Comparison of air-liquid interface transwell and airway organoid models for human respiratory virus infection studies
IntroductionComplex in vitro respiratory models, including air-liquid interface (ALI) transwell cultures and airway organoids, have emerged as promising tools for studying human respiratory virus infections. These models address several limitations of conventional two-dimensional cell line and anim...
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Frontiers Media S.A.
2025-02-01
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| author | Camilla T. Ekanger Camilla T. Ekanger Camilla T. Ekanger Nilima Dinesh Kumar Rosanne W. Koutstaal Fan Zhou Martin Beukema Joanna Waldock Simon P. Jochems Noa Mulder Cécile A. C. M. van Els Cécile A. C. M. van Els Othmar G. Engelhardt Nathalie Mantel Kevin P. Buno Karl Albert Brokstad Karl Albert Brokstad Agnete S. T. Engelsen Agnete S. T. Engelsen Rebecca J. Cox Rebecca J. Cox Barbro N. Melgert Barbro N. Melgert Anke L. W. Huckriede Puck B. van Kasteren |
| author_facet | Camilla T. Ekanger Camilla T. Ekanger Camilla T. Ekanger Nilima Dinesh Kumar Rosanne W. Koutstaal Fan Zhou Martin Beukema Joanna Waldock Simon P. Jochems Noa Mulder Cécile A. C. M. van Els Cécile A. C. M. van Els Othmar G. Engelhardt Nathalie Mantel Kevin P. Buno Karl Albert Brokstad Karl Albert Brokstad Agnete S. T. Engelsen Agnete S. T. Engelsen Rebecca J. Cox Rebecca J. Cox Barbro N. Melgert Barbro N. Melgert Anke L. W. Huckriede Puck B. van Kasteren |
| author_sort | Camilla T. Ekanger |
| collection | DOAJ |
| description | IntroductionComplex in vitro respiratory models, including air-liquid interface (ALI) transwell cultures and airway organoids, have emerged as promising tools for studying human respiratory virus infections. These models address several limitations of conventional two-dimensional cell line and animal models. However, the lack of standardized protocols for the application of these models in infection studies limits the possibilities for comparing results across different research groups. Therefore, we applied a collaborative approach to harmonize several aspects of experimental methodology between different research laboratories, aiming to assess the comparability of different models of human airway epithelium in the context of respiratory viral infections.MethodsIn this study, we compared three different models of human respiratory epithelium: a primary human bronchial epithelial cell-derived ALI transwell model, and two airway organoid models established from human airway- and lung-derived adult stem cells. We first assessed the presence of various differentiated cell types using immunofluorescence microscopy. Using a shared stock of influenza A virus, we then assessed viral growth kinetics, epithelial cytokine responses, and serum-mediated inhibition of infection.ResultsThe presence of club, goblet, and ciliated cells was confirmed in all models. We observed similar viral replication kinetics with a >4-log increase in virus titre across all models using a TCID50 assay. Following infection, a reproducible antiviral cytokine response, including a consistent increase in CXCL10, IL-6, IFN-λ1, IFN-λ2/3, and IFN-β, was detected across all models. Finally, neutralization was assessed by pre-incubation of virus with human serum. Reduced viral replication was observed across all models, resulting in a 3- to 6-log decrease in virus titres as quantified by TCID50.DiscussionIn conclusion, all three models produced consistent results regardless of the varying cell sources, culturing approaches, and infection methods. Our collaborative efforts to harmonize infection experiments and compare ALI transwell and airway organoid models described here aid in advancing our understanding and improving the standardization of these complex in vitro respiratory models for future studies. |
| format | Article |
| id | doaj-art-58f22a426f6a48649e6b3149113b6e98 |
| institution | Kabale University |
| issn | 1664-3224 |
| language | English |
| publishDate | 2025-02-01 |
| publisher | Frontiers Media S.A. |
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| series | Frontiers in Immunology |
| spelling | doaj-art-58f22a426f6a48649e6b3149113b6e982025-02-06T07:10:09ZengFrontiers Media S.A.Frontiers in Immunology1664-32242025-02-011610.3389/fimmu.2025.15321441532144Comparison of air-liquid interface transwell and airway organoid models for human respiratory virus infection studiesCamilla T. Ekanger0Camilla T. Ekanger1Camilla T. Ekanger2Nilima Dinesh Kumar3Rosanne W. Koutstaal4Fan Zhou5Martin Beukema6Joanna Waldock7Simon P. Jochems8Noa Mulder9Cécile A. C. M. van Els10Cécile A. C. M. van Els11Othmar G. Engelhardt12Nathalie Mantel13Kevin P. Buno14Karl Albert Brokstad15Karl Albert Brokstad16Agnete S. T. Engelsen17Agnete S. T. Engelsen18Rebecca J. Cox19Rebecca J. Cox20Barbro N. Melgert21Barbro N. Melgert22Anke L. W. Huckriede23Puck B. van Kasteren24Department of Biomedicine, Faculty of Medicine, University of Bergen, Bergen, NorwayCentre for Cancer Biomarkers (CCBIO), Department of Clinical Medicine, University of Bergen, Bergen, NorwayInfluenza Centre, Department of Clinical Science, University of Bergen, Bergen, NorwayDepartment of Medical Microbiology and Infection Prevention, University Medical Center Groningen, University of Groningen, Groningen, NetherlandsCenter for Infectious Disease Control, National Institute for Public Health and the Environment (RIVM), Bilthoven, NetherlandsInfluenza Centre, Department of Clinical Science, University of Bergen, Bergen, NorwayDepartment of Medical Microbiology and Infection Prevention, University Medical Center Groningen, University of Groningen, Groningen, NetherlandsInfluenza Resource Centre, Vaccines, Science Research & Innovation, Medicines and Healthcare Products Regulatory Agency (MHRA), Potters Bar, United KingdomLeiden University Center for Infectious Diseases (LU-CID), Leiden University Medical Center, Leiden, NetherlandsGroningen Research Institute for Pharmacy, Department of Molecular Pharmacology, University of Groningen, Groningen, NetherlandsCenter for Infectious Disease Control, National Institute for Public Health and the Environment (RIVM), Bilthoven, NetherlandsSection Immunology, Department Biomolecular Health Sciences, Faculty of Veterinary Medicine, Utrecht University, Utrecht, NetherlandsInfluenza Resource Centre, Vaccines, Science Research & Innovation, Medicines and Healthcare Products Regulatory Agency (MHRA), Potters Bar, United Kingdom0Sanofi, Marcy L’Etoile, France1GlaxoSmithKline (GSK), Siena, ItalyInfluenza Centre, Department of Clinical Science, University of Bergen, Bergen, Norway2Department of Safety, Chemistry and Biomedical Laboratory Sciences, Western Norway University of Applied Sciences, Bergen, NorwayDepartment of Biomedicine, Faculty of Medicine, University of Bergen, Bergen, NorwayCentre for Cancer Biomarkers (CCBIO), Department of Clinical Medicine, University of Bergen, Bergen, NorwayInfluenza Centre, Department of Clinical Science, University of Bergen, Bergen, Norway3Department of Microbiology, Haukeland University Hospital, Bergen, NorwayGroningen Research Institute for Pharmacy, Department of Molecular Pharmacology, University of Groningen, Groningen, Netherlands4Groningen Research Institute for Asthma and COPD, University Medical Center Groningen, University of Groningen, Groningen, NetherlandsDepartment of Medical Microbiology and Infection Prevention, University Medical Center Groningen, University of Groningen, Groningen, NetherlandsCenter for Infectious Disease Control, National Institute for Public Health and the Environment (RIVM), Bilthoven, NetherlandsIntroductionComplex in vitro respiratory models, including air-liquid interface (ALI) transwell cultures and airway organoids, have emerged as promising tools for studying human respiratory virus infections. These models address several limitations of conventional two-dimensional cell line and animal models. However, the lack of standardized protocols for the application of these models in infection studies limits the possibilities for comparing results across different research groups. Therefore, we applied a collaborative approach to harmonize several aspects of experimental methodology between different research laboratories, aiming to assess the comparability of different models of human airway epithelium in the context of respiratory viral infections.MethodsIn this study, we compared three different models of human respiratory epithelium: a primary human bronchial epithelial cell-derived ALI transwell model, and two airway organoid models established from human airway- and lung-derived adult stem cells. We first assessed the presence of various differentiated cell types using immunofluorescence microscopy. Using a shared stock of influenza A virus, we then assessed viral growth kinetics, epithelial cytokine responses, and serum-mediated inhibition of infection.ResultsThe presence of club, goblet, and ciliated cells was confirmed in all models. We observed similar viral replication kinetics with a >4-log increase in virus titre across all models using a TCID50 assay. Following infection, a reproducible antiviral cytokine response, including a consistent increase in CXCL10, IL-6, IFN-λ1, IFN-λ2/3, and IFN-β, was detected across all models. Finally, neutralization was assessed by pre-incubation of virus with human serum. Reduced viral replication was observed across all models, resulting in a 3- to 6-log decrease in virus titres as quantified by TCID50.DiscussionIn conclusion, all three models produced consistent results regardless of the varying cell sources, culturing approaches, and infection methods. Our collaborative efforts to harmonize infection experiments and compare ALI transwell and airway organoid models described here aid in advancing our understanding and improving the standardization of these complex in vitro respiratory models for future studies.https://www.frontiersin.org/articles/10.3389/fimmu.2025.1532144/fullinfluenza virusmucosal modelsrespiratory tractcomplex in vitro modelsharmonization |
| spellingShingle | Camilla T. Ekanger Camilla T. Ekanger Camilla T. Ekanger Nilima Dinesh Kumar Rosanne W. Koutstaal Fan Zhou Martin Beukema Joanna Waldock Simon P. Jochems Noa Mulder Cécile A. C. M. van Els Cécile A. C. M. van Els Othmar G. Engelhardt Nathalie Mantel Kevin P. Buno Karl Albert Brokstad Karl Albert Brokstad Agnete S. T. Engelsen Agnete S. T. Engelsen Rebecca J. Cox Rebecca J. Cox Barbro N. Melgert Barbro N. Melgert Anke L. W. Huckriede Puck B. van Kasteren Comparison of air-liquid interface transwell and airway organoid models for human respiratory virus infection studies Frontiers in Immunology influenza virus mucosal models respiratory tract complex in vitro models harmonization |
| title | Comparison of air-liquid interface transwell and airway organoid models for human respiratory virus infection studies |
| title_full | Comparison of air-liquid interface transwell and airway organoid models for human respiratory virus infection studies |
| title_fullStr | Comparison of air-liquid interface transwell and airway organoid models for human respiratory virus infection studies |
| title_full_unstemmed | Comparison of air-liquid interface transwell and airway organoid models for human respiratory virus infection studies |
| title_short | Comparison of air-liquid interface transwell and airway organoid models for human respiratory virus infection studies |
| title_sort | comparison of air liquid interface transwell and airway organoid models for human respiratory virus infection studies |
| topic | influenza virus mucosal models respiratory tract complex in vitro models harmonization |
| url | https://www.frontiersin.org/articles/10.3389/fimmu.2025.1532144/full |
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