Session Title: Genetics and Genomics of Rheumatic Disease II
Session Type: Abstract Submissions (ACR)
Background/Purpose: Osteoarthritis (OA) is characterized by the progressive loss of cartilage structural extracellular matrix (ECM) components, mainly collagenous and non-collagenous proteins. The release of these proteins from the tissue can vary according to the stage of the disease. Characterization of molecular differences between cartilage subtypes will provide a background for better understanding OA onset and progression. The aim of this study was to perform a quantitative proteomics approach to identify and quantify those proteins released from normal (N) and OA human articular cartilages.
Methods: Tissue explants were obtained from the dissection of 4 N and 4 OA cartilages, both from femoral heads and tibial condyles. Among the OA samples, we differentiated the wounded zones (WZ) from those corresponding to the area adjacent to the lesion, or unwounded zones (UZ). The study was approved by the local ethical committee. Cartilage shavings from each donor were cut into 6 mm discs and five discs/donor were placed into 96 wells plates and incubated during 6 days (37 °C/5% CO2). The conditioned media from each condition were collected and their proteins were digested with trypsin. Each peptide mixture was labelled with different isobaric tags using the iTRAQ reagents (ABSciex). Then, labelled peptides of the different conditions were mixed, desalted and separated by liquid chromatography (LC). The resulting fractions were grouped and resolved by reversed-phase nano-LC coupled to mass spectrometry (MS). The identification and relative quantification of the proteins was carried out with Protein Pilot 3.0 software.
Results: A mean of 206 cartilage secreted proteins was identified. Measurement of the different iTRAQ tags intensities allowed the relative quantification of almost all of them. Globally, we found 35 secreted proteins with statistically significant differences in abundance (p≤0.05) between the different OA zones (WZ and UZ) when compared with N samples: 26 were increased and 9 were decreased. We classified them in 3 sets of proteins: a group of 5 proteins (CILP, CILP2, CHI3L1, SPP1, PLA2G2A) were increased specifically in UZ samples (early OA biomarkers), a group of 6 proteins (APOA1, HSPG2, CRTAC1, COL12A1, COL15A1, PRG4) were increased only in WZ samples (late OA biomarkers) and finally a group of 15 proteins were modified in both OA zones. Although some of these proteins, like CHI3L1 and PRG4, have a previously reported putative biomarker value for OA, most of them are novel candidates of the disease onset (the first group) and progression (the second and third groups). Interesting when we compared these results with those obtained from cartilage tissue extracts analysis we found that most of the proteins increased in OA cartilage secretomes (like BGN, HAPLN1, APCS, TNC, TGFBI) are decreased in cartilage proteomes and viceversa.
Conclusion: We have identified a characteristic profile of proteins released from N and OA cartilages. We describe a panel of cartilage ECM proteins with potential biomarker value. This panel will be further explored in biological fluids (synovial fluid and serum) for the development of early diagnosis and/or anti-OA therapy monitoring strategies.
F. J. Blanco,
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ACR Meeting Abstracts - https://acrabstracts.org/abstract/proteomics-analysis-of-cartilage-secretome-a-powerful-tool-for-the-discovery-of-oa-biomarkers/