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

J Bone Miner Res. 2011 Apr;26(4):730-8. doi: 10.1002/jbmr.278.

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, α(2)-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 α(2)-integrin.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Acrylic Resins / chemical synthesis
  • Acrylic Resins / chemistry
  • Biomechanical Phenomena
  • Calcification, Physiologic / drug effects
  • Calcification, Physiologic / physiology
  • Cell Differentiation / drug effects
  • Cell Differentiation / physiology*
  • Collagen Type I / chemistry
  • Collagen Type I / genetics
  • Core Binding Factor Alpha 1 Subunit / genetics
  • Elasticity / physiology*
  • Extracellular Matrix / physiology*
  • Focal Adhesion Kinase 1 / antagonists & inhibitors
  • Focal Adhesion Kinase 1 / metabolism
  • Gene Expression / drug effects
  • Gene Expression / genetics
  • Humans
  • Integrin alpha2 / genetics
  • Integrin alpha2 / metabolism*
  • Mechanotransduction, Cellular / physiology*
  • Mesenchymal Stem Cells / cytology*
  • Mesenchymal Stem Cells / drug effects
  • Mesenchymal Stem Cells / metabolism
  • Mitogen-Activated Protein Kinase 1 / antagonists & inhibitors
  • Mitogen-Activated Protein Kinase 1 / metabolism
  • Mitogen-Activated Protein Kinase 3 / antagonists & inhibitors
  • Mitogen-Activated Protein Kinase 3 / metabolism
  • Myosin-Light-Chain Phosphatase / metabolism
  • Osteoblasts / cytology
  • Osteoblasts / drug effects
  • Osteoblasts / metabolism
  • Osteocalcin / genetics
  • Osteogenesis / physiology*
  • Phosphorylation / drug effects
  • Protein Kinase Inhibitors / pharmacology
  • RNA, Small Interfering / genetics
  • beta Catenin / metabolism
  • rho-Associated Kinases / antagonists & inhibitors
  • rho-Associated Kinases / metabolism

Substances

  • Acrylic Resins
  • CTNNB1 protein, human
  • Collagen Type I
  • Core Binding Factor Alpha 1 Subunit
  • Integrin alpha2
  • Protein Kinase Inhibitors
  • RNA, Small Interfering
  • RUNX2 protein, human
  • beta Catenin
  • Osteocalcin
  • polyacrylamide
  • Focal Adhesion Kinase 1
  • PTK2 protein, human
  • rho-Associated Kinases
  • MAPK1 protein, human
  • Mitogen-Activated Protein Kinase 1
  • Mitogen-Activated Protein Kinase 3
  • Myosin-Light-Chain Phosphatase