLR2, 3, and 4; the effects on barrier function; and their influence on neuronal cell viability. Methods RPE/choroid tissue explants were prepared from porcine eyes and cultivated in modified Ussing chambers; primary porcine RPE cells on transwell plates. Cells were basally stimulated with agonists Pam2CSK4 (Pam; TLR2), polyinosinic/polycytidylic acid (Poly I:C; TLR3), and lipopolysaccharide (LPS; TLR4) for 24 h. Supernatants were evaluated with ELISA for cytokines TNFα, IL-6, and IL-1β. Apical supernatants were applied to SHSY-5Y cells, and cell viability was evaluated in MTT assay. Barrier function was tested by measuring transepithelial electrical resistance (TER) and occludin immunostaining. Results None of the tested TLR agonists was toxic on RPE cells after 24 h of exposure. Unstimulated RPE cells secreted hardly any cytokines. Pam induced IL-6, IL-1ß, and TNFα on the basal and apical sides at all concentrations tested. Poly I:C induced IL-6 and TNFα primarily at the basal side at lower but on both sides at higher concentrations. LPS induced IL-6, IL-1ß, and TNFα apically and basally at all concentrations tested. In the RPE/choroid, a strong difference between apical and basal secretions could be found. IL-6 was constitutively secreted basally, but not apically, but was induced by all agonists on both sides. IL-1ß and TNFα alpha were strongly induced on the basal side by all agonists. TER was reduced by all agonists, with Pam and LPS being effective in all concentrations tested. Occludin expression was unaltered, but the distribution was influenced by the agonists, with a less distinct localization at the cell borders after treatment. None of the agonists or supernatants of treated RPE and RPE/choroid organ cultures exerted any effect on viability of SHSY-5Y cells. Conclusions Danger signals activating TLRs can induce polarized cytokine expression and contribute to the loss of barrier function in the RPE.

TLR2 ligand engagement upregulates airway smooth muscle TNFα-induced cytokine production

2012 ◽  
Vol 302 (9) ◽  
pp. L838-L845 ◽  
Author(s):  
Melanie Manetsch ◽  
Petra Seidel ◽  
Udo Heintz ◽  
Wenchi Che ◽  
J. Margaret Hughes ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Signaling Pathways ◽  
Airway Smooth Muscle ◽  
Inflammatory Responses ◽  
Respiratory Infections ◽  
Mapk Signaling ◽  
Chronic Obstructive ◽  
Tlr2 Expression ◽  
Chronic Airway

Airway inflammation and respiratory infections are important factors contributing to disease exacerbation in chronic airway diseases such as asthma and chronic obstructive pulmonary disease. Airway smooth muscle (ASM) cells express Toll-like receptors (TLRs) and may be involved in the amplification of airway inflammatory responses during infectious exacerbations. We determined whether infectious stimuli (mimicked using Pam3CSK4, a synthetic bacterial lipopeptide that binds to TLR2/TLR1) further enhance ASM cell inflammatory responses to TNFα in vitro and the signaling pathways involved. Human ASM cells were pretreated for 1 h with Pam3CSK4 (1 μg/ml) in the absence or presence of TNFα (10 ng/ml), and IL-6 and IL-8 release was measured after 24 h. As expected, stimulation with Pam3CSK4 or TNFα alone induced significant IL-6 and IL-8 release. Furthermore, Pam3CSK4 significantly increased TNFα-induced IL-6 and IL-8 mRNA expression and protein release and neutrophil chemotactic activity. The potentiating effect of Pam3CSK4 on TNFα-induced inflammatory responses was not due to enhanced TLR2 expression nor did it involve augmentation of NF-κB or MAPK signaling pathways. Rather, Pam3CSK4 induced cAMP response element (CRE) binding protein phosphorylation and induced CRE-mediated transcriptional regulation, suggesting that Pam3CSK4 and TNFα are acting in concert to enhance ASM cytokine secretion via parallel transcriptional pathways. Our findings suggest that ASM cells may be involved in the amplification of airway inflammatory responses during infectious exacerbations in chronic airway disease.

scholarly journals Curcumin inhibits the proliferation of airway smooth muscle cells in vitro and in vivo

2013 ◽  
Vol 32 (3) ◽  
pp. 629-636 ◽  
Author(s):  
XIN ZENG ◽  
YING CHENG ◽  
YUEJUN QU ◽  
JIDE XU ◽  
ZHIYUAN HAN ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Airway Smooth Muscle ◽  
Muscle Cells ◽  
Airway Smooth Muscle Cells

Interleukin-13 inhibits proliferation and enhances contractility of human airway smooth muscle cells without change in contractile phenotype

2011 ◽  
Vol 300 (6) ◽  
pp. L958-L966 ◽  
Author(s):  
Paul-André Risse ◽  
Taisuke Jo ◽  
Fernando Suarez ◽  
Nobuaki Hirota ◽  
Barbara Tolloczko ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Calcium Signaling ◽  
Airway Smooth Muscle ◽  
Airway Smooth Muscle Cells ◽  
Activated T Cells ◽  
Inhibition Of Proliferation ◽  
Th2 Cytokine ◽  
Number Of Cells

IL-13 is an important mediator of allergen-induced airway hyperresponsiveness. This Th2 cytokine, produced by activated T cells, mast cells, and basophils, has been described to mediate a part of its effects independently of inflammation through a direct modulation of the airway smooth muscle (ASM). Previous studies demonstrated that IL-13 induces hyperresponsiveness in vivo and enhances calcium signaling in response to contractile agonists in vitro. We hypothesized that IL-13 drives human ASM cells (ASMC) to a procontractile phenotype. We evaluated ASM phenotype through the ability of the cell to proliferate, to contract, and to express contractile protein in response to IL-13. We found that IL-13 inhibits human ASMC proliferation (expression of Ki67 and bromodeoxyuridine incorporation) in response to serum, increasing the number of cells in G0/G1 phase and decreasing the number of cells in G2/M phases of the cell cycle. IL-13-induced inhibition of proliferation was not dependent on signal transducer and activator of transcription-6 but was IL-13Rα2 receptor dependent and associated with a decrease of Kruppel-like factor 5 expression. In parallel, IL-13 increased calcium signaling and the stiffening of human ASMC in response to 1 μM histamine, whereas the stiffening response to 30 mM KCl was unchanged. However, Western blot analysis showed unchanged levels of calponin, smooth muscle α-actin, vinculin, and myosin. We conclude that IL-13 inhibits proliferation via the IL-13Rα2 receptor and induces hypercontractility of human ASMC without change of the phenotypic markers of contractility.

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