title: Proteomic Analysis of Human Tendon and Ligament: Solubilization and Analysis of Insoluble Extracellular Matrix in Connective Tissues.
abstract: Connective tissues such as tendon, ligament and cartilage are mostly composed of extracellular matrix (ECM). These tissues are insoluble, mainly due to the highly cross-linked ECM proteins such as collagens. Difficulties obtaining suitable samples for mass spectrometric analysis render the application of modern proteomic technologies difficult. Complete solubilization of them would not only elucidate protein composition of normal tissues but also reveal pathophysiology of pathological tissues. Here we report complete solubilization of human Achilles tendon and yellow ligament, which is achieved by chemical digestion combined with successive protease treatment including elastase. The digestion mixture was subjected to liquid chromatography-mass spectrometry. The low specificity of elastase was overcome by accurate mass analysis achieved using FT-ICR-MS. In addition to the detailed proteome of both tissues, we also quantitatively determine the major protein composition of samples, by measuring peak area of some characteristic peptides detected in tissue samples and in purified proteins. As a result, differences between human Achilles tendon and yellow ligament were elucidated at molecular level.
authors.affiliation: Department of Orthopedics, Institute of Biomedical Sciences, Tokushima University Graduate School , 3-18-15 Kuramoto-cho, Tokushima 770-8503, Japan.
authors.forename: Nori
authors.initials: N
authors.lastname: Sato
authors.affiliation: Division of Disease Proteomics, Institute for Enzyme Research, Tokushima University , 3-18-15 Kuramoto-cho, Tokushima 770-8503, Japan.
authors.forename: Takako
authors.initials: T
authors.lastname: Taniguchi
authors.affiliation: Department of Orthopedics, Institute of Biomedical Sciences, Tokushima University Graduate School , 3-18-15 Kuramoto-cho, Tokushima 770-8503, Japan.
authors.forename: Yuichiro
authors.initials: Y
authors.lastname: Goda
authors.affiliation: Department of Orthopedics, Institute of Biomedical Sciences, Tokushima University Graduate School , 3-18-15 Kuramoto-cho, Tokushima 770-8503, Japan.
authors.forename: Hirofumi
authors.initials: H
authors.lastname: Kosaka
authors.affiliation: Department of Orthopedics, Institute of Biomedical Sciences, Tokushima University Graduate School , 3-18-15 Kuramoto-cho, Tokushima 770-8503, Japan.
authors.forename: Kosaku
authors.initials: K
authors.lastname: Higashino
authors.affiliation: Department of Orthopedics, Institute of Biomedical Sciences, Tokushima University Graduate School , 3-18-15 Kuramoto-cho, Tokushima 770-8503, Japan.
authors.forename: Toshinori
authors.initials: T
authors.lastname: Sakai
authors.affiliation: Department of Orthopedics, Institute of Biomedical Sciences, Tokushima University Graduate School , 3-18-15 Kuramoto-cho, Tokushima 770-8503, Japan.
authors.forename: Shinsuke
authors.initials: S
authors.lastname: Katoh
authors.affiliation: Department of Orthopedics, Institute of Biomedical Sciences, Tokushima University Graduate School , 3-18-15 Kuramoto-cho, Tokushima 770-8503, Japan.
authors.forename: Natsuo
authors.initials: N
authors.lastname: Yasui
authors.affiliation: Department of Orthopedics, Institute of Biomedical Sciences, Tokushima University Graduate School , 3-18-15 Kuramoto-cho, Tokushima 770-8503, Japan.
authors.forename: Koichi
authors.initials: K
authors.lastname: Sairyo
authors.affiliation: Division of Disease Proteomics, Institute for Enzyme Research, Tokushima University , 3-18-15 Kuramoto-cho, Tokushima 770-8503, Japan.
authors.forename: Hisaaki
authors.initials: H
authors.lastname: Taniguchi
mesh_terms: D000125:Achilles Tendon / Q000737:chemistry*; D002853:Chromatography, Liquid; D003238:Connective Tissue / Q000737:chemistry*; D005109:Extracellular Matrix / Q000737:chemistry*; D006801:Humans; D008022:Ligaments / Q000737:chemistry*; D013058:Mass Spectrometry; D010447:Peptide Hydrolases / Q000378:metabolism; D020543:Proteome / Q000032:analysis*; D040901:Proteomics / Q000379:methods; D012995:Solubility
chemical_list: D020543:Proteome; D010447:Peptide Hydrolases
keywords: Achilles tendon; collagen; elastin; proteome; yellow ligament