title: Role of apoptosis inhibition in various chondrocyte culture systems.
abstract: Apoptosis may limit the utility of cell-based therapies for articular cartilage disorders. We tested the hypothesis that chondrocyte apoptosis can be reduced by optimizing the conditions employed to expand chondrocyte numbers in culture. Chondrocyte apoptosis was examined in monolayer and suspension culture, in the presence of fetal bovine serum (FBS) or autologous serum, and for culture periods of 2 or 4 days. The effect of these variables was assessed by measuring cell viability, Annexin V labeling and mitochondrial membrane potential. After 2 days of culture, the greatest increase in viable cell number (3.7-fold) occurred in monolayer cultures with autologous serum. After 4 days of culture, the greatest increase in cell number (9.0-fold) occurred in monolayer cultures supplemented with FBS. By Annexin V staining, the proportion of cells undergoing apoptosis after 2 days was not affected by the type of serum used or by culture in monolayer versus suspension. After 4 days of culture, the proportion of apoptotic cells was significantly reduced (35% to 13%, p<0.02) in suspension cultures with autologous serum. Apoptosis assessed by loss of mitochondrial membrane potential was decreased in the presence of autologous serum. These data suggest that suspension culture with autologous serum is useful in simultaneously maintaining cell proliferation and minimizing apoptosis in cultured human articular chondrocytes.
authors.affiliation: Department of Orthopaedic Surgery, Kochi Medical School, Oko-cho, Nankoku, Kochi 783-8505, Japan. takahast@kochi-ms.ac.jp
authors.forename: Toshiaki
authors.initials: T
authors.lastname: Takahashi
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Yamamoto
authors.forename: Yasuhiro
authors.initials: Y
authors.lastname: Ogawa
authors.forename: Shoji
authors.initials: S
authors.lastname: Yoshida
authors.forename: Sayo
authors.initials: S
authors.lastname: Kataoka
authors.forename: Manas
authors.initials: M
authors.lastname: Majumdar
authors.forename: Elisabeth
authors.initials: E
authors.lastname: Morris
authors.forename: Stephen B
authors.initials: SB
authors.lastname: Tripple
mesh_terms: D017304:Annexin A5 / Q000378:metabolism; D017209:Apoptosis / Q000502:physiology*; D002452:Cell Count; D018929:Cell Culture Techniques / Q000379:methods; D002470:Cell Survival; D002478:Cells, Cultured; D019902:Chondrocytes / Q000378:metabolism / Q000502:physiology*; D005434:Flow Cytometry; D016650:Fluorescein-5-isothiocyanate / Q000378:metabolism; D006801:Humans; D008564:Membrane Potentials; D008928:Mitochondria / Q000378:metabolism / Q000502:physiology; D013997:Time Factors
chemical_list: D017304:Annexin A5; D016650:Fluorescein-5-isothiocyanate