Identification and Validation of Biomarkers in the Development and Progression of Graves' Ophthalmopathy.

Abstract

Purpose Graves' ophthalmopathy (GO) is the most common extrathyroidal manifestation of Graves' disease. This study aimed to identify immune-related genes involved in GO pathogenesis and evaluate their potential as peripheral blood biomarkers for early diagnosis and disease severity assessment. Methods Bulk RNA sequencing was performed on peripheral blood mononuclear cells from 16 GO patients and six healthy controls. Differential expression, functional enrichment, weighted gene co-expression network analysis (WGCNA), and immune infiltration analyses were conducted. Key genes were validated by quantitative real-time PCR and flow cytometry in independent cohorts. Results A total of 2858 differentially expressed genes (DEGs) were identified, with significant enrichment in immune-related pathways. Integrating DEGs, WGCNA modules, and pathway analyses identified 90 candidate hub genes. Among these, XAF1, SAMD9L, and CD2 showed high diagnostic accuracy for GO. Notably, only CX3CR1 and IL-7R expression correlated significantly with disease severity; IL-7R was significantly downregulated after immunosuppressive therapy in responsive patients. Flow cytometry confirmed that IL-7R expression in CD4+ T cells and memory B cells was positively associated with GO severity. Conclusions This study identifies IL-7R and CX3CR1 as novel severity-associated biomarkers in GO, providing insights into its immunopathogenesis and offering potential peripheral blood-based targets for diagnosis, prognosis, and therapy.