Cystic fibrosis transmembrane conductance regulator is involved in airway epithelial wound repair

Abstract

The role of cystic fibrosis (CF) transmembrane conductance regulator (CFTR) in airway epithelial wound repair was investigated using normal human bronchial epithelial (NHBE) cells and a human airway epithelial cell line (Calu-3) of serous gland origin. Measurements of wound repair were performed using continuous impedance sensing to determine the time course for wound closure. Control experiments showed that the increase in impedance corresponding to cell migration into the wound was blocked by treatment with the actin polymerization inhibitor, cytochalasin D. Time lapse imaging revealed that NHBE and Calu-3 cell wound closure was dependent on cell migration, and that movement occurred as a collective sheet of cells. Selective inhibition of CFTR activity with CFTRinh-172 or short hairpin RNA silencing of CFTR expression produced a significant delay in wound repair. The CF cell line UNCCF1T also exhibited significantly slower migration than comparable normal airway epithelial cells. Inhibition of CFTR-dependent anion transport by treatment with CFTRinh-172 slowed wound closure to the same extent as silencing CFTR protein expression, indicating that ion transport by CFTR plays a critical role in migration. Moreover, morphologic analysis of migrating cells revealed that CFTR inhibition or silencing significantly reduced lamellipodia protrusion. These findings support the conclusion that CFTR participates in airway epithelial wound repair by a mechanism involving anion transport that is coupled to the regulation of lamellipodia protrusion at the leading edge of the cell.

Cx26 regulates proliferation of repairing basal airway epithelial cells

“…Recent data have reported the critical role of the cystic fibrosis transmembrane conductance regulator (Cftr) gene in airway epithelial wound healing (Schiller et al, 2010;Trinh et al, 2012). Although the mechanisms connecting Cftr deficiency to defective klf4 induction and enhanced HAEC activation remain to be established, our results point to a disturbed repair process of CF HAECs characterized by fast but desynchronized initiation of BC activation and proliferation in a primary 3D model of epithelium damage.…”

Quantitative Proteomics Reveals an Altered Cystic FibrosisIn VitroBronchial Epithelial Secretome

“…Overall, analysis of C3 showed increased proteolysis and (45). Additionally, the UNCCF1T cell line exhibits significantly slower migration (46). Some of the altered biological processes identified in our in vitro CF secretions were also identified in BALF from adults with CF and control subjects analyzed by shotgun proteomics and gene ontology (21).…”

“…Several proteases and protease inhibitors were quantitatively increased in CF apical secretions, including serine proteases and SERPINs, whereas the protease gelsolin and the antiprotease Serpin B5 exhibited decreased levels. Because unregulated proteases can damage respiratory epithelium and impair ciliary function (57), this continues the theme of a state of repair induced by inflammation (44), as previously demonstrated in UNCCF1T cells (46). MMP-9 belongs to a family of MMP zinc-and calcium-dependent proteolytic enzymes that regulate ECM turnover, especially during tissue repair after injury (58).…”

“…A variety of proteins traditionally associated with the ECM also exhibited increased levels in SILAC analysis of CF secretions. In general, ECM proteins serve as an anchor/scaffold for cell proliferation and differentiation, and their increased levels in CF secretions contribute to the theme of inherent and ongoing repair in CF airway epithelium (44,46,59). SILAC/MS/MS demonstrated that fibronectin, an ECM protein typically secreted basally in airway epithelium, was one of the most increased and abundant proteins in CF apical secretions, and ELISA validated a significant increase in fibronectin abundance in CF apical secretions from both life-extended and primary ALI cultures.…”

Development of a Lab-on-a-Chip Device for Rapid Nanotoxicity Assessment In Vitro

“…Impedance based sensors have been in studies for many applications, such as detection of cell migration [158,159], cell growth and proliferations [160], cell health [161,162], cytotoxicity [163], nanotoxicity [164], drug effects [165], and circulating tumor cell detection [166]. Many chip based electrochemical measurement devices for single cell analysis have also been fabricated earlier, and the challenge of capturing single cell has been mentioned.…”