Single cell RNA sequencing of large and small airway surface epithelium in newborn cystic fibrosis (CF) and non-CF pigs
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ABSTRACT:
ORGANISM(S): Sus scrofa
PROVIDER: GSE150211 | GEO | 2021/03/16
REPOSITORIES: GEO
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Project description:We did bulk RNA sequencing in newborn cystic fibrosis (CF) and non-CF pig kidney. We compared kidney gene expression profiling between non-CF and CF pigs. RNA sequencing results showed that there is not significant difference between non-CF and CF in terms of gene expression, suggesting that CFTR knockout does not affect kidney development in newborn pigs.
2024-06-07 | GSE217122 | GEO
Project description:Cystic fibrosis (CF) is a life-shortening disease caused by mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene. Although bacterial lung infection and the resulting inflammation cause most of the morbidity and mortality, how loss of CFTR first disrupts airway host defense has remained uncertain. We asked what abnormality impairs elimination when a bacterium lands on the pristine surface of a newborn CF airway? To investigate this defect, we interrogated the viability of individual bacteria immobilized on solid grids and placed on the airway surface. As a model we studied CF pigs, which spontaneously develop hallmark features of CF lung disease. At birth, their lungs lack infection and inflammation, but have a reduced ability to eradicate bacteria. Here we show that in newborn wild-type pigs, the thin layer of airway surface liquid (ASL) rapidly killed bacteria in vivo, when removed from the lung, and in primary epithelial cultures. Lack of CFTR reduced bacterial killing. We found that ASL pH was more acidic in CF, and reducing pH inhibited the antimicrobial activity of ASL. Reducing ASL pH diminished bacterial killing in wild-type pigs, and increasing ASL pH rescued killing in CF pigs. These results directly link the initial host defense defect to loss of CFTR, an anion channel that facilitates HCO3- transport. Without CFTR, airway epithelial HCO3- secretion is defective, ASL pH falls and inhibits antimicrobial function, and thereby impairs killing of bacteria that enter the newborn lung. These findings suggest that increasing ASL pH might prevent the initial infection in patients with CF and that assaying ASL pH or bacterial killing could report on the benefit of therapeutic interventions. 11 samples of trachea primary airway epithelial cultures representing CFTR+/+ and CFTR-/- pigs. Pig samples representing 14 bronchus and 12 trachea tissue samples submitted in GSE21071.
2012-03-29 | E-GEOD-36906 | biostudies-arrayexpress
Project description:Cystic fibrosis (CF) is a life-shortening disease caused by mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene. Although bacterial lung infection and the resulting inflammation cause most of the morbidity and mortality, how loss of CFTR first disrupts airway host defense has remained uncertain. We asked what abnormality impairs elimination when a bacterium lands on the pristine surface of a newborn CF airway? To investigate this defect, we interrogated the viability of individual bacteria immobilized on solid grids and placed on the airway surface. As a model we studied CF pigs, which spontaneously develop hallmark features of CF lung disease. At birth, their lungs lack infection and inflammation, but have a reduced ability to eradicate bacteria. Here we show that in newborn wild-type pigs, the thin layer of airway surface liquid (ASL) rapidly killed bacteria in vivo, when removed from the lung, and in primary epithelial cultures. Lack of CFTR reduced bacterial killing. We found that ASL pH was more acidic in CF, and reducing pH inhibited the antimicrobial activity of ASL. Reducing ASL pH diminished bacterial killing in wild-type pigs, and increasing ASL pH rescued killing in CF pigs. These results directly link the initial host defense defect to loss of CFTR, an anion channel that facilitates HCO3- transport. Without CFTR, airway epithelial HCO3- secretion is defective, ASL pH falls and inhibits antimicrobial function, and thereby impairs killing of bacteria that enter the newborn lung. These findings suggest that increasing ASL pH might prevent the initial infection in patients with CF and that assaying ASL pH or bacterial killing could report on the benefit of therapeutic interventions.
2012-03-30 | GSE36906 | GEO
Project description:Chronic infection of the lungs by bacteria such as Pseudomonas aeruginosa plays an important role in the natural history of Cystic Fibrosis (CF). Altered mucin secretions and glycosylations are hallmarks of bacterial infections but their exact contributions to the development of chronic infection and/or inflammation remain to be established. We studied the lung environment in newborn CFTR-/- pigs, which closely mirrors the human CF phenotype at the levels of mucin glycosylation, mucociliary transport (MCT) and immune response to P. aeruginosa infections. We show an increased sialylation of mucins from the tracheal mucosa of CFTR-/- pigs, which is also associated to increased Pseudomonas aeruginosa adherence. RNA-seq analysis did not show any altered expression of inflammatory genes, nor of sialylation enzymes. CFTR-/- piglets were unable to clear P. aeruginosa from the airways, leading to increased bacteria time of residence in the airways. We propose that alterations in mucin sialylation and MCT directly contribute to the early development of a favorable environment for bacterial colonization of the CF airways in the absence of an intrinsic inflammation.
2020-06-01 | GSE99862 | GEO
Project description:Lung disease causes most of the morbidity and mortality in cystic fibrosis (CF). However, understanding its pathogenesis has been hindered by lack of an animal model with characteristic features of CF. To overcome this problem, we recently generated pigs with targeted CFTR genes. We now report that within months of birth, CF pigs spontaneously develop hallmark features of CF lung disease including airway inflammation, remodeling, mucus accumulation, and infection. Their lungs contained multiple bacterial species, suggesting an equal opportunity host defense defect. In humans, the temporal and/or causal relationships between inflammation and infection have remained uncertain. To investigate these processes, we studied newborn pigs. Their lungs showed no inflammation, but were less often sterile than controls. Moreover, after intrapulmonary bacterial challenge, CF pigs failed to eradicate bacteria as effectively as wild- type pigs. These results suggest that impaired bacterial elimination is the pathogenic event that initiates a cascade of inflammation and pathology in CF lungs. Finding that CF pigs have a bacterial host defense defect within hours of birth provides an exciting opportunity to further investigate pathogenesis and to test therapeutic and preventive strategies before secondary consequences develop. Pig model of human CF lung disease
2010-09-17 | E-GEOD-21071 | biostudies-arrayexpress
Project description:Lung disease causes most of the morbidity and mortality in cystic fibrosis (CF). However, understanding its pathogenesis has been hindered by lack of an animal model with characteristic features of CF. To overcome this problem, we recently generated pigs with targeted CFTR genes. We now report that within months of birth, CF pigs spontaneously develop hallmark features of CF lung disease including airway inflammation, remodeling, mucus accumulation, and infection. Their lungs contained multiple bacterial species, suggesting an equal opportunity host defense defect. In humans, the temporal and/or causal relationships between inflammation and infection have remained uncertain. To investigate these processes, we studied newborn pigs. Their lungs showed no inflammation, but were less often sterile than controls. Moreover, after intrapulmonary bacterial challenge, CF pigs failed to eradicate bacteria as effectively as wild- type pigs. These results suggest that impaired bacterial elimination is the pathogenic event that initiates a cascade of inflammation and pathology in CF lungs. Finding that CF pigs have a bacterial host defense defect within hours of birth provides an exciting opportunity to further investigate pathogenesis and to test therapeutic and preventive strategies before secondary consequences develop.
2010-09-17 | GSE21071 | GEO
Project description:Our laboratory has held a long interest in the glycosylation changes seen on the surface of airway epithelia of patients with the disease cystic fibrosis (CF). Experiments from our laboratory have detailed a CF glycosylation phenotype of increased Fuca1,3/4 and decreased Fuca1,2 and sialic acid on the surfaces of immortalized and primary CF cells compared to non-CF cells. Further, we have shown that gene transfer and subsequent expression of a wild type CF plasmid in CF airway cells results in correction or reversal of this glycosylation phenotype. We hypothesize that the changes in glycosylation seen in CF cells are key in the pathophysiology of the cystic fibrosis airway disease. For example, it has been shown that Pseudomonas aeruginosa, a bacterium that has a predilection for colonizing CF airways, adheres to asialylated glycolipids and glycoconjugates with terminal Fuca1,3/4. One focus of our laboratory is to elucidate the etiology of the glycosylation changes seen in CF cells and the mechanism by which these changes are reversed by wild type CFTR gene transfer. We propose to study the gene expression of immortalized and primary CF and non-CF airway epithelial cells: 1. CF/T43 vs. BEAS-2B cells. These are two widely used immortalized airway cell lines that we have used extensively in the past. 2. C38 cells; C38 cells are IB3 cells expressing wtCFTR. The experimental focus is to elucidate the etiology of the glycosylation changes seen in Cystic Fibrosis (CF) cells and the mechanism by which these changes are reversed by wild type CFTR gene transfer. To do so, the gene expression of immortalized and primary CF and non-CF airway epithelial cells were compared and studied. Cell lines used were CF/T43 and BEAS-2B, both widely used immortalized airway cell lines. Other cell lines studied included C38 cell lines (clonal derivatives of IB3 cells expressing wtCFTR).
2011-01-07 | E-GEOD-26482 | biostudies-arrayexpress
Project description:A deficiency in cystic fibrosis transmembrane conductance regulator (CFTR) function in cystic fibrosis (CF) leads to chronic lung disease. However, the molecular mechanisms are not well understood and therapies that can help all patients remain elusive. CF is associated with abnormalities in fatty acids, ceramides and cholesterol, therefore we examined the impact of CFTR deficiency on lipid metabolism and pro-inflammatory signaling in airway epithelium using mass spectrometric, protein array and RNAseq analyses. We observed a striking imbalance in fatty acid and ceramide metabolism, associated with chronic oxidative stress under basal conditions in CF mouse lung and well differentiated bronchial epithelial cell cultures of CFTR knock out pig and CF patients. Cell autonomous features of all three CF models included high ratios of ω-6- to ω-3-polyunsaturated fatty acids and long- to very long- chain ceramide species (LCC/VLCC). The anti-oxidants glutathione (GSH) and deferoxamine partially corrected the lipid profile indicating that oxidative stress may promote the lipid abnormalities. CFTR-targeted modulators reduced the lipid imbalance and apparent oxidative stress, confirming the CFTR dependence of lipid ratios. RNA sequencing and protein array analysis revealed higher expression and shedding of cytokines and growth factors from CF epithelial cells compared to non-CF cells, consistent with sterile inflammation and tissue remodeling under basal conditions. Treatment with antioxidants or CFTR modulators that mimic the approved combination therapies, Orkambi and Trikafta, did not suppress the inflammatory phenotype. These results suggest that anti-inflammatory therapies may provide additional benefit for CF patients taking CFTR modulator drugs.
2021-01-01 | GSE154802 | GEO
Project description:Our laboratory has held a long interest in the glycosylation changes seen on the surface of airway epithelia of patients with the disease cystic fibrosis (CF). Experiments from our laboratory have detailed a CF glycosylation phenotype of increased Fuca1,3/4 and decreased Fuca1,2 and sialic acid on the surfaces of immortalized and primary CF cells compared to non-CF cells. Further, we have shown that gene transfer and subsequent expression of a wild type CF plasmid in CF airway cells results in correction or reversal of this glycosylation phenotype. We hypothesize that the changes in glycosylation seen in CF cells are key in the pathophysiology of the cystic fibrosis airway disease. For example, it has been shown that Pseudomonas aeruginosa, a bacterium that has a predilection for colonizing CF airways, adheres to asialylated glycolipids and glycoconjugates with terminal Fuca1,3/4. One focus of our laboratory is to elucidate the etiology of the glycosylation changes seen in CF cells and the mechanism by which these changes are reversed by wild type CFTR gene transfer.
2011-01-07 | GSE26482 | GEO
Project description:We have compared gene expression in human nasal brushing cells from 19 cystic fibrosis (CF) patients and 19 healthy controls using a 5.2K cDNA microarray. Our aim is to identify new disease biomarkers for the Cystic Fibrosis Gene Therapy Consortium. These markers will be used to report more effectively on the response to the administration of gene therapy in vivo. Cystic Fibrosis is a recessive genetic disease caused by mutations in the cystic fibrosis conductance regulator (CFTR) gene which encodes a chloride ion channel. The most common mutation is the ∆F508 mutation, present on 70% of CF chromosomes in Caucasian populations. The disease affects many organs in the body such as the pancreas, liver, sweat glands, small intestine and reproductive tracts but is most commonly associated with progressive, inflammatory lung disease. The current average life expectancy of CF patients is 35 years. Gene therapy is being developed as a treatment for CF airway disease, however, means of measuring the efficiency and efficacy of gene therapy in vivo are lacking. This is mainly due to the difficulty in measuring the chloride conductance of CFTR in cells and tissues. Furthermore, clinical assays for measuring improvements in lung function are insensitive. Surrogate markers of inflammation and CFTR function will therefore be important for the effective assessment of gene therapy in vivo. We have analysed gene expression in human nasal epithelium as this is considered an accessible surrogate for the conducting airways where disease manifests in the majority of patients. Additionally, this tissue will be sampled in clinical trials.
2011-04-30 | E-MEXP-436 | biostudies-arrayexpress