Unknown,Transcriptomics,Genomics,Proteomics

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New validated reference genes for gene expression studies in the differentiating human airway epithelium

ABSTRACT: The aim of this study was to identify stably expressed genes suitable for RT-qPCR normalization in an in vitro model of human airway epithelium (AE). RNA sequencing was performed to characterize global gene expression profiles across AE cultures representing various sexes, AE origin (bronchial or nasal) and stages of epithelial differentiation. Strand-specific poly(A)-selected libraries were prepared using the TruSeq Stranded mRNA protocol and sequenced at Macrogen Europe BV (Amsterdam, Netherlands) on the Illumina NovaSeq 6000 platform in paired-end mode (2 × 150 bp), generating approximately 100 million reads per sample. The deposited files include raw paired-end FASTQ data generated from 2 primary and 27 human AE cells differentiated in vitro in ALI (air-liquid interface) culture. These data were used for transcriptome-wide assessment of gene expression stability and selection of candidate reference genes for downstream validation.

INSTRUMENT(S): Illumina NovaSeq 6000

ORGANISM(S): Homo sapiens

SUBMITTER: Joanna Jurczak

PROVIDER: E-MTAB-16674 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Background: In asthma, airway epithelium remodeling can already be detected during childhood, and epithelial cells are more susceptible to virus and oxidative stress. Their exact role in natural history and severity of children allergic respiratory disease remains however surprisingly unexplored. Aim: To analyze dysfunctions of epithelium in dust mite allergic respiratory disease (rhinitis ¬± asthma) in children. Methods: Expression profilings of nasal epithelial cells collected by brushing were performed on Affymetrix Hugene 1.0 ST arrays. All allergic patients were sensitized to dust mite. 19 patients had an isolated allergic rhinitis (AR). 14 patients had AR associated with asthma. Patients were compared to 12 controls, their severity and control being assessed according to NAEPP and ARIA criteria. Infections by respiratory viruses were excluded by real-time PCR measurements. Results: 61 probes were able to distinguish allergic rhinitis children from healthy controls. A majority of these probes was under the control of Th2 cytokines, as evidenced by parallel experiments performed on primary cultures of nasal epithelial cells. In uncontrolled asthmatic patients, we observed not only an enhanced expression of these Th2-responsive transcripts, but also a down-regulation of interferon-responsive genes. Conclusion: Our study identifies a Th2 driven epithelial phenotype common to all dust mite allergic children. Besides, it suggests that epithelium is involved in the severity of the disease. Expression profiles observed in uncontrolled asthmatic patients suggest that severity of asthma is linked at the same time to atopy and to impaired viral response. Nasal epithelium gene expression profiling of dust mite allergic children with isolated rhinitis, rhinitis associated with asthma and controls. 38 samples classified in 4 categories : 14 isolated rhinitis (R), 6 rhinitis with uncontrolled asthma (UA), 7 rhinitis with controlled asthma (CA) and 11 healthy subjects (C )

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New validated reference genes for gene expression studies in the differentiating human airway epithelium

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