Background Juvenile idiopathic arthritis (JIA) is the one of the most common chronic childhood diseases with a prevalence of around 1/1000 . Over time JIA can result in persistent joint inflammation leading to chronic pain and stiffness, joint deformity and damage. Disease aetiology remains unknown. Investigation of disease pathology at the level of the synovial membrane (SM) is required if we want to begin to understand the disease at the molecular and biochemical level.
Objectives Proteomics enables us to better understand the disease process. Thus we undertook proteomic analysis of the SM from early disease-stage, treatment naïve JIA patients in order to identify protein expression profiles that could define JIA subgroups and help explain disease pathology.
Methods SM biopsies were acquired from 16 newly diagnosed, untreated JIA patients (8 polyarticular and 8 oligoarticular) under general anaesthetic. After a 2 year follow up none of patients in the oligoarticular group had extended and so this group of patients were termed persistent oligoarticular. Protein was extracted from SM in lysis buffer. Each SM sample was minimally labeled with either Cy5 or Cy3 fluorescent dyes and an internal pooled standard (Cy2) was included on each gel. Proteins were separated by 2-D Fluorescence Difference Gel Electrophoresis (2D DIGE). Gel images were analysed and proteins displaying a 2-fold or greater change in expression levels between oligoarticular and polyarticular patients were identified by mass spectrometry (MS) with expression further verified by Western blotting. Proteins were then localised within the SM by immunohistochemistry.
Results 2D DIGE analysis revealed an average of 460 spots per pH 4-7, 11cm gel which were matched across all 16 gels. ANOVA revealed 21 spots that were differentially expressed (2 fold or more change in expression levels) between polyarticular and oligoarticular patients (p≤0.05); 14 spots were expressed at 2-fold or higher in the oligoarticular group and 7 were expressed at 2-fold or higher in the polyarticular group. Principal component analysis (PCA) indicated that the synovial membrane proteome forms 2 distinct groups of patients in an unsupervised fashion. Hierarchical cluster analysis (HCA) was used to examine protein expression patterns and Pearson ranked correlation revealed two distinctive clusters of proteins.
Conclusions The data indicates that SM proteome profiles could be used to stratify patients based on disease subgroup. The ability to identify, quantify and localise these specific target proteins in our patient samples would not only provide a novel insight into JIA disease pathology but may also provide potential therapeutic targets which could in turn be used to guide diagnosis and treatment.
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Disclosure of Interest None Declared
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