Transcriptome analysis of ciliary differentiation in airway epithelium
Transcriptome analysis of ciliary differentiation in airway epithelium
Fresh nasal brushings and air-liquid interface (ALI) cultures of nasal epithelial cells are frequently used as a model for airways research but the process of cell differentiation and ciliation in ALI cultures is poorly understood. We aimed to characterise ALI culture, in terms of gene expression and cell phenotype, over 63 days.
ALI cultures of three healthy volunteers were harvested at 1, 4, 8, 14, 21, 28 and 63 days. We assessed the transcriptome and morphology of ciliary differentiation using RNA-seq, high-speed video microscopy (HSVM) and scanning electron microscopy (SEM). Specific gene markers were used to assess cell type changes throughout the culturing process. Differential gene expression analysis and functional enrichment analysis were done to identify changes in biological processes during culturing. Furthermore, the transcriptomes of ALI-cultures were compared to ‘fresh’ nasal epithelium in RNAlater.
We present the ciliary differentiation from proliferate (day 4), to deuterosomal (day 8), to multi-ciliated cells (day 14) in ALI culture. Motile cilia were first detected on day 7 with HSVM, and SEM showed widespread ciliation from day 24 onwards. In addition, we present changes in biological processes involved with e.g. the cell cycle (day 4) and cilium assembly (day 8). It was found that ‘fresh’ nasal epithelium in RNAlater showed more similarity with ALI cultures from day 14 onwards.
This work gives insights into the transcriptomic changes and ciliary differentiation throughout ALI cell culturing providing the timepoint of fully differentiated multi-ciliated cells. In addition, we determined which ALI culture time point showed the most in vivo similarity compared to nasal brushings.
Legebeke, Jelmer
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Horton, Katie
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Wheway, Gabrielle
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Wai, Htoo
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Jackson, Claire
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Coles, Janice
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Holloway, John
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Lucas, Jane
5cb3546c-87b2-4e59-af48-402076e25313
Baralle, Diana
faac16e5-7928-4801-9811-8b3a9ea4bb91
Legebeke, Jelmer
f6062b8c-22ac-465c-9528-3bac881137d0
Horton, Katie
64d9e720-63d4-48c2-8663-e8ccb16f5b6f
Wheway, Gabrielle
2e547e5d-b921-4243-a071-2208fd4cc090
Wai, Htoo
4428517b-33b3-42cb-9818-ca64763ab7bc
Jackson, Claire
64cdd6fa-74c3-4ac6-94ef-070620a6efd9
Coles, Janice
fb9d20aa-93b9-42b3-9b9e-bab2f565ea60
Holloway, John
4bbd77e6-c095-445d-a36b-a50a72f6fe1a
Lucas, Jane
5cb3546c-87b2-4e59-af48-402076e25313
Baralle, Diana
faac16e5-7928-4801-9811-8b3a9ea4bb91
Legebeke, Jelmer, Horton, Katie, Wheway, Gabrielle, Wai, Htoo, Jackson, Claire, Coles, Janice, Holloway, John, Lucas, Jane and Baralle, Diana
(2021)
Transcriptome analysis of ciliary differentiation in airway epithelium.
European Respiratory Journal, 58 (63), [PA2398].
(doi:10.1183/13993003.congress-2021.PA2398).
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Meeting abstract
Abstract
Fresh nasal brushings and air-liquid interface (ALI) cultures of nasal epithelial cells are frequently used as a model for airways research but the process of cell differentiation and ciliation in ALI cultures is poorly understood. We aimed to characterise ALI culture, in terms of gene expression and cell phenotype, over 63 days.
ALI cultures of three healthy volunteers were harvested at 1, 4, 8, 14, 21, 28 and 63 days. We assessed the transcriptome and morphology of ciliary differentiation using RNA-seq, high-speed video microscopy (HSVM) and scanning electron microscopy (SEM). Specific gene markers were used to assess cell type changes throughout the culturing process. Differential gene expression analysis and functional enrichment analysis were done to identify changes in biological processes during culturing. Furthermore, the transcriptomes of ALI-cultures were compared to ‘fresh’ nasal epithelium in RNAlater.
We present the ciliary differentiation from proliferate (day 4), to deuterosomal (day 8), to multi-ciliated cells (day 14) in ALI culture. Motile cilia were first detected on day 7 with HSVM, and SEM showed widespread ciliation from day 24 onwards. In addition, we present changes in biological processes involved with e.g. the cell cycle (day 4) and cilium assembly (day 8). It was found that ‘fresh’ nasal epithelium in RNAlater showed more similarity with ALI cultures from day 14 onwards.
This work gives insights into the transcriptomic changes and ciliary differentiation throughout ALI cell culturing providing the timepoint of fully differentiated multi-ciliated cells. In addition, we determined which ALI culture time point showed the most in vivo similarity compared to nasal brushings.
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e-pub ahead of print date: 25 November 2021
Identifiers
Local EPrints ID: 481553
URI: http://eprints.soton.ac.uk/id/eprint/481553
ISSN: 0903-1936
PURE UUID: 42e3d0c1-6ada-44a5-8039-9701e518554a
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Date deposited: 01 Sep 2023 17:01
Last modified: 18 Mar 2024 03:49
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Author:
Jelmer Legebeke
Author:
Katie Horton
Author:
Htoo Wai
Author:
Claire Jackson
Author:
Janice Coles
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