Temporal whole-transcriptomic analysis of characterized in vitro and ex vivo primary nasal epithelia
Temporal whole-transcriptomic analysis of characterized in vitro and ex vivo primary nasal epithelia
Air-liquid interface (ALI) cell culture of primary airway progenitors enables the differentiation and recapitulation of a pseudostratified epithelium in vitro, providing a highly useful tool for researching respiratory health and disease. Previous studies into gene expression in ALI-cultures compared to ex vivo nasal brushings have been limited in the number of time points and/or the number of genes studied. In this study physiological and global transcriptomic changes were assessed in an extended in vitro 63-day human healthy nasal epithelium ALI-culture period and compared to ex vivo nasal brushing samples. Ex vivo nasal brushing samples formed distinct transcriptome clusters to in vitro ALI-cultured nasal epithelia, with from day 14 onwards ALI samples best matching the ex vivo samples. Immune response regulation genes were not expressed in the in vitro ALI-culture compared to the ex vivo nasal brushing samples, likely because the in vitro cultures lack an airway microbiome, lack airborne particles stimulation, or did not host an immune cell component. This highlights the need for more advanced co-cultures with immune cell representation to better reflect the physiological state. During the first week of ALI-culture genes related to metabolism and proliferation were increased. By the end of week 1 epithelial cell barrier function plateaued and multiciliated cell differentiation started, although widespread ciliation was not complete until day 28. These results highlight that time-points at which ALI-cultures are harvested for research studies needs to be carefully considered to suit the purpose of investigation (transcriptomic and/or functional analysis).
air-liquid interface culture, airway cilia, physiological analysis, primary nasal epithelium, whole transcriptome analysis
907511
Legebeke, Jelmer
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Horton, Katie Leanne
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Jackson, Claire
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Coles, Janice
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Harris, Amanda
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Wai, Htoo
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Holloway, John
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Wheway, Gabrielle
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Baralle, Diana
faac16e5-7928-4801-9811-8b3a9ea4bb91
Lucas, Jane
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15 June 2022
Legebeke, Jelmer
f6062b8c-22ac-465c-9528-3bac881137d0
Horton, Katie Leanne
0e8b1fe0-65ae-41d2-815e-d8ee76ee9433
Jackson, Claire
64cdd6fa-74c3-4ac6-94ef-070620a6efd9
Coles, Janice
fb9d20aa-93b9-42b3-9b9e-bab2f565ea60
Harris, Amanda
c6eaa1d1-daa7-4229-bcd0-08eb840410fd
Wai, Htoo
4428517b-33b3-42cb-9818-ca64763ab7bc
Holloway, John
4bbd77e6-c095-445d-a36b-a50a72f6fe1a
Wheway, Gabrielle
2e547e5d-b921-4243-a071-2208fd4cc090
Baralle, Diana
faac16e5-7928-4801-9811-8b3a9ea4bb91
Lucas, Jane
5cb3546c-87b2-4e59-af48-402076e25313
Legebeke, Jelmer, Horton, Katie Leanne, Jackson, Claire, Coles, Janice, Harris, Amanda, Wai, Htoo, Holloway, John, Wheway, Gabrielle, Baralle, Diana and Lucas, Jane
(2022)
Temporal whole-transcriptomic analysis of characterized in vitro and ex vivo primary nasal epithelia.
Frontiers in Cell and Developmental Biology, 10, , [907511].
(doi:10.3389/fcell.2022.907511).
Abstract
Air-liquid interface (ALI) cell culture of primary airway progenitors enables the differentiation and recapitulation of a pseudostratified epithelium in vitro, providing a highly useful tool for researching respiratory health and disease. Previous studies into gene expression in ALI-cultures compared to ex vivo nasal brushings have been limited in the number of time points and/or the number of genes studied. In this study physiological and global transcriptomic changes were assessed in an extended in vitro 63-day human healthy nasal epithelium ALI-culture period and compared to ex vivo nasal brushing samples. Ex vivo nasal brushing samples formed distinct transcriptome clusters to in vitro ALI-cultured nasal epithelia, with from day 14 onwards ALI samples best matching the ex vivo samples. Immune response regulation genes were not expressed in the in vitro ALI-culture compared to the ex vivo nasal brushing samples, likely because the in vitro cultures lack an airway microbiome, lack airborne particles stimulation, or did not host an immune cell component. This highlights the need for more advanced co-cultures with immune cell representation to better reflect the physiological state. During the first week of ALI-culture genes related to metabolism and proliferation were increased. By the end of week 1 epithelial cell barrier function plateaued and multiciliated cell differentiation started, although widespread ciliation was not complete until day 28. These results highlight that time-points at which ALI-cultures are harvested for research studies needs to be carefully considered to suit the purpose of investigation (transcriptomic and/or functional analysis).
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907511_Manuscript
- Accepted Manuscript
More information
Accepted/In Press date: 12 May 2022
Published date: 15 June 2022
Additional Information:
Funding Information:
The National PCD Centre in Southampton is commissioned and funded by NHS England; PCD research is supported by National Institute for Health Research (NIHR) Southampton Biomedical Research Centre (BRC), Southampton Biomedical Imaging Unit, NIHR Southampton Clinical Research Facility, National Institute for Health Research (RfPB PB-PG-1215-20014; and 200470) and The AAIR Charity (Reg. No. 1129698). KH is funded by Wessex Medical Research and NIHR Southampton BRC (NIHR-INF-0010). JL is funded by NIHR Southampton BRC (NIHR-INF-0932). The Baralle Laboratory was supported by NIHR Research Professorship to DB (RP-2016-07-011).
Publisher Copyright:
Copyright © 2022 Legebeke, Horton, Jackson, Coles, Harris, Wai, Holloway, Wheway, Baralle and Lucas.
Keywords:
air-liquid interface culture, airway cilia, physiological analysis, primary nasal epithelium, whole transcriptome analysis
Identifiers
Local EPrints ID: 457586
URI: http://eprints.soton.ac.uk/id/eprint/457586
ISSN: 2296-634X
PURE UUID: fbf5d447-2f44-4488-937d-4259828f7011
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Date deposited: 13 Jun 2022 16:48
Last modified: 17 Mar 2024 03:53
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Contributors
Author:
Jelmer Legebeke
Author:
Katie Leanne Horton
Author:
Claire Jackson
Author:
Janice Coles
Author:
Amanda Harris
Author:
Htoo Wai
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