Fig. 1.
Altered morphology and confluence of bronchial epithelial cells carrying rs35699176. a Schematic of the rs35699176 targeted region and sequences of the human ZNF77 locus, including the chromatogram derived from Sanger sequencing of the mutant allele. Gene expression of ZNF77 in 16HBE cells and 16HBErs35699176 cell lines is equivalent (right panel). b Transepithelial electrical resistance (TER) does not increase after 8 days in monolayers formed by 16HBErs35699176 cells (panel b, solid line, ***P < 0.0001) indicating failure to form confluent cell layers. c Microscopy reveals alterations in 16HBErs35699176 monolayer morphology. Phase contrast microscopy shows rounded, non-adherent cells of 16HBErs35699176; confocal, surface rendered projection, stained with Cell Mask Deep Red shows holes in the monolayer and single optical section from projection. Images were processed using Metamorph and IMARIS v8.0.1 software. Asterisks denote lack of confluency. Scale bar represents 20 µm. d Fold change in gene expression of caveolin, occludin and E-cadherin genes after 6 h (solid columns) and 6 h (hashed columns) in 16HBE and 16HBErs35699176 cells co-cultured with A. fumigatus. Data in graphs are represented as mean ± standard deviation of a minimum of three experiments performed in biological and technical triplicates. Differences in gene expression and transepithelial resistance for each time point were analysed using the Mann–Whitney U test, ***P < 0.001