title: Efficient expansion of mouse primary tenocytes using a novel collagen gel culture method.
abstract: Development of regenerative therapies for damaged tendons remains a great challenge, largely because of lack of information regarding the mechanisms responsible for differentiation of tenocytes. Mouse tenocytes have not been fully characterized owing to the absence of efficient and reproducible methods for their in vitro expansion without losing phenotypic features. The objective of the study was to establish an improved and reliable method for stable primary culture of mouse tenocytes by using collagen gel. Achilles and tail tendon tissues were harvested and embedded in collagen gel. After 10 days of continuous culture, the gel was digested and cells were passaged on tissue culture-treated plastic dishes. Mouse tenocytes cultured in collagen gel exhibited significantly shorter doubling time and higher numbers of proliferation when maintained on the plastic dishes compared with those cultured without using gel. Transmission electron microscopic analyses showed that cultured tenocytes retained some morphological features of tenocytes in tendon tissues, such as cell-cell junctional complex formation, well-developed rough endoplasmic reticulum, and mitochondria in their cytoplasm. mRNA expression of tenocyte markers (tenomodulin, type I collagen, periostin, and scleraxis) was higher in cells cultured in collagen gel than in those cultured in the absence of gel. Our results show that tenocytes cultured using the collagen gel method express typical lineage markers and exhibit improved growth characteristics, thus providing a stable platform for studying molecular mechanisms that control their differentiation.
authors: Department of Pharmacology, School of Dental Medicine, Tsurumi University, 2-1-3 Tsurumi, Tsurumi-ku, Yokohama, 230-8501, Japan, shimada-a@tsurumi-u.ac.jp.
authors: Akemi
authors: A
authors: Shimada
authors: Satoshi
authors: S
authors: Wada
authors: Kouji
authors: K
authors: Inoue
authors: Hisashi
authors: H
authors: Ideno
authors: Taichi
authors: T
authors: Kamiunten
authors: Koichiro
authors: K
authors: Komatsu
authors: Akira
authors: A
authors: Kudo
authors: Yoshiki
authors: Y
authors: Nakamura
authors: Tetsuji
authors: T
authors: Sato
authors: Kazuhisa
authors: K
authors: Nakashima
authors: Akira
authors: A
authors: Nifuji
mesh_terms: D000125:Achilles Tendon / Q000166:cytology*; D000818:Animals; D051792:Basic Helix-Loop-Helix Proteins / Q000096:biosynthesis; D015815:Cell Adhesion Molecules / Q000096:biosynthesis; D049109:Cell Proliferation; D064987:Cell- and Tissue-Based Therapy / Q000379:methods; D002478:Cells, Cultured; D024042:Collagen Type I / Q000096:biosynthesis; D018870:Endoplasmic Reticulum, Rough / Q000502:physiology; D005782:Gels / Q000494:pharmacology*; D008565:Membrane Proteins / Q000096:biosynthesis; D051379:Mice; D008810:Mice, Inbred C57BL; D046529:Microscopy, Electron, Transmission; D008928:Mitochondria; D061251:Primary Cell Culture / Q000379:methods*; D013708:Tendon Injuries / Q000628:therapy; D019108:Tight Junctions / Q000502:physiology
chemical_list: D051792:Basic Helix-Loop-Helix Proteins; D015815:Cell Adhesion Molecules; D024042:Collagen Type I; D005782:Gels; D008565:Membrane Proteins; C082838:Postn protein, mouse; C503815:Scx protein, mouse; C486026:Tnmd protein, mouse