ENaC, iNOS, mucins, and wound healing in cystic fibrosis airway epithelial and submucosal cells

Hussain, Rashida; Umer, Hafiz Muhammad; Björkqvist, Maria; Roomans, Godfried M.

doi:10.1002/cbi3.10014

Cited by 4 publications

(7 citation statements)

References 71 publications

(107 reference statements)

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“…2A ). These epithelial cells produce the mucins MUC1, MUC2, MUC4, and MUC5B ( 37 , 38 ). Once biofilms developed, the coculture was treated with a dispersal agent, such as sodium nitrite, for 15 min.…”

Section: Resultsmentioning

confidence: 99%

Dispersal of Epithelium-Associated Pseudomonas aeruginosa Biofilms

Zemke

D'Amico

Snell

et al. 2020

mSphere

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Pseudomonas aeruginosa grows in highly antibiotic-tolerant biofilms during chronic airway infections. Dispersal of bacteria from biofilms may restore antibiotic susceptibility or improve host clearance. We describe models to study biofilm dispersal in the nutritionally complex environment of the human airway. P. aeruginosa was cocultured in the apical surface of airway epithelial cells (AECs) in a perfusion chamber. Dispersal, triggered by sodium nitrite, a nitric oxide (NO) donor, was tracked by live cell microscopy. Next, a static model was developed in which biofilms were grown on polarized AECs without flow. We observed that NO-triggered biofilm dispersal was an energy-dependent process. From the existing literature, NO-mediated biofilm dispersal is regulated by DipA, NbdA, RbdA, and MucR. Interestingly, altered signaling pathways appear to be used in this model, as deletion of these genes failed to block NO-induced biofilm dispersal. Similar results were observed using biofilms grown in an abiotic model on glass with iron-supplemented cell culture medium. In cystic fibrosis, airway mucus contributes to the growth environment, and a wide range of bacterial phenotypes are observed; therefore, we tested biofilm dispersal in a panel of late cystic fibrosis clinical isolates cocultured in the mucus overlying primary human AECs. Finally, we examined dispersal in combination with the clinically used antibiotics ciprofloxacin, aztreonam and tobramycin. In summary, we have validated models to study biofilm dispersal in environments that recapitulate key features of the airway and identified combinations of currently used antibiotics that may enhance the therapeutic effect of biofilm dispersal. IMPORTANCE During chronic lung infections, Pseudomonas aeruginosa grows in highly antibiotic-tolerant communities called biofilms that are difficult for the host to clear. We have developed models for studying P. aeruginosa biofilm dispersal in environments that replicate key features of the airway. We found that mechanisms of biofilm dispersal in these models may employ alternative or additional signaling mechanisms, highlighting the importance of the growth environment in dispersal events. We have adapted the models to accommodate apical fluid flow, bacterial clinical isolates, antibiotics, and primary human airway epithelial cells, all of which are relevant to understanding bacterial behaviors in the context of human disease. We also examined dispersal agents in combination with commonly used antipseudomonal antibiotics and saw improved clearance when nitrite was combined with the antibiotic aztreonam.

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Section: Resultsmentioning

confidence: 99%

Dispersal of Epithelium-Associated Pseudomonas aeruginosa Biofilms

Zemke

D'Amico

Snell

et al. 2020

mSphere

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“…It is worth to note that cell proliferation and migration were not assayed in wounded conditions [ 165 ], differently from other studies which nevertheless found a difference in cell migration during wound repair, as noted above. Interestingly, CFTRinh-172 induced a delay in wound repair, not only in non-CF cells (16HBE and corrected CFBE cells) but also in CFBE monolayers [ 165 ], indicating that this effect may not be related to the ion channel function of CFTR, but most probably is an unspecific effect of the inhibitor. A non-specific effect of CFTRinh-172 has also been found in other cell types of airway and non-airway origin [ 185 ].…”

Section: Airway Epithelial Regeneration Wound Repair and Cfmentioning

confidence: 92%

“…Ex-vivo xenograft models have been used in the context of re-differentiation of a native airway epithelium leading to regeneration and mucosa reconstitution by basal-like cells [ 161 ]. On the other hand, in-vitro models have been used to study wound repair through simplistic models of injury and repair [ 138 , 162 , 163 , 164 , 165 , 166 ]. These studies have been mainly conducted in submerged cultures on plastic, with either immortalized or primary cells lines, leaving aside all the fact that in vivo we deal instead with a polarized pseudostratified airway epithelium.…”

Section: Airway Epithelial Regeneration Wound Repair and Cfmentioning

confidence: 99%

“…The scratch assay was instrumental in determining whether CFTR levels corresponded to certain wound closure rates [ 165 ]. Wound repair was found to be significantly slower in CFBE cells compared to corrected CFBE cells (CFBE41o-pCep4, overexpressing wt-CFTR), but the comparison between CFBE and 16HBE cells showed no significant difference in the repair.…”

Section: Airway Epithelial Regeneration Wound Repair and Cfmentioning

confidence: 99%

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Pathophysiology of Lung Disease and Wound Repair in Cystic Fibrosis

Conese

Gioia

2021

Pathophysiology

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Cystic fibrosis (CF) is an autosomal recessive, life-threatening condition affecting many organs and tissues, the lung disease being the chief cause of morbidity and mortality. Mutations affecting the CF Transmembrane Conductance Regulator (CFTR) gene determine the expression of a dysfunctional protein that, in turn, triggers a pathophysiological cascade, leading to airway epithelium injury and remodeling. In vitro and in vivo studies point to a dysregulated regeneration and wound repair in CF airways, to be traced back to epithelial CFTR lack/dysfunction. Subsequent altered ion/fluid fluxes and/or signaling result in reduced cell migration and proliferation. Furthermore, the epithelial-mesenchymal transition appears to be partially triggered in CF, contributing to wound closure alteration. Finally, we pose our attention to diverse approaches to tackle this defect, discussing the therapeutic role of protease inhibitors, CFTR modulators and mesenchymal stem cells. Although the pathophysiology of wound repair in CF has been disclosed in some mechanisms, further studies are warranted to understand the cellular and molecular events in more details and to better address therapeutic interventions.

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“…U87 and U251 cells (1×10 6 cells/well) were seeded into 6-well plates. After reaching 70% confluence, the cells were transfected with miR-10b-5p mimic or inhibitor, and incubated for 48 h. Using a 200-µl sterile pipette tip, three scratches were created in each well (15). Next, the cells were washed twice with PBS and incubated in medium supplemented with 2% FBS for 24 h. In order to assess the cell migration, images of the wells were captured at 0 and 24 h in three randomly selected microscopic fields (Olympus Corporation, Tokyo, Japan).…”

Section: Methodsmentioning

confidence: 99%

MicroRNA‑10b‑5p downregulation inhibits the invasion of glioma cells via modulating homeobox B3 expression

Xiong

et al. 2019

Exp Ther Med

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MicroRNA-10b (miR-10b) has been reported to be specifically upregulated in glioma tissues and cell lines. The aim of the present study was to investigate the effect of miR-10b-5p on the proliferation and invasion of glioma cells, and the possible underlying molecular mechanism. Cell viability was evaluated using an MTT assay, and flow cytometry was performed for cell cycle analysis. The effects of miR-10b-5p on glioma cell migration and invasion were assessed using wound healing and Transwell assays, respectively. The activity of matrix metalloproteinase 2 (MMP2) was also determined using zymography. Furthermore, homeobox B3 (HOXB3) mRNA expression in glioma cells was examined using quantitative polymerase chain reaction analysis. The protein expression levels of HOXB3, high mobility group box 1 (HMGB1) and Ras homolog family member C (RhoC) were further measured using western blotting. It was observed that glioma cells transfected with miR-10b-5p inhibitor exhibited significantly decreased proliferation. The wound healing and Transwell assays demonstrated that the miR-10b-5p inhibitor reduced the ability of glioma cells to migrate and invade, while transfection with miR-10b-5p mimic exhibited the opposite effect. HOXB3 was downregulated by miR-10b-5p at both the mRNA and protein levels. In addition, the expression of proteins associated with migration and invasion, including HMGB1, RhoC and MMP2, was upregulated in glioma cells transfected with miR-10b-5p mimic, while these proteins were downregulated in cells transfected with miR-10b-5p inhibitor. Taken together, the findings of the present study indicated that miR-10b-5p downregulation suppressed glioma cell proliferation and invasion, possibly by modulating HOXB3, which may provide a novel bio-target for glioma therapy.

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ENaC, iNOS, mucins, and wound healing in cystic fibrosis airway epithelial and submucosal cells

Cited by 4 publications

References 71 publications

Dispersal of Epithelium-Associated Pseudomonas aeruginosa Biofilms

Dispersal of Epithelium-Associated Pseudomonas aeruginosa Biofilms

Pathophysiology of Lung Disease and Wound Repair in Cystic Fibrosis

MicroRNA‑10b‑5p downregulation inhibits the invasion of glioma cells via modulating homeobox B3 expression

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