Dataset Information

Proto-oncogene mutations in middle ear cholesteatoma contribute to its pathogenesis.

ABSTRACT:

Background

Chronic inflammation causes bone destruction in middle ear cholesteatomas (MECs). However, the causes of their neoplastic features remain unknown. The present study demonstrated for the first time that neoplastic features of MEC are based on proto-oncogene mutations.

Results

DNA was extracted from MEC and blood samples of five patients to detect somatic mutations using depth-depth exome sequencing. Exons with somatic variants were analyzed using an additional 17 MEC/blood test pairs. Variants detected in MECs but not in blood were considered pathogenic variant candidates. We analyzed the correlation between proto-oncogene (NOTCH1 and MYC) variants and the presence of bone destruction and granulation tissue formation. MYC and NOTCH1 variants were detected in two and five of the 22 samples, respectively. Two of the NOTCH1 variants were located in its specific functional domain, one was truncating and the other was a splice donor site variant. Mutations of the two genes in attic cholesteatomas (n = 14) were significantly related with bone destruction (p = 0.0148) but not with granulation tissue formation (p = 0.399).

Conclusions

This is the first study to demonstrate a relationship between neoplastic features of MEC and proto-oncogene mutations.

SUBMITTER: Satoh C

PROVIDER: S-EPMC10647096 | biostudies-literature | 2023 Nov

REPOSITORIES: biostudies-literature

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Publications

Proto-oncogene mutations in middle ear cholesteatoma contribute to its pathogenesis.

Satoh Chisei C Yoshiura Koh-Ichiro KI Mishima Hiroyuki H Yoshida Haruo H Takahashi Haruo H Kumai Yoshihiko Y

BMC medical genomics 20231115 1

<h4>Background</h4>Chronic inflammation causes bone destruction in middle ear cholesteatomas (MECs). However, the causes of their neoplastic features remain unknown. The present study demonstrated for the first time that neoplastic features of MEC are based on proto-oncogene mutations.<h4>Results</h4>DNA was extracted from MEC and blood samples of five patients to detect somatic mutations using depth-depth exome sequencing. Exons with somatic variants were analyzed using an additional 17 MEC/blo ...[more]

PMID: 37968650

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Project description:IntroductionMiddle ear cholesteatoma is a chronic middle ear disease characterized by severe hearing loss and adjacent bone erosion, resulting in numerous complications. This study sought to identify pathways involved in N6-methyladenosine (m6A) modification of circRNA in middle ear cholesteatoma.MethodsA m6A circRNA epitranscriptomic microarray analysis was performed in middle ear cholesteatoma tissues (n = 5) and normal post-auricular skin samples (n = 5). Bioinformatics analyses subsequently explored the biological functions (Gene Ontology, GO) and signaling pathways (Kyoto Encyclopedia of Genes and Genomes, KEGG) underlying middle ear cholesteatoma pathogenesis. Methylated RNA immunoprecipitation qPCR (MeRIP-qPCR) was performed to verify the presence of circRNAs with m6A modifications in middle ear cholesteatoma and normal skin samples.ResultsMicroarray analysis identified 3,755 circRNAs as significantly differentially modified by m6A methylation in middle ear cholesteatoma compared with the normal post-auricular skin. Among these, 3,742 were hypermethylated (FC ≥ 2, FDR < 0.05) and 13 were hypomethylated (FC ≤ 1/2, FDR < 0.05). GO analysis terms with the highest enrichment score were localization, cytoplasm, and ATP-dependent activity for biological processes, cellular components, and molecular functions respectively. Of the eight hypermethylated circRNA pathways, RNA degradation pathway has the highest enrichment score. Peroxisome Proliferator-Activated Receptor (PPAR) signaling pathway was hypomethylated. To validate the microarray analysis, we conducted MeRIP-qPCR to assess the methylation levels of five specific m6A-modified circRNAs: hsa_circRNA_061554, hsa_circRNA_001454, hsa_circRNA_031526, hsa_circRNA_100833, and hsa_circRNA_022382. The validation was highly consistent with the findings from the microarray analysis.ConclusionOur study firstly presents m6A modification patterns of circRNAs in middle ear cholesteatoma. This finding suggests a direction for circRNA m6A modification research in the etiology of cholesteatoma and provides potential therapeutic targets for the treatment of middle ear cholesteatoma.

Project description:BackgroundCholesteatoma disease is an expanding lesion in the middle ear. Hearing loss and facial paralysis alongside with other intracranial complications are found. No pharmaceutical treatment is available today and recurrence after surgical extraction occurs. We investigated possible TLR4-based mechanisms promoting recurrence and explore possible treatments strategies.MethodsWe isolated fibroblasts and epidermal stem cells from cholesteatoma tissue and healthy auditory canal skin. Subsequently, their expression under standard culture conditions and after stimulation with LPS was investigated by RT-qPCR. Cell metabolism and proliferation were analysed upon LPS treatment, with and without TLR4 antagonist. An indirect co-culture of fibroblasts and epidermal stem cells isolated from cholesteatoma tissue was utilized to monitor epidermal differentiation upon LPS treatment by RT-qPCR and immunocytochemistry.ResultsUnder standard culture conditions, we detected a tissue-independent higher expression of IL-1β and IL-8 in stem cells, an upregulation of KGF and IGF-2 in both cell types derived from cholesteatoma and higher expression of TLR4 in stem cells derived from cholesteatoma tissue. Upon LPS challenge, we could detect a significantly higher expression of IL-1α, IL-1β, IL-6 and IL-8 in stem cells and of TNF-a, GM-CSF and CXCL-5 in stem cells and fibroblasts derived from cholesteatoma. The expression of the growth factors KGF, EGF, EREG, IGF-2 and HGF was significantly higher in fibroblasts, particularly when derived from cholesteatoma. Upon treatment with LPS the metabolism was elevated in stem cells and fibroblasts, proliferation was only enhanced in fibroblasts derived from cholesteatoma. This could be reversed by the treatment with a TLR4 antagonist. The cholesteatoma fibroblasts could be triggered by LPS to promote the epidermal differentiation of the stem cells, while no LPS treatment or LPS treatment without the presence of fibroblasts did not result in such a differentiation.ConclusionWe propose that cholesteatoma recurrence is based on TLR4 signalling imprinted in the cholesteatoma cells. It induces excessive inflammation of stem cells and fibroblasts, proliferation of perimatrix fibroblasts and the generation of epidermal cells from stem cells thru paracrine signalling by fibroblasts. Treatment of the operation site with a TLR4 antagonist might reduce the chance of cholesteatoma recurrence. Video Abstract.

Dataset Information

Proto-oncogene mutations in middle ear cholesteatoma contribute to its pathogenesis.

Background

Results

Conclusions

Publications

Proto-oncogene mutations in middle ear cholesteatoma contribute to its pathogenesis.

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