Analysis of Molecular Expression in Adipose Tissue-Derived Mesenchymal Stem Cells : Prospects for Use in the Treatment of Intervertebral Disc Degeneration

Abstract

ObjectiveRecent studies have shown encouraging progress toward the use of autogenic and allogenic mesenchymal stem cells (MSCs) to arrest, or even lead to partial regeneration in, intervertebral disc (IVD) degeneration. However, this technology is still in its infancy, and further development is required. The aim of this study was to analyze whether rat adipose-derived mesenchymal stem cells (ADMSC) can differentiate towards IVD-like cells after treatment with transforming growth factor β3 (TGF-β3) in vitro. We also performed quantitative analysis of gene expression for ADMSC only, ADMSCs treated with TGF-β3, and co-cultured ADMSCs treated with TGF-β3.MethodsADMSCs were sub-cultured to homogeneity and used in fluorocytometry assays for CD11, CD45, and CD90/Thy1. ADMSCs were differentiated in spheroid culture towards the chondrogenic lineage by the presence of TGF-β3, dexamethasone, and ascorbate. We also co-cultured pure ADMSCs and nucleus pulposus cells in 24-well plates, and performed immunohistochemical staining, western blotting, and RT-PCR for quantitativeanalysis of gene expression.ResultsResults of fluorocytometry were positive for CD90/Thy1 and negative for CD11 and CD45. TGF-β3-mediated induction of ADMSCs led to the expression of the differentiation markers of intervertebral disc-like cells, such as aggrecan, collagen II, and sox-9. Co-cultured ADMSCs treated with TGF-β3 showed higher expression of differentiation markers and greater extracellular matrix production compared with ADMSCs treated with TGF-β3 alone.ConclusionADMSC treated with TGF-β3 may be an attractive source for regeneration therapy in degenerative IVD. These findings may also help elucidate the pathologic mechanism of MSC therapy in the degeneration of IVD in vivo.