Rheumatoid arthritis (RA) is a chronic systemic autoimmune disorder characterized by persistent synovial inflammation, progressive cartilage and bone destruction, and wide-ranging immune dysregulation. The disease burden is substantial and rising globally. Beyond classical immune mechanisms, recent research implicates non-apoptotic regulated cell death pathways and disturbed metal ion homeostasis in RA pathogenesis. Two such pathways are ferroptosis — an iron-dependent form of lipid peroxidation-driven cell death — and cuproptosis — a copper-dependent form of mitochondrial proteotoxic stress. Both pathways intersect with oxidative stress and mitochondrial dysfunction, features already associated with RA synovitis and immune cell activation.
The authors applied an integrative bioinformatic pipeline to publicly available datasets to identify candidate regulators shared between ferroptosis and cuproptosis in RA. Bulk RNA-seq data from peripheral CD14+ monocytes (GSE294225) included samples from 15 healthy controls and 9 patients with active RA (DAS28 > 2.7). A single-cell RNA-seq dataset (GSE296117) was also used to map candidate gene expression within synovial cell populations.
Analytic methods combined differential expression analysis (DESeq2 criteria reported as adjusted P < 0.05 and |log2 fold change| > 1), weighted gene co-expression network analysis (WGCNA) to identify RA-associated gene modules, functional enrichment of gene sets, protein–protein interaction (PPI) network construction to identify hubs, immune cell infiltration deconvolution, and single-cell expression profiling to locate candidate genes in synovial cell types.
Using the specified thresholds, the bulk monocyte analysis yielded 1,410 significantly differentially expressed genes (DEGs) between RA and healthy samples. WGCNA identified a co-expression module associated with RA that was enriched for processes including oxidative stress response, mitochondrial dysfunction, and regulated cell death pathways. These enrichment signals provided the rationale for intersecting module genes with curated ferroptosis- and cuproptosis-related gene lists to find overlapping candidates.
Overlap analysis between the RA-associated module and ferroptosis/cuproptosis gene sets highlighted three upregulated hub genes: FTH1 (ferritin heavy chain 1), SOD2 (superoxide dismutase 2), and CDKN2A (cyclin-dependent kinase inhibitor 2A). These genes exhibited high module membership within the RA-associated WGCNA module and significant differential expression in RA monocytes. PPI network analysis supported their status as hub nodes in the intersecting gene set.
Functional enrichment of the intersecting gene sets emphasized pathways relevant to metal-dependent regulated cell death and RA biology: oxidative stress response, iron and copper homeostasis, mitochondrial respiration and dysfunction, and cellular senescence. The combined signal suggests mechanistic links by which altered iron and copper handling and mitochondrial stress might modulate immune cell behavior and synovial pathology.
Immune cell infiltration deconvolution analyses associated the identified hub genes with specific immune patterns in RA: increased signatures of pro-inflammatory monocytes and macrophages and reduced regulatory T cell signals. These correlations align with the known roles of monocyte/macrophage populations and regulatory T cells in sustaining or restraining synovial inflammation, respectively.
Single-cell RNA-sequencing interrogation (dataset GSE296117) localized expression of the hub genes predominantly to synovial macrophages and fibroblasts in RA tissue. Both cell types are central effectors of joint damage and inflammation: macrophages as cytokine-producing innate responders and fibroblast-like synoviocytes (FLS) as locally aggressive, matrix-degrading cells. The single-cell data therefore provide tissue-context evidence supporting the potential relevance of FTH1, SOD2, and CDKN2A to joint pathology.
The integrative network analysis reported here identifies FTH1, SOD2, and CDKN2A as candidate genes at the intersection of ferroptosis and cuproptosis pathways in RA. The study links these genes to oxidative stress, disrupted metal ion homeostasis, mitochondrial dysfunction, specific immune infiltration patterns, and expression in synovial macrophages and fibroblasts. The authors propose these genes as candidate biomarkers and potential therapeutic targets, noting that all data and analysis scripts are publicly available (bulk RNA-seq accession GSE294225, single-cell accession GSE296117, and an accompanying GitHub repository).
Limitations reported in the source include reliance on transcriptomic datasets and bioinformatic inference; functional validation and mechanistic experiments were not reported in the article. The source provides no additional experimental validation data beyond the described computational analyses.
Overall, the study presents a systems-level argument that intersecting metal-dependent regulated cell death pathways — specifically ferroptosis and cuproptosis — merit further experimental and translational investigation in rheumatoid arthritis, with FTH1, SOD2, and CDKN2A prioritized for follow-up.