Stem cells 2013 July-10


Sequential differentiation of mesenchymal stem cells in an agarose scaffold promotes a physis-like zonal alignment of chondrocytes.

J Orthop Res.2012 Nov;30(11):1753-1759.

Schmitt JF, See KH, Yang Z, Hui JHP, Lee EH

Department of Orthopaedic Surgery, Yong Loo Lin School of Medicine & Tissue Engineering Program, National University of Singapore, Singapore


Background: The epiphyseal growth plate (physis) is a cartilaginous tissue responsible for elongation of the long bones in children. Chondrocytes of the physis differentiate and mature in defined linear zones.  Disrupted function of any of these zones would lead to abnormal bone growth resulting in shortening or angular deformity.

Objective: The current study is an attempt to recreate the physis in vitro by the differentiation of human bone marrow derived mesenchymal stem cells (hBMSCs) into zonal physeal cartilage in an agarose scaffold supplemented with growth factors.

Design: hBMSCs were embedded in a layered agarose scaffold with only the surface of the scaffold in direct contact with the culture medium. The cells were differentiated using a two-step system involving the sequential addition of TGFb followed by BMP2.

Results: The resultant samples displayed a heterogenous population of physis-like collagen type 2 positive cells including proliferating chondrocytes and mature chondrocytes showing hypertrophy, expression of early bone markers and matrix mineralization. Histological analysis revealed a physis-like linear zonal alignment of chondrocytes in varying stages of differentiation. The less mature chondrocytes were seen at the base of the construct while hypertrophic chondrocytes and matrix mineralization was observed closer to the surface of the construct. The formation of a distinct late hypertrophy, matrix mineralization zone was observed in the samples treated by TGFb followed by BMP. The expression profiling study revealed that in the BMP samples, BMP receptors and signaling factors were upregulated while TGFb receptors were downregulated. This was not seen in the TGF/BMP samples suggesting that continuous presence of TGFb blocks BMP function through down regulation of BMP signaling factors.

Conclusion: This protocol could be used to examine the effects of various culture conditions and growth factors on the differentiation, proliferation, maturation and zonal alignment of physeal chondrocytes facilitating the development of improved and targeted therapeutics for physeal defects.

PMID: 22517299

 LEE Eng Hin(Figures adapted from the original publication)

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