Journal
INFLAMMATION RESEARCH
Volume 63, Issue 11, Pages 907-917Publisher
SPRINGER BASEL AG
DOI: 10.1007/s00011-014-0764-y
Keywords
Stem cells; Osteogenesis; Growth kinetics; Adipose tissues; Differentiation
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Funding
- Canadian Institute of Health Research Grant (CIHR)
- Lebanese National Council for Scientific Research (LNCSR)
- Canadian Institute of Health research (CIHR)
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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. 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. 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. Osteogenic potential of ASCs is impaired by successive passaging and may not serve as a useful clinical source of osteogenic ASCs past P2.
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