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Zinc deficiency as a codeterminant for airway epithelial barrier dysfunction in an ex vivo model of COPD.

ABSTRACT: There is now convincing evidence that the airway epithelium drives the pathogenesis of COPD. A major aspect of this is the disease-related reduction in barrier function that is potentiated by dysregulation of tight junction (TJ) protein complexes. However, a significant number of studies using in vitro smoke exposure models have not observed alterations in barrier permeability. We have previously shown that zinc (Zn) is an influential cytoprotective factor for the airway epithelium, and its depletion by cigarette smoke produces disease-related modifications consistent with inflammatory changes in COPD. We hypothesized that Zn deficiency is a significant co-stimulus with cigarette smoke extract (CSE) for potentiating the leaky barrier phenotype exhibited in COPD. We employed an ex vivo model of differentiated human airway epithelium exposed to Zn depletion and CSE to determine the contribution of Zn in maintaining normal epithelial permeability. Western blot analysis demonstrated a significant downregulation of the TJ proteins such as ZO-1 (-1.93-fold, P<0.05) and Claudin-1 (-3.37-fold, P<0.01) with the combination exposure. Assessment of barrier function via paracellular ionic conductance and tracer permeability also showed that Zn depletion was an important factor, which potentiated an increase in epithelial permeability (P<0.001 for both) compared to Zn depletion or CSE exposures in isolation. Visual inspection of the epithelium using transmission electron microscopy revealed a marked reduction in junction complexes between the adjacent airway epithelial cells treated with a combination of Zn depletion and CSE. These observations identify Zn deficiency as a significant codeterminant with CSE as a factor leading to an increase in airway epithelial permeability. Hence, as Zn dyshomeostasis has been reported in the airway epithelium exposed to chronic cigarette smoke and inflammation, targeting these phenomena may represent a promising strategy to ameliorate the leaky barrier phenotype that is synonymous with COPD.

SUBMITTER: Roscioli E

PROVIDER: S-EPMC5723110 | biostudies-literature | 2017

REPOSITORIES: biostudies-literature

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Zinc deficiency as a codeterminant for airway epithelial barrier dysfunction in an ex vivo model of COPD.

Roscioli Eugene E Jersmann Hubertus Pa HP Lester Susan S Badiei Arash A Fon Andrew A Zalewski Peter P Hodge Sandra S

International journal of chronic obstructive pulmonary disease 20171205

There is now convincing evidence that the airway epithelium drives the pathogenesis of COPD. A major aspect of this is the disease-related reduction in barrier function that is potentiated by dysregulation of tight junction (TJ) protein complexes. However, a significant number of studies using in vitro smoke exposure models have not observed alterations in barrier permeability. We have previously shown that zinc (Zn) is an influential cytoprotective factor for the airway epithelium, and its depl ...[more]

PMID: 29255357

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Project description:BackgroundIndividuals with respiratory disease are being increasingly exposed to wildfire smoke as populations encroach further into forested regions and climate change continues to bring higher temperatures with lower rainfall. Frequent exposures have significant potential to accelerate conditions such as chronic obstructive pulmonary disease (COPD) which is characterised by an exaggerated inflammatory response to environmental stimuli. Here we employ models of human airway epithelium exposed to wildfire smoke-extract (WFSE) to examine modulation in airway epithelial cell (AEC) survival, fragility and barrier function.MethodsSubmerged cultures of small airway epithelial cells (SAEC) and differentiated air-liquid interface (ALI) cultures of primary bronchial AEC (bAEC) were treated for 1-24 h with 1-10% WFSE generated from plant species found in the Australian bushland. Autophagy (LC3-II and Sequestosome), apoptosis (Poly-(ADP)-Ribose Polymerase (PARP) cleavage) and tight junction proteins were measured using western blot. Barrier function was assessed via permeability of fluorescein tracers and measuring trans-epithelial electrical resistance. The production of IL-6 was assessed using ELISA.ResultsPrimary epithelial models exposed to WFSE exhibited a significant blockade in autophagy as evidenced by an increase in LC3-II coupled with a concomitant elevation in Sequestosome abundance. These exposures also induced significant PARP cleavage indicative of apoptotic changes. ALI cultures of bAEC treated with 5% WFSE demonstrated barrier dysfunction with significant increases in paracellular molecular permeability and ionic conductance, and a reduction in the abundance of the tight junction proteins ZO-1 and Claudin-1. These cultures also exhibited increased IL-6 secretion consistent with the aberrant and pro-inflammatory repair response observed in the COPD airways. Further, blocks in autophagy and barrier disruption were significantly elevated in response to WFSE in comparison to similar exposures with cigarette smoke-extract.ConclusionWFSE inhibits autophagic flux and induces barrier dysfunction in the airway epithelium. As autophagy is a central regulator of cellular repair, viability, and inflammation, targeting the block in autophagic flux may ameliorate the consequences of wildfire smoke-exposure for individuals with pre-existing respiratory conditions.

Project description:BackgroundThe airway epithelial barrier function is disrupted in the airways of asthmatic patients. Abnormal mitochondrial biogenesis is reportedly involved in the pathogenesis of asthma. However, the role of mitochondrial biogenesis in the airway barrier dysfunction has not been elucidated yet. This study aimed to clarify whether the peroxisome proliferator-activated receptor γ coactivator-1alpha (PGC-1α), a central regulator of mitochondrial biogenesis, is involved in the disruption of the airway barrier function induced by aeroallergens.MethodsBEAS-2B cells were exposed to house dust mite (HDM) and the expressions of PGC-1α and E-cadherin, a junctional protein, were examined by immunoblotting. The effect of SRT1720, a PGC-1α activator, was investigated by immunoblotting, immunocytochemistry, and measuring the transepithelial electrical resistance (TEER) on the HDM-induced reduction in mitochondrial biogenesis markers and junctional proteins in airway bronchial epithelial cells. Furthermore,the effects of protease activated receptor 2 (PAR2) inhibitor, GB83, Toll-like receptor 4 (TLR4) inhibitor, lipopolysaccharide from Rhodobacter sphaeroides (LPS-RS), protease inhibitors including E64 and 4-(2-Aminoethyl) benzenesulfonyl fluoride hydrochloride (AEBSF) on the HDM-induced barrier dysfunction were investigated.ResultsThe amounts of PGC-1α and E-cadherin in the HDM-treated cells were significantly decreased compared to the vehicle-treated cells. SRT1720 restored the expressions of PGC-1α and E-cadherin reduced by HDM in BEAS-2B cells. Treatment with SRT1720 also significantly ameliorated the HDM-induced reduction in TEER. In addition, GB83, LPS-RS, E64 and AEBSF prevented the HDM-induced reduction in the expression of PGC1α and E-cadherin.ConclusionsThe current study demonstrated that HDM disrupted the airway barrier function through the PAR2/TLR4/PGC-1α-dependent pathway. The modulation of this pathway could be a new approach for the treatment of asthma.

Dataset Information

Zinc deficiency as a codeterminant for airway epithelial barrier dysfunction in an ex vivo model of COPD.

Publications

Zinc deficiency as a codeterminant for airway epithelial barrier dysfunction in an ex vivo model of COPD.

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