title: Mesenchymal stem cell isolation and characterization from human spinal ligaments.
abstract: Mesenchymal stem cells (MSCs) have a fibroblast-like morphology, multilineage potential, long-term viability and capacity for self-renewal. While several articles describe isolating MSCs from various human tissues, there are no reports of isolating MSCs from human spinal ligaments, and their localization in situ. If MSCs are found in human spinal ligaments, they could be used to investigate hypertrophy or ossification of spinal ligaments. To isolate and characterize MSCs from human spinal ligaments, spinal ligaments were harvested aseptically from eight patients during surgery for lumbar spinal canal stenosis and ossification of the posterior longitudinal ligament. After collagenase digestion, nucleated cells were seeded at an appropriate density to avoid colony-to-colony contact. Cells were cultured in osteogenic, adipogenic or chondrogenic media to evaluate their multilineage differentiation potential. Immunophenotypic analysis of cell surface markers was performed by flow cytometry. Spinal ligaments were processed for immunostaining using MSC-related antibodies. Cells from human spinal ligaments could be extensively expanded with limited senescence. They were able to differentiate into osteogenic, adipogenic or chondrogenic cells. Flow cytometry revealed that their phenotypic characteristics met the minimum criteria of MSCs. Immunohistochemistry revealed the localization of CD90-positive cells in the collagenous matrix of the ligament, and in adjacent small blood vessels. We isolated and expanded MSCs from human spinal ligaments and demonstrated localization of MSCs in spinal ligaments. These cells may play an indispensable role in elucidating the pathogenesis of numerous spinal diseases.
authors.affiliation: Department of Orthopaedic Surgery, Hirosaki University Graduate School of Medicine, 5 Zaifu-cho, Hirosaki, Aomori 036-8562, Japan.
authors.forename: Toru
authors.initials: T
authors.lastname: Asari
authors.forename: Ken-Ichi
authors.initials: K
authors.lastname: Furukawa
authors.forename: Sunao
authors.initials: S
authors.lastname: Tanaka
authors.forename: Hitoshi
authors.initials: H
authors.lastname: Kudo
authors.forename: Hiroki
authors.initials: H
authors.lastname: Mizukami
authors.forename: Atsushi
authors.initials: A
authors.lastname: Ono
authors.forename: Takuya
authors.initials: T
authors.lastname: Numasawa
authors.forename: Gentaro
authors.initials: G
authors.lastname: Kumagai
authors.forename: Shigeru
authors.initials: S
authors.lastname: Motomura
authors.forename: Soroku
authors.initials: S
authors.lastname: Yagihashi
authors.forename: Satoshi
authors.initials: S
authors.lastname: Toh
mesh_terms: D017667:Adipocytes / Q000166:cytology; D050156:Adipogenesis; D002452:Cell Count; D018929:Cell Culture Techniques; D002454:Cell Differentiation; D002469:Cell Separation; D016922:Cellular Senescence; D019902:Chondrocytes / Q000166:cytology; D006801:Humans; D007150:Immunohistochemistry; D008022:Ligaments / Q000166:cytology*; D059630:Mesenchymal Stem Cells / Q000166:cytology* / Q000502:physiology; D010006:Osteoblasts / Q000166:cytology; D013131:Spine