Annulus fibrosus cells can induce mineralization: an in vitro study

Abstract Background context There is still no consensus as to whether the calcification observed in degenerate intervertebral discs (IVDs) is a cause or a consequence of disc degeneration. Purpose To investigate the mineralization potential of healthy (independent of other associated changes) annulu...

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Bibliographic Details
Published in:The spine journal 2013-04, Vol.13 (4), p.443-453
Main Authors: Nosikova, Yaroslavna, MASc, Santerre, J. Paul, PhD, Grynpas, Marc D., PhD, Kandel, Rita A., MD
Format: Article
Language:English
Subjects:
Alkaline phosphatase
Alkaline Phosphatase - metabolism
Animals
Annulus fibrosus
Calcinosis - etiology
Calcinosis - metabolism
Cattle
Cells, Cultured
Hydroxyapatite
Intervertebral Disc - cytology
Intervertebral Disc - metabolism
Intervertebral Disc Degeneration - metabolism
Mineralization
Orthopedics
β-Glycerophosphate
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Summary:Abstract Background context There is still no consensus as to whether the calcification observed in degenerate intervertebral discs (IVDs) is a cause or a consequence of disc degeneration. Purpose To investigate the mineralization potential of healthy (independent of other associated changes) annulus fibrosus (AF) cells under controlled in vitro conditions. Study design/setting In vitro study to investigate the mineralization potential of the AF cells. Methods Annulus fibrosus cells, isolated from bovine IVDs, were grown in monolayer. The effect of cell density, culture time, age of cell source, and passage on the percentage of AF cells with alkaline phosphatase activity (ALPa) was evaluated. Gene expression of mineralization-associated markers was determined. Cells were immunostained for Type I, II, and X collagens. To study mineralization potential, AF cells and AF cells that were sorted into two populations, high (top 5%±1%) or low (bottom 5%±1%) ALPa expressors, were grown in the presence of β-glycerophosphate for 2 weeks. Results The percentage of AF cells that express ALPa changes with time in culture and seeding density for primary immature and mature cell sources but not for passaged cells. Gene expression of ALP, matrix metallopeptidase-13 (MMP-13), osteopontin, and runt-related transcription factor 2 was upregulated by Day 7. Under mineralization-inducing conditions, high ALPa expressors and unsorted AF cells formed von Kossa–positive nodules, composed of hydroxyapatite as determined by electron diffraction analysis. Low ALPa expressors had significantly fewer von Kossa–positive nodules (p
ISSN:1529-9430
1878-1632
DOI:10.1016/j.spinee.2012.11.050