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Abstract

Objective

This study was undertaken to identify new biomarkers of osteoarthritis (OA) by proteomics analysis and to develop specific immunoassays to detect and quantify them.

Methods

Proteomics analysis was performed in urine samples from 10 women (mean ± SD age 76.0 ± 5.0 years) undergoing knee replacement surgery due to severe OA and 5 healthy women (mean ± SD age 25.6 ± 2.6 years). Protein content was analyzed by 2-dimensional differential gel electrophoresis. Protein spots that exhibited an OA:control abundance ratio of ≥1.5 were identified by mass spectrometry. Specific enzyme-linked immunosorbent assays were developed and validated in serum obtained from 236 healthy subjects ages 20–64 years and from 76 patients with severe radiologic knee OA (mean ± SD age 68.8 ± 11.9 years). Immunohistochemical analysis was performed on articular cartilage from tibial plateaus.

Results

Thirteen proteins within spots that were significantly modified between groups were identified. Two peptides of fibulin 3, named Fib3-1 and Fib3-2, were of particular interest. Two antisera directed against these peptides were used to develop immunoassays. Compared with age-matched healthy subjects, median levels of serum Fib3-1 and Fib3-2 were elevated in OA patients (54.6 pM versus 85.1 pM [P < 0.0001] and 144.4 pM versus 191.4 pM [P < 0.0001], respectively). Using area under the receiver operating characteristic curve analysis, we demonstrated that Fib3-1 and Fib3-2 levels discriminate between OA and normal populations. Immunostaining revealed the presence of Fib3-1 and Fib3-2 in chondrocytes and in the extracellular matrix of the superficial layer of the fibrillated cartilage.

Conclusion

Our findings indicate that Fib3-1 and Fib3-2 are potential biochemical markers for the diagnosis of OA.