Matrix stiffness regulation of integrin-mediated mechanotransduction during osteogenic differentiation of human mesenchymal stem cells

Authors

  • Yu-Ru V Shih,

    1. Institute of Clinical Medicine, National Yang-Ming University, Taipei, Taiwan
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  • Kuo-Fung Tseng,

    Corresponding author
    1. Department of Orthopaedics, Cheng Hsin General Hospital, Taipei, Taiwan
    • Department of Orthopaedics, Cheng Hsin General Hospital, No.45, Cheng Hsin Street, Beitou District, Taipei City, Taiwan, Republic of China.
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  • Hsiu-Yu Lai,

    1. Institute of Clinical Medicine, National Yang-Ming University, Taipei, Taiwan
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  • Chi-Hung Lin,

    1. Institute of Microbiology and Immunology, National Yang-Ming University, Taipei, Taiwan
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  • Oscar K Lee

    Corresponding author
    1. Institute of Clinical Medicine, National Yang-Ming University, Taipei, Taiwan
    2. Department of Orthopaedics and Traumatology, Taipei Veterans General Hospital, Taipei, Taiwan
    3. Stem Cell Research Center, National Yang-Ming University, Taipei, Taiwan
    • Department of Orthopaedics and Traumatology, Taipei Veterans General Hospital, 201, Sec. 2, Shipai Road, Beitou District, Taipei City, Taiwan 11217, Republic of China.
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Abstract

Mesenchymal stem cells (MSCs) cultured on extracellular matrices with different stiffness have been shown to possess diverse lineage commitment owing to the extracellular mechanical stimuli sensed by the cells. The aim of this study was to further delineate how matrix stiffness affects intracellular signaling through the mechanotransducers Rho kinase (ROCK) and focal adhesion kinase (FAK) and subsequently regulates the osteogenic phenotype of MSCs. MSCs were cultured in osteogenic medium on tunable polyacrylamide hydrogels coated with type I collagen with elasticities corresponding to Young's modulus of 7.0 ± 1.2 and 42.1 ± 3.2 kPa. Osteogenic differentiation was increased on stiffer matrices, as evident by type I collagen, osteocalcin, and Runx2 gene expressions and alizarin red S staining for mineralization. Western blot analysis demonstrated an increase in kinase activities of ROCK, FAK, and ERK1/2 on stiffer matrices. Inhibition of FAK, an important mediator of osteogenic differentiation, and inhibition of ROCK, a known mechanotransducer of matrix stiffness during osteogenesis, resulted in decreased expression of osteogenic markers during osteogenic induction. In addition, FAK affects osteogenic differentiation through ERK1/2, whereas ROCK regulates both FAK and ERK1/2. Furthermore, α2-integrin was upregulated on stiffer matrices during osteogenic induction, and its knockdown by siRNA downregulated the osteogenic phenotype through ROCK, FAK, and ERK1/2. Taken together, our results provide evidence that the matrix rigidity affects the osteogenic outcome of MSCs through mechanotransduction events that are mediated by α2-integrin. © 2011 American Society for Bone and Mineral Research.

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