To comprehensively investigate the global lactylation proteomics profile in patients with rheumatoid arthritis (RA) and to meticulously validate the specifically sorted candidate lactylated modification sites to understand their potential pathogenic roles. Protein lactylation is a recently identified post-translational modification that has been rarely investigated in the context of rheumatoid arthritis, despite its known critical involvement in cellular processes. Therefore, this study aimed to thoroughly explore how these unique lactylation proteomics alterations could potentially contribute to the complex development and ongoing progression of the disease. 

Synovial tissue samples strategically collected from a well-defined cohort of ten rheumatoid arthritis patients and six osteoarthritis (OA) control patients were systematically subjected to advanced lactylation proteomics analysis utilizing high-throughput affinity enrichment paired with high-resolution liquid chromatography-tandem mass spectrometry (LC-MS/MS). Extensive bioinformatics evaluations, specifically Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG), and COG/KOG enrichment analyses, were rigorously applied to determine the functional pathways of the dysregulated proteins. Furthermore, to verify the findings, four distinct candidate lactylated modification sites were precisely validated using targeted immunoprecipitation techniques, and additional in vitro functional assessments were conducted on transfected human rheumatoid arthritis fibroblast-like synoviocytes. 

The comprehensive analysis successfully identified a robust total of 566 distinct lactylated modification sites and 250 corresponding lactylated proteins specifically within the rheumatoid arthritis patient group. When directly compared to the osteoarthritis patient cohort, researchers discovered 24 significantly upregulated and 2 downregulated lactylated modification sites, alongside 18 prominently upregulated and 1 downregulated lactylated modification proteins. These notably dysregulated lactylated proteins were heavily enriched in critical biological processes such as the positive regulation of plasma membrane repair and neutrophil extracellular trap formation. Crucially, targeted immunoprecipitation conclusively confirmed that the relative abundance of FTH1-K69la and PKM2-K166la was substantially greater in rheumatoid arthritis patients, with FTH1-K69la showing a direct positive correlation with the erythrocyte sedimentation rate. 

This pioneering study successfully describes and mapping a general, comprehensive landscape of lactylation proteomics specifically within the complex environment of the rheumatoid arthritis synovium. By rigorously screening out and thoroughly validating specific potential lactylated modification proteins, most notably the significantly upregulated FTH1-K69la and PKM2-K166la sites, the research delivers essential preliminary evidence highlighting the functional importance of protein lactylation. Ultimately, these crucial discoveries provide a strong foundational basis for future, more targeted therapeutic investigations aimed at addressing and potentially mitigating the underlying pathogenic mechanisms driving rheumatoid arthritis development and progression.