OBJECTIVE: The effect of in vitro expansion of human adipose-derived stem cells (ASCs) on stem cell properties is controversial. We examined serial subcultivation with expansion on the ability of ASCs to grow and differentiate into osteoblastic lineages. DESIGN: Flow cytometric analysis, growth kinetics, cell population doubling time, light microscopy and confocal analysis, and osteogenesis induction were performed to assess growth and osteogenic potential of subcultivated ASCs at passages 2 (P2), P4 and P6. RESULTS: Flow cytometric analysis revealed that ASCs at P2 express classical mesenchymal stem cell markers including CD44, CD73, and CD105, but not CD14, CD19, CD34, CD45, or HLA-DR. Calcium deposition and alkaline phosphatase activity were the highest at P2 but completely abrogated at P4. Increased passage number impaired cell growth; P2 cultures exhibited exponential growth, while cells at P4 and P6 showed near linear growth with cell population doubling times increased from 3.2 at P2 to 4.8 d at P6. Morphologically, cells in various subcultivation stages showed flattened shape at low density but spindle-like structures at confluency as judged by phalloidin staining. CONCLUSIONS: Osteogenic potential of ASCs is impaired by successive passaging and may not serve as a useful clinical source of osteogenic ASCs past P2.
OBJECTIVE: The effect of in vitro expansion of human adipose-derived stem cells (ASCs) on stem cell properties is controversial. We examined serial subcultivation with expansion on the ability of ASCs to grow and differentiate into osteoblastic lineages. DESIGN: Flow cytometric analysis, growth kinetics, cell population doubling time, light microscopy and confocal analysis, and osteogenesis induction were performed to assess growth and osteogenic potential of subcultivated ASCs at passages 2 (P2), P4 and P6. RESULTS: Flow cytometric analysis revealed that ASCs at P2 express classical mesenchymal stem cell markers including CD44, CD73, and CD105, but not CD14, CD19, CD34, CD45, or HLA-DR. Calcium deposition and alkaline phosphatase activity were the highest at P2 but completely abrogated at P4. Increased passage number impaired cell growth; P2 cultures exhibited exponential growth, while cells at P4 and P6 showed near linear growth with cell population doubling times increased from 3.2 at P2 to 4.8 d at P6. Morphologically, cells in various subcultivation stages showed flattened shape at low density but spindle-like structures at confluency as judged by phalloidin staining. CONCLUSIONS: Osteogenic potential of ASCs is impaired by successive passaging and may not serve as a useful clinical source of osteogenic ASCs past P2.
Authors: James B Mitchell; Kevin McIntosh; Sanjin Zvonic; Sara Garrett; Z Elizabeth Floyd; Amy Kloster; Yuan Di Halvorsen; Robert W Storms; Brian Goh; Gail Kilroy; Xiying Wu; Jeffrey M Gimble Journal: Stem Cells Date: 2005-12-01 Impact factor: 6.277
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Authors: J G Toma; M Akhavan; K J Fernandes; F Barnabé-Heider; A Sadikot; D R Kaplan; F D Miller Journal: Nat Cell Biol Date: 2001-09 Impact factor: 28.824
Authors: Laura de Girolamo; Silvia Lopa; Elena Arrigoni; Matteo F Sartori; Franz W Baruffaldi Preis; Anna T Brini Journal: Cytotherapy Date: 2009 Impact factor: 5.414
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Authors: Alina Mieczkowska; Adriana Schumacher; Natalia Filipowicz; Anna Wardowska; Maciej Zieliński; Piotr Madanecki; Ewa Nowicka; Paulina Langa; Milena Deptuła; Jacek Zieliński; Karolina Kondej; Alicja Renkielska; Patrick G Buckley; David K Crossman; Michael R Crowley; Artur Czupryn; Piotr Mucha; Paweł Sachadyn; Łukasz Janus; Piotr Skowron; Sylwia Rodziewicz-Motowidło; Mirosława Cichorek; Michał Pikuła; Arkadiusz Piotrowski Journal: Sci Rep Date: 2018-07-27 Impact factor: 4.379