Analysis of N6-methyladenosine modification and immune infiltration characterization in thyroid associated ophthalmopathy

Abstract

Abstract Background Thyroid-associated ophthalmopathy (TAO) is a potentially sight-threatening ocular disease that affects 50% of patients with Graves’ disease. N6-methyladenosine (m6A) modification is post-transcriptional RNA modification in mammalian mRNA. Exploring the role of m6A regulators and the immune infiltration characterization in TAO is essential to understand its pathogenesis. Methods We excavated differentially expressed m6A regulators from the TAO microarray data set (GSE58331) in the Gene Expression Omnibus (GEO). Based on R software (v4.2.2), we performed bioinformatics analyses including differential expression analysis, training models construction such as support vector machine (SVM), random forest (RF), and nomogram models for the evaluation of TAO occurrence, consensus clustering algorithm for the evaluation of clusters quantity, principal component analysis (PCA) for quantification of m6A modification level in individual patients, Gene Ontology (GO) functional annotation, Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis, and single sample gene set enrichment analysis (ssGSEA) for functional enrichment analysis, etc. Results Four significant m6A regulators (IGFBP2, IGFBP3, RBM15B, and FTO) were screened by differential expression analysis between the TAO group and the healthy group. RM identified that the four m6A regulators above were associated with the risk of TAO. Furthermore, a nomogram was conducted to provide benefits to patients. Based on filtered m6A regulators, we divided the TAO group into two m6A subtypes (cluster A/B) by consensus clustering. The PCA algorithm calculated the m6A score of each sample to quantify m6A patterns. GO functional annotation proved that m6A-related differentially expressed genes (DEGs) enriched in biological processes (BP), cellular components (CC), and molecular functions (MF). KEGG enrichment analysis showed m6A-related DEGs were particularly abundant in the endocytosis, protein processing in endoplasmic reticulum, ubiquitin mediated proteolysis, and protein export. ssGSEA presented the difference in immune infiltration between m6Acluster A/B and gene cluster A/B. In addition, we also analyzed the relationship between m6Aclusters or gene clusters and cytokines. Conclusions This article analyzed the effect of m6A modification in the pathogenesis of TAO, focusing on immune infiltration. These findings may be able to guide the future strategy of immunotherapy.