The effect of Link N on differentiation of human bone marrow-derived mesenchymal stem cellsReport as inadecuate

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Arthritis Research and Therapy

, 14:R267

First Online: 10 December 2012Received: 28 June 2012Revised: 16 November 2012Accepted: 05 December 2012


IntroductionWe previously showed that Link N can stimulate extracellular matrix biosynthesis by intervertebral disc IVD cells, both in vitro and in vivo, and is therefore a potential stimulator of IVD repair. The purpose of the present study was to determine how Link N may influence human mesenchymal stem cell MSC differentiation, as a prelude to using Link N and MSC supplementation in unison for optimal repair of the degenerated disc.

MethodsMSCs isolated from the bone marrow of three osteoarthritis patients were cultured in chondrogenic or osteogenic differentiation medium without or with Link N for 21 days. Chondrogenic differentiation was monitored by proteoglycan staining and quantitation by using Alcian blue, and osteogenic differentiation was monitored by mineral staining and quantitation by using Alzarin red S. In addition, proteoglycan secretion was monitored with the sulfated glycosaminoglycan GAG content of the culture medium, and changes in gene expression were analyzed with real-time reverse transcription RT PCR.

ResultsLink N alone did not promote MSC chondrogenesis. However, after MSCs were supplemented with Link N in chondrogenic differentiation medium, the quantity of GAG secreted into the culture medium, as well as aggrecan, COL2A1, and SOX9 gene expression, increased significantly. The gene expression of COL10A1 and osteocalcin OC were downregulated significantly. When MSCs were cultured in osteogenic differentiation medium, Link N supplementation led to a significant decrease in mineral deposition, and alkaline phosphatase ALP, OC, and RUNX2 gene expression.

ConclusionsLink N can enhance chondrogenic differentiation and downregulate hypertrophic and osteogenic differentiation of human MSCs. Therefore, in principle, Link N could be used to optimize MSC-mediated repair of the degenerated disc.


ALPalkaline phosphatase

COL2A1collagen, type 2, alpha 1

DMMB1,9-dimethylmethylene blue

ECMextracellular matrix

GAGsulfated glycosaminoglycan

GAPDHglyceraldehyde-3-phosphate dehydrogenase

IVDintervertebral disc

MSCmesenchymal stem cell

NPnucleus pulposus


PCRpolymerase chain reaction

RUNX2Runt-related transcription factor 2

SOX9SRY sex-determining region Y-box 9.

Electronic supplementary materialThe online version of this article doi:10.1186-ar4113 contains supplementary material, which is available to authorized users.

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Author: John Antoniou - Hong Tian Wang - Abdulrahman M Alaseem - Lisbet Haglund - Peter J Roughley - Fackson Mwale


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