Multimodal Immune Profiling in Thyroid Eye Disease

Introduction/Rationale: Thyroid Eye Disease (TED) is an autoimmune orbital disorder affecting up to half of patients with Graves’ disease. It progresses from inflammation to fibrosis, leading to proptosis, diplopia, and tissue remodeling. While fibroblast diversity is known to influence disease course, the interplay between immune and stromal compartments remains poorly understood. Here, we apply integrated single-cell and TCR sequencing to define cellular programs and immune–stromal interactions across disease stages in TED.

Methods: Orbital tissue was obtained during decompression surgery from twelve TED patients (active: Clinical Activity Score ≥3, n=6; inactive: CAS < 3, n=6) and two non-inflammatory controls. Single-cell RNA and paired V(D)J sequencing characterized cellular heterogeneity and clonality. Differential expression, pathway enrichment, and ligand–receptor interaction analyses were performed to identify mechanisms associated with disease activity and fibrosis.

Results: Analysis revealed coordinated immune and fibroblast remodeling across disease stages. Active TED was marked by inflammatory–adipogenic PLIN2⁺/TNFSF8⁺ fibroblasts, elevated cytokine signaling, and expanded CD8⁺ T cell clones expressing activation markers (REL, NFKB1, CD69). Inactive TED showed a shift toward fibrotic and contractile fibroblast states (SPARC, GPX3, TAGLN) and reduced T cell clonality. Cross-talk modeling identified a RORA⁺ fibroblast subset co-expressing TSHR and IGF1R, potentially mediating T cell–fibroblast signaling via ELL2-associated pathways.

Conclusion: This study provides a multimodal framework linking immune activation with stromal remodeling in TED. By integrating transcriptional and clonotypic data, we uncover immune–stromal circuits that define inflammatory and fibrotic phases, revealing candidate targets for stage-specific intervention.