CF (1565) vs CFTR-corrected (1564) iPSC-airway at ALI
Ontology highlight
ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE203008 | GEO | 2023/03/02
REPOSITORIES: GEO
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Project description:Platform: iPSC-derived airway plated on 2D-air liquid interface through basal cell intermediate. Purpose of experiment: To examine the role of mutant CFTR on IPSC-derived airway epithelium (ie. immune dysregulation; dysregulated calcium channel signaling etc). Description of samples:Typical phenotype CF F508del (1565) and syngeneic CFTR-corrected (1564) iPSCs- derived airway epithelium at D90 (D15 CD47hi/CD26lo; replated in 3DMG cultured in 210DCIY x 2 weeks, single-cell passaged and cultured in PNExPlus+dual smad media in 3D, s/p 2 NGFR sorts, cultured at air-liquid interface at 2D)
2023-03-02 | GSE203006 | GEO
Project description:To interrogate the intrinsic effect of CFTR on airway epithelium; to interrogate the presence of FOXi1+ASLC3+ ionocyte population. Description of samples:1567 (F508del CF patient of rapid progressive phenotye) and 1566 (isogenic CFTR-repaired pair). iPSC-derived airway epithelium, s/p D15CD47hi/CD26lo sort, 210DCYI media x
2023-03-02 | GSE203007 | GEO
Project description:This SuperSeries is composed of the SubSeries listed below.
2023-03-02 | GSE203009 | GEO
Project description:Purpose: To identify the cell types that contribute to CFTR expression and function within the proximal-distal axis of the normal human lung. Methods: Single cell RNA-seq (scRNA-seq) was performed on freshly isolated human large and small airway epithelial cells. Single cell-based RNA in situ hybridization (scRNA-ISH) and quantitative RT-PCR (scqRT-PCR) were performed for validation. In vitro culture systems correlated CFTR function to cell types. Lentiviruses were used for cell-type specific transduction of wild-type CFTR in CF cells. Results: scRNA-seq identified secretory cells as dominating CFTR expression in normal human large and particularly small airways, followed by basal cells. Ionocytes expressed the highest CFTR levels but were rare, while expression in ciliated cells was infrequent and low. scRNA-ISH, and scqRT-PCR confirmed the scRNA-seq findings. CF lungs exhibited distributions of CFTR and ionocytes similar to normal controls. CFTR-mediated Cl- secretion tracked secretory cell, not ionocyte, densities. Further, the nucleotide-purinergic regulatory system that controls CFTR-mediated hydration was associated with secretory cells. Lentiviral transduction of wild-type CFTR produced CFTR-mediated Cl- secretion in CF airway secretory cells but not ciliated cells. Conclusions: Secretory cells dominate CFTR expression and function in human airway superficial epithelia. CFTR therapies may need to restore CFTR function to multiple cell types, with a focus on secretory cells.
2020-12-16 | GSE160674 | GEO
Project description:Purpose: To identify the cell types that contribute to CFTR expression and function within the proximal-distal axis of the normal human lung. Methods: Single cell RNA-seq (scRNA-seq) was performed on freshly isolated human large and small airway epithelial cells. Single cell-based RNA in situ hybridization (scRNA-ISH) and quantitative RT-PCR (scqRT-PCR) were performed for validation. In vitro culture systems correlated CFTR function to cell types. Lentiviruses were used for cell-type specific transduction of wild-type CFTR in CF cells. Results: scRNA-seq identified secretory cells as dominating CFTR expression in normal human large and particularly small airways, followed by basal cells. Ionocytes expressed the highest CFTR levels but were rare, while expression in ciliated cells was infrequent and low. scRNA-ISH, and scqRT-PCR confirmed the scRNA-seq findings. CF lungs exhibited distributions of CFTR and ionocytes similar to normal controls. CFTR-mediated Cl- secretion tracked secretory cell, not ionocyte, densities. Further, the nucleotide-purinergic regulatory system that controls CFTR-mediated hydration was associated with secretory cells. Lentiviral transduction of wild-type CFTR produced CFTR-mediated Cl- secretion in CF airway secretory cells but not ciliated cells. Conclusions: Secretory cells dominate CFTR expression and function in human airway superficial epithelia. CFTR therapies may need to restore CFTR function to multiple cell types, with a focus on secretory cells.
2020-12-16 | GSE160673 | GEO
Project description:Atmospheric oxygen tension is increasingly recognized to be hyperoxic for the maintenance of in vitro cell cultures. Oxygen has broad implications on energy metabolism, cellular growth and many regulatory functions. Primary airway epithelial cells (AECs) differentiated at air-liquid interface (ALI) are the gold standard for preclinical assessment of cystic fibrosis transmembrane conductance regulatory (CFTR) modulator efficacy in CF. The fidelity of CF AECs cultured at atmospheric oxygen tension rather than at reduced oxygen tension which more closely reflect the in vivo environment has not been studied to date. This study examined the impact of ambient (21%) and low (2%) oxygen tension on the expansion and differentiation of nasal epithelial cells (hNECs) derived from 11 participants (8 with CF and 3 non-CF). hNECs expanded under normoxic and chronic hypoxic conditions demonstrated epithelial cobblestone morphology and similar proliferation rate. hNECs differentiated at hypoxia demonstrated poorer differentiation capacity (significantly thinner epithelium and lower TEER) and a shift from ciliated to secretory epithelial phenotype. Hypoxic differentiated hNECs had significantly shorter cilia length, slower beating frequency but had improved cilia coordination. CFTR functional response is altered in hypoxic differentiated hNECs. This study highlights the need to reconsider the oxygen tension used in CF primary cell cultures so as to preserve the characteristics and functional response of the cell models as we progress towards personalised medicine in CF.
2023-03-25 | PXD038893 | Pride
Project description:Background. Accumulating evidence indicates that CFTR modulators can be effective for CF sufferers. However, in vivo the microRNAs that regulate CFTR are increased in the CF lung milieu and this can nullify the beneficial effects of CFTR modulators. Methods. A panel of target site blockers (TSBs) designed against the CFTR 3’untranslated region (UTR) were tested for their ability to increase CFTR expression by CFTR 3’UTR luciferase assay and western blot. The effect of two lead TSBs on CFTR activity were tested alone or in combination with selected CFTR modulators in four separate p.Phe508del/p.Phe508del in vitro and ex vivo models (CFBE41o-, Cufi-1, primary bronchial epithelial cells, iPSC-derived lung organoids). Respirable poly-lactic-co-glycolic acid (PLGA) nanoparticles encapsulating the TSBs were formulated and tested. Findings. TSBs that target the miR-145-5p or miR-223-3p binding sites at positons 298-305 and 166-173 in the CFTR 3’UTR, respectively, increased CFTR expression and CFTR anion channel activity, and enhance the effects of Lumacaftor/Ivacaftor or Lumacftor/Tezacaftor in CF BECs. PLGAs encapsulating the TSBs promote CFTR expression in primary BECs and retain their biophysical characteristics following nebulization. Interpretation. Alone or in combination with CFTR modulators, CFTR-targeting TSBs encapsulated in PLGA nanoparticles and nebulized to the lung could overcome microRNA-mediated inhibition of CFTR in CF bronchial epithelium. This strategy represents a promising drug-device combination therapy for the treatment for CFTR dysfunction in the lung.
2019-03-27 | GSE128861 | GEO
Project description:Speciation leads to adaptive changes in organ cellular anatomy and physiology. These evolutionary changes create challenges for studying rare cell type functions that diverge between human and mice. Rare CFTR-rich pulmonary ionocytes exist throughout the cartilaginous airways of humans, but limited presence and divergent biology in the proximal trachea of mice has prevented the use of traditional transgenic models to elucidate ionocyte functions in the airway. Here we describe the creation and use of novel conditional genetic ferret models to dissect pulmonary ionocyte biology and function by enabling ionocyte lineage tracing (FOXI1-CreERT2::ROSA-TG), ionocyte ablation (FOXI1-KO), and ionocyte-specific deletion of CFTR (FOXI1-CreERT2::CFTRL/L). By comparing these models to cystic fibrosis (CF) ferrets, we demonstrate that ionocytes control airway surface liquid (ASL) absorption, secretion, pH, and mucus viscosity—leading to reduced ASL volume and impaired mucociliary clearance in CF, FOXI1-KO, and FOXI1-CreERT2::CFTRL/L ferrets. These processes were regulated by CFTR- dependent ionocyte transport of Cl– and HCO3–, as determined electrophysiologically and by single-cell imaging with a conditionally activated halide fluorescent sensor. Single-cell transcriptomics, using pulse-seq of lineage-traced ionocyte-enriched cultures, revealed three subtypes of pulmonary ionocytes with unique biologic functions and a common rare cell progenitor of ionocytes, tuft, and neuroendocrine cells. Thus, rare pulmonary ionocytes perform critical CFTR-dependent functions in the proximal airway that are hallmark features of CF airway disease. These studies provide a road map for using conditional genetics in the first non- rodent mammal to address gene function, cell biology, and disease processes that have greater evolutionary conservation between humans and ferrets.
2023-08-01 | GSE233654 | GEO
Project description:Early life viral infections are responsible for pulmonary exacerbations that can contribute to disease progression in young children with CF. The most common respiratory viruses detected in the CF airway are human rhinoviruses (RV) and susceptibility to infection has been attributed to dysregulated airway epithelial responses, although evidence has been conflicting. Here, we exposed airway epithelial cells from children with and without CF to RV in vitro. Using RNA-Seq, we profiled the transcriptomic differences of CF and non-CF airway epithelial cells at baseline and in response to RV. There were only modest differences between CF and non-CF cells at baseline. In response to RV there were 1442 and 896 differentially expressed genes in CF and non-CF airway epithelial cells respectively. The core antiviral responses in CF and non-CF airway epithelial cells were mediated through interferon signaling although type 1 and 3 interferon proteins were reduced in CF airway epithelial cells following viral challenge consistent with previous reports. The transcriptional responses in CF airway epithelial cells were more complex than in non-CF airway epithelial cells with more over-represented biological pathways ranging from cytokine signaling to metabolic and biosynthetic pathways. Network analysis highlighted that the differentially expressed genes of CF airway epithelial cells transcriptional responses were highly interconnected and formed a more complex network than observed in non-CF airway epithelial cells. These data provide novel insights to the CF airway epithelial cells responses to RV infection and highlight potential pathways that could be targeted to improve antiviral and anti-inflammatory responses in CF.
2020-07-03 | GSE138167 | GEO