Synthesis of embryonic tendon-like tissue by human marrow stromal/mesenchymal stem cells requires a three-dimensional environment and transforming growth factor β3

Matrix Biol. 2010 Oct;29(8):668-77. doi: 10.1016/j.matbio.2010.08.005. Epub 2010 Aug 22.

Abstract

Tendon-like tissue generated from stem cells in vitro has the potential to replace tendons and ligaments lost through injury and disease. However, thus far, no information has been available on the mechanism of tendon formation in vitro and how to accelerate the process. We show here that human mesenchymal stem cells (MSCs) and bone marrow-derived mononuclear cells (BM-MNCs) can generate tendon-like tissue in 7days mediated by transforming growth factor (TGF) β3. MSCs cultured in fixed-length fibrin gels spontaneously synthesized narrow-diameter collagen fibrils and exhibited fibripositors (actin-rich, collagen fibril-containing plasma membrane protrusions) identical to those that occur in embryonic tendon. In contrast, BM-MNCs did not synthesize tendon-like tissue under these conditions. We performed real-time PCR analysis of MSCs and BM-MNCs. MSCs upregulated genes encoding type I collagen, TGFβ3, and Smad2 at the time of maximum contraction of the tendon-like tissue (7days). Western blot analysis showed phosphorylation of Smad2 at maximum contraction. The TGFβ inhibitor SB-431542, blocked the phosphorylation of Smad2 and stopped the formation of tendon-like tissue. Quantitative PCR showed that BM-MNCs expressed very low levels of TGFβ3 compared to MSCs. Therefore we added exogenous TGFβ3 protein to BM-MNCs in fibrin gels, which resulted in phosphorylation of Smad2, synthesis of collagen fibrils, the appearance of fibripositors at the plasma membrane, and the formation of tendon-like tissue. In conclusion, MSCs that self-generate TGFβ signaling or the addition of TGFβ3 protein to BM-MNCs in fixed-length fibrin gels spontaneously make embryonic tendon-like tissue in vitro within 7days.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adipocytes / cytology
  • Adipocytes / metabolism
  • Adult
  • Antigens, CD / metabolism
  • Bone Marrow Cells / cytology*
  • Bone Marrow Cells / drug effects
  • Bone Marrow Cells / metabolism
  • Cell Differentiation / drug effects
  • Cell Differentiation / physiology
  • Chondrocytes / cytology
  • Chondrocytes / metabolism
  • Collagen Type I / genetics
  • Collagen Type I, alpha 1 Chain
  • Extracellular Matrix / ultrastructure
  • Fibrillar Collagens / metabolism
  • Fibrillar Collagens / ultrastructure
  • Fibrin / metabolism
  • Fibrinogen / metabolism
  • Gels / metabolism
  • Gels / pharmacology
  • Gene Expression / genetics
  • Humans
  • Integrin beta Chains / genetics
  • Mesenchymal Stem Cells / cytology*
  • Mesenchymal Stem Cells / drug effects
  • Mesenchymal Stem Cells / metabolism
  • Monocyte-Macrophage Precursor Cells / cytology
  • Monocyte-Macrophage Precursor Cells / drug effects
  • Monocyte-Macrophage Precursor Cells / metabolism
  • Osteoblasts / cytology
  • Osteoblasts / metabolism
  • Phosphorylation / drug effects
  • Signal Transduction / drug effects
  • Signal Transduction / physiology
  • Smad2 Protein / genetics
  • Smad2 Protein / metabolism
  • Stromal Cells / cytology*
  • Stromal Cells / drug effects
  • Stromal Cells / metabolism
  • Tendons / cytology
  • Tendons / embryology*
  • Tendons / metabolism
  • Thrombin / metabolism
  • Transforming Growth Factor beta3 / antagonists & inhibitors
  • Transforming Growth Factor beta3 / genetics
  • Transforming Growth Factor beta3 / metabolism*
  • Transforming Growth Factor beta3 / pharmacology
  • Young Adult

Substances

  • Antigens, CD
  • Collagen Type I
  • Collagen Type I, alpha 1 Chain
  • Fibrillar Collagens
  • Gels
  • Integrin beta Chains
  • SMAD2 protein, human
  • Smad2 Protein
  • Transforming Growth Factor beta3
  • Fibrin
  • Fibrinogen
  • Thrombin