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Molecular & Cellular Proteomics 5:1984-1995, 2006.
© 2006 by The American Society for Biochemistry and Molecular Biology, Inc.

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Methodologies

Establishment of a Reliable Method for Direct Proteome Characterization of Human Articular Cartilage^*

Jean-Baptiste Vincourt^,, Frédéric Lionneton, Gueorgui Kratassiouk^¶, François Guillemin^¶, Patrick Netter, Didier Mainard and Jacques Magdalou

From the Laboratoire de Physiopathologie et Pharmacologie Articulaires, Faculté de Médecine, Unité Mixte de Recherche (UMR) 7561 CNRS-Université Henry Poincaré (UHP), 9 Avenue de la Forêt de Haye, BP 184, 54505 Vandoeuvre-lès-Nancy Cedex, France and ^¶ Centre Alexis Vautrin, Centre de Recherche en Automatisme de Nancy UMR 7039, CNRS-Institut National Polytechnique Lorrain-UHP, 54500 Vandoeuvre-lès-Nancy, France

Articular cartilage consists mainly of extracellular matrix, mostly made of collagens and proteoglycans. These macromolecules have so far impaired the detailed two-dimensional electrophoresis-based proteomic analysis of articular cartilage. Here we describe a method for selective protein extraction from cartilage, which excludes proteoglycans and collagen species, thus allowing direct profiling of the protein content of cartilage by two-dimensional electrophoresis. Consistent electrophoretic patterns of more than 600 protein states were reproducibly obtained after silver staining from 500 mg of human articular cartilage from joints with diverse pathologies. The extraction yield increased when the method was applied to a chondrosarcoma sample, consistent with selective extraction of cellular components. Nearly 200 of the most intensely stained protein spots were analyzed by MALDI-TOF mass spectrometry after trypsin digestion. They represented 127 different proteins with diverse functions. Our method provides a rapid, efficient, and pertinent alternative to previously proposed approaches for proteomic characterization of cartilage phenotypes. It will be useful for detecting protein expression patterns that relate pathophysiological processes of cartilaginous tissues such as osteoarthritis and chondrosarcoma.

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