INCREASING CONCENTRATIONS OF PROTEOLYTIC DEBRIS OF GLYCATED AND OXIDISED PROTEINS IN PATIENTS WITH INCREASING SEVERITY OF OSTEOARTHRITIS

Abstract

Background: Osteoarthritis (OA) is a progressive degenerative condition that causes pain and impairs the mobility of more than 10% of the UK population. Over 50,000 total knee replacements (TKR) are carried out each year for patients with the most severe symptoms. The aim of this study was to assess if markers of protein damage in the synovial fluid and plasma of patients with OA increase with severity of symptoms. These markers may then be of use in assessing disease presence and progression to assist with subsequent management. Proteins damaged by glycation (modified by sugars) and oxidation undergo cellular proteolysis. The proteolytic debris thereby formed - called glycation and oxidation free adducts (glycated and oxidised amino acids) - is released into the synovium and plasma for urinary excretion. In this study the concentrations of the glycation free adducts, Nε-carboxymethyl-lysine (CML) and methylglyoxal-derived hydroimidazolone (MG-H1), and the oxidation free adduct methionine sulfoxide (MetSO) were measured in the synovial fluid and plasma of patients with severe OA (sOA) and early-stage OA (eOA).

Methods: Patients were recruited from those attending the Rheumatology clinics at Ipswich Hospital, Ipswich and the Orthopaedic clinics at University Hospital, Coventry. Volunteer subjects were recruited to the normal healthy control group. The age (years; mean ± SD) for patient and control subject groups was: controls 45 ± 6 (n = 8), sOA 70 ± 12 (n = 8), eOA 50 ± 14 (n = 6). Patients found to have eOA changes (Outerbridge grade I/II) during routine arthroscopy were recruited to the eOA group. Synovial fluid and venous blood samples were taken with informed consent. All synovial fluid samples were taken from the knee joint. The concentrations of glycation and oxidation free adducts were assayed by stable isotopic dilution analysis liquid chromatography with tandem mass spectrometric detection (LC-MS/MS) in ultrafiltrate of synovial fluid and plasma. Significance of difference between study groups was assessed by ANOVA and Student’s t-test.

Results: The concentrations (nM; mean ± SD) of MetSO, CML and MG-H1 in synovial fluid were all markedly increased in OA patients with more severe disease. MetSO free adduct: eOA 459 ± 107 and sOA 2889 ± 1064 (p< 0.001). CML free adduct: eOA 77 ± 24 and 224 ± 51 (p< 0.001). MG-H1 free adduct: eOA 387 ± 182 and sOA 674 ±199 (p< 0.05). Analysis of plasma of these patients also showed increases in the concentrations of corresponding glycation and oxidation free adducts compared to those of normal healthy controls.

Discussion: The concentration of glycation and oxidation free adducts increased with severity of symptoms in the synovial fluid of patients with OA. This probably occurs by down regulation of protective gene expression in OA. Measurement of plasma protein glycation and oxidation free adducts may be useful in assessing progression and severity of OA. In the future, these markers may guide non-operative management and facilitate earlier joint-preserving surgery.