Elucidating the Molecular Composition of Cartilage by Proteomics

Abstract

Articular cartilage consists of chondrocytes and two major components, a collagen-rich framework and highly abundant proteoglycans. Most prior studies defining the zonal distribution of cartilage have extracted proteins with guanidine-HCl. However, an unextracted collagen-rich residual is left after extraction. In addition, the high abundance of anionic polysaccharide molecules extracted from cartilage adversely affects the chromatographic separation. In this study, we established a method for removing chondrocytes from cartilage sections with minimal extracellular matrix protein loss. The addition of surfactant to guanidine-HCl extraction buffer improved protein solubility. Ultrafiltration removed interference from polysaccharides and salts. Almost four times more collagen peptides were extracted by the in situ trypsin digestion method. However, as expected, proteoglycans were more abundant within the guanidine-HCl extraction. These different methods were used to extract cartilage sections from different cartilage layers (superficial, intermediate and deep), joint types (knee and hip), and disease states (healthy and osteoarthritic) and the extractions were evaluated by quantitative and qualitative proteomic analyses. The results of this study led to the identifications of the potential biomarkers of OA, OA progression, and the joint specific biomarkers.

Chondrocyte secretome: a source of novel insights and exploratory biomarkers of osteoarthritis

“…MMP-1 and 3, and TIMP-1, are particularly abundant. TIMP-3 appears to be more abundant in normal hip cartilage than knee, but MMP-1 is contrarily abundant in normal knee cartilage [37]. According to these studies, MMP-13…”

“…Furthermore, the C-terminal processed domain of PCPE (CT-PCPE) may have metalloproteinase inhibitory activity. PCOLCE1 is increased during chondrocyte differentiation of MSCs [26] and in OA cartilage [37], but is decreased with IL-1β [14,15] and with TGF-β1 [24].…”

“…HTRA1 is preferentially localized in the deep layer of cartilage [34] and increased during chondrocyte differentiation of MSCs [26], and by TGF-β1 stimulation [24]. HTRA1 is reduced 4-fold in OA vs normal chondrocytes [35], with no difference detected between OA and normal decellularized cartilage [37].…”

“…In this part of the review, we focused on proteins identified either directly in the secretome of cartilage explants [12][13][14][15][16][17][18][19][20] or chondrocytes cultures [18,[21][22][23][24][25][26][27][28][29][30][31]), both of which are listed in Table 1. We further complete this list with proteins recently identified by proteomic analysis performed directly on fresh chondrocytes or cartilage tissue [32][33][34][35][36] (sometimes de-cellularized [37]) whereby different locations in the joint were compared, or healthy joint tissues were compared with 6 OA tissues. The characteristics of all these studies are summarized in Table 2.…”

“…This protein is more abundant in knee compared to hip cartilage [37] and in OA compared to healthy cartilage [27] and is considered to be an early OA biomarker in the Stenberg' proteomic study, which compared early and severe OA secretomes [25].…”