Background/Purpose: Non-Esterified Fatty Acids (NEFA) are important lipid mediators which can play a role in a number of biological functions other than energy supply. NEFA are known to regulate gene expression in monocytes and macrophages, as well as to modulate of CD4⁺ T-cell function. Thus, we hypothesized that altered NEFA levels may underlie rheumatoid arthritis (RA) pathogenesis.

Methods: NEFA serum levels (palmitic, stearic, palmitoleic, oleic, linoleic, g-linoleic, arachidonic –AA–, linolenic, eicosapentaenoic –EPA– and docosahexaenoic –DHA–) were quantified by LC-MS/MS after methyl-tert–butylether (MTBE)-extraction in 124 RA patients (all fulfilling 2010 ACR/EULAR RA criteria, 61.2% RF+, 59.6% ACPA+) and 56 healthy controls (HC). Moreover, 13 prospectively-followed RA patients undergoing TNFa-blockade were recruited. CD4⁺ T-cell phenotype was studied by flow cytometry. TNFa, IL-8, VEGF, GM-CSF, IFNg, IL-17, MCP-1, IP-10, leptin and resistin serum levels were quantified by immunoassays. The effect of NEFA on IFNg production by PBMC was evaluated in vitro.

Results: Lower levels of palmitic (p<0.0001), palmitoleic (p=0.002), oleic (p=0.010), arachidonic (p=0.027), EPA (p<0.0001) and DHA (p<0.0001) were found in RA, some NEFA being altered at onset. No differences in total NEFA level were observed (p=0.157), thus pointing to an altered NEFA profile in RA. Cluster analysis identified a FFA profile (NEFA^low) characterized by increased stearic and decreased EPA and DHA levels to be overrepresented in RA patients compared to HC (p=0.002) (Figure 1). NEFA^low profile was associated with clinical features (RF, shared epitope and erosions), increased IFNg expression in CD4⁺ T-cells (p=0.002) and a Th1-enhanced serum milieu (IFNg, MCP-1 and IP-10, all p<0.005). In vitro assays demonstrated that imbalanced NEFA could underlie IFNg production by CD4⁺ T-cells. Finally, changes on NEFA levels were associated with clinical response upon TNFa-blockade, decreasing EPA and DHA being related to a poor clinical outcome.

Conclusion: An altered NEFA profile can be found in RA patients associated with clinical characteristics of aggressive disease and Th1-enhanced response. Impaired NEFA levels may underlie a poor clinical outcome upon TNFa-blockade. A complex NEFA profile was observed, specific NEFA being altered independently of their chemical properties. These results support the relevance of lipidomic studies in RA and provide a rationale for new therapeutic targets.