The cystic fibrosis (CF) field is the beneficiary of five species of animal models that lack functional cystic fibrosis transmembrane conductance regulator (CFTR) channel. These models are rapidly informing mechanisms of disease pathogenesis and CFTR function regardless of how faithfully a given organ reproduces the human CF phenotype. New approaches of genetic engineering with RNA-guided nucleases are rapidly expanding both the potential types of models available and the approaches to correct the CFTR defect. The application of new CRISPR/Cas9 genome editing techniques are similarly increasing capabilities for in vitro modeling of CFTR functions in cell lines and primary cells using air-liquid interface cultures and organoids. Gene editing of CFTR mutations in somatic stem cells and induced pluripotent stem cells is also transforming gene therapy approaches for CF. This short review evaluates several areas that are key to building animal and cell systems capable of modeling CF disease and testing potential treatments.

Neutrophils are the first immune cells to kill invading microbes at sites of infection using a variety of processes, including the release of proteases, phagocytosis and the production of neutrophil extracellular traps (NETs). NET formation, or NETosis, is a specific and highly efficient process, which is induced by a variety of stimuli leading to expulsion of DNA, proteases and antimicrobial peptides to the extracellular space. However, uncontrolled NETosis may lead to adverse effects and exert tissue damage in pathological conditions. Here, we show that the ATP channel pannexin1 (Panx1) is functionally expressed by bone marrow-derived neutrophils (BMDNs) of wild-type (WT) mice and that ATP contributes to NETosis induced in vitro by the calcium ionophore A23187 or phorbol 12-myristate 13-acetate (PMA). Interestingly, neutrophils isolated from Panx1−/− mice showed reduced and/or delayed induction of NETosis. Brilliant blue FCF dye (BB-FCF), a Panx1 channel inhibitor, decreased NETosis in wild-type neutrophils to the extent observed in Panx1−/− neutrophils. Thus, we demonstrate that ATP and Panx1 channels contribute to NETosis and may represent a therapeutic target.

To elucidate the function of a gene in bacteria it is vital that targeted gene inactivation (allelic replacement) can be achieved. Allelic replacement is often carried out by disruption of the gene of interest by insertion of an antibiotic-resistance marker followed by subsequent transfer of the mutant allele to the genome of the host organism in place of the wild-type gene. However, due to their intrinsic resistance to many antibiotics only selected antibiotic-resistance markers can be used in members of the genus Burkholderia, including the Burkholderia cepacia complex (Bcc). Here we describe the construction of improved antibiotic-resistance cassettes that specify resistance to kanamycin, chloramphenicol or trimethoprim effectively in the Bcc and related species. These were then used in combination with and/or to construct a series enhanced suicide vectors, pSHAFT2, pSHAFT3 and pSHAFT-GFP to facilitate effective allelic replacement in the Bcc. Validation of these improved suicide vectors was demonstrated by the genetic inactivation of selected genes in the Bcc species Burkholderia cenocepacia and B. lata, and in the non-Bcc species, B. thailandensis.

Pathological remodeling of the airway epithelium is commonly observed in Cystic Fibrosis (CF). The different cell types that constitute the airway epithelium are regenerated upon injury to restore integrity and maintenance of the epithelium barrier function. The molecular signature of tissue repair in CF airway epithelial cells has, however, not well been investigated in primary cultures. We therefore collected RNA-seq data from well-differentiated primary cultures of bronchial human airway epithelial cells (HAECs) of CF (F508del/F508del) and non-CF (NCF) origins before and after mechanical wounding, exposed or not to flagellin. We identified the expression changes with time of repair of genes, the products of which are markers of the different cell types that constitute the airway epithelium (basal, suprabasal, intermediate, secretory, goblet and ciliated cells as well as ionocytes). Researchers in the CF field may benefit from this transcriptomic profile, which covers the initial steps of wound repair and revealed differences in this process between CF and NCF cultures.