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In vitro platform to model the function of ionocytes in the human airway epithelium.

  • Vilà-González, Marta
  • Pinte, Laetitia
  • Fradique, Ricardo
  • Causa, Erika
  • Kool, Heleen
  • Rodrat, Mayuree
  • Morell, Carola Maria
  • Al-Thani, Maha
  • Porter, Linsey
  • Guo, Wenrui
  • Maeshima, Ruhina
  • Hart, Stephen L
  • McCaughan, Frank
  • Granata, Alessandra
  • Sheppard, David N
  • Floto, R Andres
  • Rawlins, Emma L
  • Cicuta, Pietro
  • Vallier, Ludovic
Publication Date
Apr 16, 2024
Apollo - University of Cambridge Repository
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BACKGROUND: Pulmonary ionocytes have been identified in the airway epithelium as a small population of ion transporting cells expressing high levels of CFTR (cystic fibrosis transmembrane conductance regulator), the gene mutated in cystic fibrosis. By providing an infinite source of airway epithelial cells (AECs), the use of human induced pluripotent stem cells (hiPSCs) could overcome some challenges of studying ionocytes. However, the production of AEC epithelia containing ionocytes from hiPSCs has proven difficult. Here, we present a platform to produce hiPSC-derived AECs (hiPSC-AECs) including ionocytes and investigate their role in the airway epithelium. METHODS: hiPSCs were differentiated into lung progenitors, which were expanded as 3D organoids and matured by air-liquid interface culture as polarised hiPSC-AEC epithelia. Using CRISPR/Cas9 technology, we generated a hiPSCs knockout (KO) for FOXI1, a transcription factor that is essential for ionocyte specification. Differences between FOXI1 KO hiPSC-AECs and their wild-type (WT) isogenic controls were investigated by assessing gene and protein expression, epithelial composition, cilia coverage and motility, pH and transepithelial barrier properties. RESULTS: Mature hiPSC-AEC epithelia contained basal cells, secretory cells, ciliated cells with motile cilia, pulmonary neuroendocrine cells (PNECs) and ionocytes. There was no difference between FOXI1 WT and KO hiPSCs in terms of their capacity to differentiate into airway progenitors. However, FOXI1 KO led to mature hiPSC-AEC epithelia without ionocytes with reduced capacity to produce ciliated cells. CONCLUSION: Our results suggest that ionocytes could have role beyond transepithelial ion transport by regulating epithelial properties and homeostasis in the airway epithelium. / This work was supported by the Wellcome Trust Sir Henry Wellcome Postdoctoral Fellowship (218663/Z/19/Z to MVG), the UK Cystic Fibrosis Trust (IH001 to MVG, RAF; SRC 016 to LPi, RF, PC; SRC 021 to DNS; VIA 028 to HK, ELR; RM, SLH), the EU ITN PhyMot (to EC), the European Research Council Grant New-Chol (ERC: 741707 to LV), the MRC (MR/P009581/1 to HK, ELR), the Medical Research Foundation Fellowship (to MA-T, AG), the NC3Rs (Training Fellowship NC/R001987/1 to CMM; Project Grant NC/S001204/1 to LPo, WG, FM), the Roy Castle Lung Cancer Foundation grant (2015/10/McCaughan to LPo, WG, FM), the Cystic Fibrosis Foundation (to RM, SLH) and the core support grant from the Wellcome Trust (203151/Z/16/Z) and the UKRI Medical Research Council (MC_PC_17230) for the Wellcome MRC – Cambridge Stem Cell Institute. This paper presents independent research supported by the NIHR Cambridge BRC. The NIHR Cambridge Biomedical Research Centre (BRC) is a partnership between Cambridge University Hospitals NHS Foundation Trust and the University of Cambridge, funded by the National Institute for Health Research (NIHR). The views expressed are those of the author(s) and not necessarily those of the NIHR or the Department of Health and Social Care.

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