Elucidating the Molecular Composition of Cartilage by Proteomics

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Elucidating the Molecular Composition of Cartilage by Proteomics. / Hsueh, Ming Feng; Khabut, Areej; Kjellström, Sven; Önnerfjord, Patrik; Kraus, Virginia Byers.

In: Journal of Proteome Research, Vol. 15, No. 2, 05.02.2016, p. 374-388.

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Hsueh, Ming Feng ; Khabut, Areej ; Kjellström, Sven ; Önnerfjord, Patrik ; Kraus, Virginia Byers. / Elucidating the Molecular Composition of Cartilage by Proteomics. In: Journal of Proteome Research. 2016 ; Vol. 15, No. 2. pp. 374-388.

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TY - JOUR

T1 - Elucidating the Molecular Composition of Cartilage by Proteomics

AU - Hsueh, Ming Feng

AU - Khabut, Areej

AU - Kjellström, Sven

AU - Önnerfjord, Patrik

AU - Kraus, Virginia Byers

PY - 2016/2/5

Y1 - 2016/2/5

N2 - 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 osteoarthritis (OA), OA progression, and the joint specific biomarkers.

AB - 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 osteoarthritis (OA), OA progression, and the joint specific biomarkers.

KW - cartilage

KW - extracellular matrix proteins

KW - guanidine-HCl extraction

KW - in situ trypsin digestion

KW - multiple reaction monitoring

KW - proteomics

UR - http://www.scopus.com/inward/record.url?scp=84957628373&partnerID=8YFLogxK

U2 - 10.1021/acs.jproteome.5b00946

DO - 10.1021/acs.jproteome.5b00946

M3 - Article

VL - 15

SP - 374

EP - 388

JO - Journal of Proteome Research

T2 - Journal of Proteome Research

JF - Journal of Proteome Research

SN - 1535-3893

IS - 2

ER -