High-throughput quantitation of serological dimethylarginines by LC/MS/MS: Potential cardiovascular biomarkers for rheumatoid arthritis

Rheumatoid arthritis (RA) is a chronic autoimmune disorder characterized by systemic inflammation of the joints and extra-articular tissues. The incidence of cardiovascular disease (CVD) remains the main cause of morbidity and mortality in patients with RA. Despite the development of new therapeutics targeting the articular mani-festations, the relief of the cardiovascular burden is still an unmet medical need during the management of RA. So, the early prognosis of RA-associated CVD plays a crucial role in improving the clinical outcomes of RA pa-tients. Recently, circulating dimethylarginines have gained attention as potential biomarkers for CVDs. Here, we present the development and validation of a high-throughput liquid chromatography-tandem mass spectrometric (LC/MS/MS) method for simultaneous quantification of creatinine, arginine, and dimethylarginines in human serum within 2 mins by isotope dilution mass spectrometry. This method employed a protein precipitation method for rapid sample preparation, trichloroacetic acid (TCA)-based ion pairing chromatography for fast analyte separation, and multiple reaction monitoring (MRM) with stable isotope-labeled internal standards (ISs) for simultaneous quantitation. To assure the quality, our method was validated against the FDA guidelines for lower limit of quantitation (0.2 mu M), linearity (square of coefficient correlation>0.99), precision (intra-&inter-assay imprecision < 10 %), accuracy (intra-&inter-assay inaccuracy < 10 %), sample preparation recovery (recovery >= 90 %), stability (instability < 10 %), matrix effect (signal suppression < 55 %), and carryover ( < 0.01 %). Afterward, we applied the validated method to a retrospective cross-sectional study. We aimed to evaluate the utility of serological dimethylarginines as potential cardiovascular biomarkers in the development of RA-associated CVD. Our results revealed that the serological ratio of asymmetric dimethylarginine (ADMA) and symmetric dimethylarginine (SDMA), an indicator of physiological arginine methylation status, was significantly elevated in patients with RA. This finding might provide value in detecting CVD to improve clinical outcomes in RA management.

Automated summary

Rheumatoid arthritis (RA) is an autoimmune disorder characterized by joint and tissue inflammation. Cardiovascular disease (CVD) is the main cause of morbidity and mortality in RA patients. The development of new therapeutics for joint symptoms has not adequately addressed the cardiovascular burden in RA management. Recent attention has focused on circulating dimethylarginines as potential biomarkers for CVD. The study presented a high-throughput method for simultaneous quantification of creatinine, arginine, and dimethylarginines in human serum. The results revealed that the serological ratio of asymmetric dimethylarginine (ADMA) and symmetric dimethylarginine (SDMA) was significantly elevated in RA patients, suggesting its potential value in detecting CVD and improving clinical outcomes in RA management.