Xenotransplantation of interferon-gamma-pretreated clumps of a human mesenchymal stem cell/extracellular matrix complex induces mouse calvarial bone regeneration

Stem Cell Res Ther. 2017 Apr 26;8(1):101. doi: 10.1186/s13287-017-0550-1.


Background: Three-dimensional cultured clumps of a mesenchymal stem cell (MSC)/extracellular matrix (ECM) complex (C-MSC) consists of cells and self-produced ECM. C-MSC can regulate the cellular function in vitro and induce successful bone regeneration using ECM as a cell scaffold. Potentiating the immunomodulatory capacity of C-MSCs, which can ameliorate the allo-specific immune response, may be helpful in developing beneficial "off-the-shelf" cell therapy for tissue regeneration. It is well reported that interferon (IFN)-γ stimulates the immunosuppressive properties of MSC via upregulation of the immunomodulatory enzyme IDO. Therefore, the aim of this study was to investigate the effect of IFN-γ on the immunomodulatory capacity of C-MSC in vitro and to test the bone regenerative activity of C-MSC or IFN-γ-pretreated C-MSC (C-MSCγ) xenografts in a mice calvarial defect model.

Methods: Human bone marrow-derived MSCs were seeded at a density of 2.0 × 105 cells/well into 24-well plates and cultured with growth medium supplemented with 50 μg/mL L-ascorbic acid for 4 days. To obtain C-MSC, confluent cells that had formed on the cellular sheet were scratched using a micropipette tip and were then torn off. The cellular sheet was rolled to make a round clump of cells. C-MSC was stimulated with IFN-γ and IDO expression, immunosuppressive capacity, and immunophenotype were evaluated in vitro. Moreover, C-MSC or C-MSCγ was xenotransplanted into immunocompetent or immunodeficient mice calvarial defect models without artificial scaffold, respectively.

Results: IFN-γ stimulated IDO expression in C-MSC. C-MSCγ, but not C-MSC, attenuated CD3/CD28-induced T cell proliferation and its suppressive effect was reversed by an IDO inhibitor. C-MSCγ showed upregulation of HLA-DR expression, but its co-stimulatory molecule, CD86, was not detected. Xenotransplantation of C-MSCγ into immunocompetent mice calvarial defect induced bone regeneration, whereas C-MSC xenograft failed and induced T cell infiltration in the grafted area. On the other hand, both C-MSC and C-MSCγ xenotransplantation into immunodeficient mice caused bone regeneration.

Conclusions: Xenotransplantation of C-MSCγ, which exerts immunomodulatory properties via the upregulation of IDO activity in vitro, may attenuate xenoreactive host immune response, and thereby induce bone regeneration in mice. Accordingly, C-MSCγ may constitute a promising novel allograft cell therapy for bone regeneration.

Keywords: C-MSC; IDO; IFN-γ; Xenotransplantation.

MeSH terms

  • Animals
  • Ascorbic Acid / pharmacology
  • Bone Marrow Cells / cytology
  • Bone Regeneration / physiology*
  • Cell Proliferation / drug effects
  • Cells, Cultured
  • Disease Models, Animal
  • HLA-DR Antigens / metabolism
  • Humans
  • Indoleamine-Pyrrole 2,3,-Dioxygenase / genetics
  • Indoleamine-Pyrrole 2,3,-Dioxygenase / metabolism
  • Interferon-gamma / pharmacology*
  • Leukocytes, Mononuclear / cytology
  • Leukocytes, Mononuclear / metabolism
  • Male
  • Mesenchymal Stem Cell Transplantation*
  • Mesenchymal Stem Cells / cytology
  • Mesenchymal Stem Cells / drug effects*
  • Mesenchymal Stem Cells / metabolism
  • Mice
  • Mice, Inbred C57BL
  • Mice, Inbred NOD
  • Mice, SCID
  • Skull / physiology*
  • T-Lymphocytes / cytology
  • T-Lymphocytes / immunology
  • T-Lymphocytes / metabolism
  • Transplantation, Heterologous


  • HLA-DR Antigens
  • Indoleamine-Pyrrole 2,3,-Dioxygenase
  • Interferon-gamma
  • Ascorbic Acid