Abstract

ObjectiveAutoantibodies to aminoacyl-tRNA synthetases (ARSs) are useful in the diagnosis of idiopathic inflammatory myopathy (IIM) with interstitial pneumonia (IP). We developed an enzyme-linked immunosorbent assay (ELISA) system using a mixture of recombinant ARS antigens and tested its utility in a multicenter study. Methods: We prepared six recombinant ARSs: GST-Jo-1, His-PL-12, His-EJ and GST-KS expressed in Escherichia coli, and His-PL-7 and His-OJ expressed in Hi-5 cells. After confirming their antigenic activity, with the exception of His-OJ, we developed our ELISA system in which the five recombinant ARSs (without His-OJ) were mixed. Efficiency was confirmed using the sera from 526 Japanese patients with connective tissue disease (CTD) (IIM n = 250, systemic lupus erythematosus n = 91, systemic sclerosis n = 70, rheumatoid arthritis n = 75, Sjögren’s syndrome n = 27 and other diseases n = 13), 168 with idiopathic interstitial pneumonia (IIP) and 30 healthy controls collected from eight institutes. IIPs were classified into two groups; idiopathic pulmonary fibrosis (IPF) (n = 38) and non-IPF (n = 130). Results were compared with those of RNA immunoprecipitation. Results: Sensitivity and specificity of the ELISA were 97.1% and 99.8%, respectively when compared with the RNA immunoprecipitation assay. Anti-ARS antibodies were detected in 30.8% of IIM, 2.5% of non-myositis CTD, and 10.7% of IIP (5.3% of IPF and 12.3% of non-IPF). Anti-ARS-positive non-IPF patients were younger and more frequently treated with glucocorticoids and/or immunosuppressants than anti-ARS-negative patients. Conclusion: A newly established ELISA detected anti-ARS antibodies as efficiently as RNA immunoprecipitation. This system will enable easier and wider use in the detection of anti-ARS antibodies in patients with IIM and IIP.

Highlights

  • A number of autoantibodies can be detected in sera from patients with idiopathic inflammatory myopathy (IIM), some of which are specific to IIM

  • Because we hypothesized that poor antigenic activity of recombinant PL-7 and OJ was due to a lack of posttranslational modification or proper structural folding, we prepared both fusion proteins expressed in eukaryotic Hi-5 cells using the baculovirus system

  • We tested a variety of antigen mixtures to estimate the most appropriate ratio and concentration to use, and we found that anti-aminoacyl-tRNA synthetases (ARSs)-positive sera showed reactivity with all five different ARSs with the highest sensitivity and specificity occurring at antigen concentrations of 0.63, 1.25, 1.25, 0.63, and 2.5 mg/mL (6.25 mg/mL in total) for histidyl, threonyl, alanyl, glycyl, and asparaginyl-tRNA synthetases, respectively

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Summary

Introduction

A number of autoantibodies can be detected in sera from patients with idiopathic inflammatory myopathy (IIM), some of which are specific to IIM (known as myositis-specific autoantibodies: MSAs). Detection of these autoantibodies is closely associated with IIM clinical manifestations [1,2]. Patients with anti-ARSs show a spectrum of common clinical manifestations known as anti-synthetase syndrome (ASS), including myositis, interstitial pneumonia (IP), nonerosive arthritis, fever, Raynaud’s phenomenon, and mechanic’s hands. Anti-ARS antibodies are useful in diagnosing IIM and in predicting late-onset myopathy in IPproceeding patients and the clinical course of IP in myositis

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Conclusion

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