Differences in human mesenchymal stem cell secretomes during chondrogenic induction.

Mesenchymal stem cells (MSCs) can be induced towards chondrogenesis through the application of chondrogenic stimuli such as transforming growth factor-β (TGF-β) or by multiaxial mechanical load. Previous work has showed that the chondrogenic effect of multiaxial load on MSCs is mediated by the endogenous production of TGF-β1 by stimulated cells. This work compared the effects of TGF-β1 stimulation and multiaxial mechanical load on the secretomes of stimulated cells. MSCs were seeded into fibrin-poly(ester-urethane) scaffolds and chondrogenically stimulated with either TGF-β1 or mechanical load. The culture media was collected and analysed for 174 proteins using a cytokine antibody array. The results of the secretome analysis were then confirmed at a gene expression level by real-time PCR. As results implicated nitric oxide (NO), the media nitrite content was also determined as an indirect measurement of media NO levels. Results showed that TGF-β1 stimulation and mechanical load lead to similar changes in factors such as BLC, VEGF and MMP13, whilst differences in detected levels were seen for factors including leptin, MDC, MIP3α and LAP. Gene expression analysis confirmed significant changes in four factors: angiopoietin 2, GROα, MMP13 and osteoprotegerin. After one week in culture the media nitrite content was significantly higher in loaded groups than both control and TGF-β1 stimulated groups, suggesting this may be a major therapeutic target. These data show that despite clear similarities, TGF-β1 stimulation and load have distinct effects on MSCs and are not analogous. This study has identified a number of potentially novel targets for tissue engineering, these data may also be useful for improving rehabilitation protocols e.g. after microfracture.