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Interferon gamma and T cells inhibit osteogenesis induced by allogeneic mesenchymal stromal cells.

ABSTRACT: The mesenchymal stromal cells (MSCs) are reported to be immunoprivileged and osteogenic. We hypothesized that the use of allogeneic MSCs for bone repair was possible if they displayed an ability to induce similar osteogenesis in syngeneic as well as in allogeneic hosts. To test this hypothesis we used a cloned bone marrow derived cell, termed D1, isolated from Balb/c mice. The D1 cells were subcutaneously injected in syngeneic Balb/c, allogeneic immunocompetent B6, allogeneic T-cell deficient NCr nude, and allogeneic B6 Pfp-/- Rag2-/- mice that lack matured T and B cells as well as NK-cell cytolytic functions. D1 cells formed ectopic bones only in syngeneic or allogeneic immunocompromised hosts but not in allogeneic B6 hosts. The lack of T cells alone in allogeneic NCr mice was sufficient to promote osteogenesis in allogeneic environment. We observed a significantly higher number of T cells, B cells, macrophages and significantly higher expression of interferon gamma (IFN-?) in B6 allogeneic implants as compared to the syngeneic implants. These factors correlated with severe inhibition of expression of alkaline phosphatase, osteocalcin, and runx2 genes in the implants from B6 mice. Our data suggest that strategies to inhibit T cells and IFN-? functions will be useful for bone repair mediated by allogeneic MSCs.

SUBMITTER: Dighe AS

PROVIDER: S-EPMC3510319 | biostudies-other | 2013 Feb

REPOSITORIES: biostudies-other

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Interferon gamma and T cells inhibit osteogenesis induced by allogeneic mesenchymal stromal cells.

Dighe Abhijit S AS Yang Scott S Madhu Vedavathi V Balian Gary G Cui Quanjun Q

Journal of orthopaedic research : official publication of the Orthopaedic Research Society 20120810 2

The mesenchymal stromal cells (MSCs) are reported to be immunoprivileged and osteogenic. We hypothesized that the use of allogeneic MSCs for bone repair was possible if they displayed an ability to induce similar osteogenesis in syngeneic as well as in allogeneic hosts. To test this hypothesis we used a cloned bone marrow derived cell, termed D1, isolated from Balb/c mice. The D1 cells were subcutaneously injected in syngeneic Balb/c, allogeneic immunocompetent B6, allogeneic T-cell deficient NC ...[more]

PMID: 22886855

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Project description:BackgroundMesenchymal stromal cells (MSCs) are believed to be hypoimmunogeneic with potential use for allogeneic administration.MethodsBone marrow was harvested from Connemara (n = 1), Standardbred (n = 6), and Thoroughbred (n = 3) horses. MSCs were grouped by their level of expression of major histocompatibility factor II (MHC II). MSCs were then sub-grouped by those MSCs derived from universal blood donor horses. MSCs were isolated and cultured using media containing fetal bovine serum until adequate numbers were acquired. The MSCs were cultured in xenogen-free media for 48 h prior to use and during all assays. Autologous and allogeneic MSCs were then directly co-cultured with responder leukocytes from the Connemara horse in varying concentrations of MSCs to leukocytes (1:1, 1:10, and 1:100). MSCs were also cultured with complement present and heat-inactivated complement to determine whether complement alone would decrease MSC viability. MSCs underwent haplotyping of their equine leukocyte antigen (ELA) to determine whether the MHC factors were matched or mismatched between the donor MSCs and the responder leukocytes.ResultsAll allogeneic MSCs were found to be ELA mismatched with the responder leukocytes. MHC II-low and universal blood donor MSCs caused no peripheral blood mononuclear cell (PBMC) proliferation, no increase in B cells, and no activation of CD8 lymphocytes. Universal blood donor MSCs stimulated a significant increase in the number of T regulatory cells. Neutrophil interaction with MSCs showed that universal blood donor and MHC II-high allogeneic MSCs at the 6 h time point in co-culture caused greater neutrophil activation than the other co-culture groups. Complement-mediated cytotoxicity did not consistently cause MSC death in cultures with active complement as compared to those with inactivated complement. Gene expression assays revealed that the universal blood donor group and the MHC II-low MSCs were more metabolically active both in the anabolic and catabolic gene categories when cultured with allogeneic lymphocytes as compared to the other co-cultures. These upregulated genes included CD59, FGF-2, HGF, IDO, IL-10, IL-RA, IL-2, SOX2, TGF-β1, ADAMSTS-4, ADAMSTS-5, CCL2, CXCLB/IL-8, IFNγ, IL-1β, and TNFα.ConclusionsMHC II-low MSCs are the most appropriate type of allogeneic MSC to prevent activation of the innate and cell-mediated component of the adaptive immune systems and have increased gene expression as compared to other allogeneic MSCs.

Project description:BackgroundThe intermediate filament protein nestin is used as a marker for neural stem cells, and its expression is inversely correlated with cellular differentiation. More recently, nestin expression has also been described in other cell types including multipotential mesenchymal stromal cells (MSCs). In this study, we examined the expression of nestin in equine, canine and human bone marrow-derived MSCs undergoing osteogenic differentiation, to determine whether nestin levels were attenuated as the cells acquired a more mature phenotype. In addition, the expression of nestin may be under the influence of cellular hypoxia, as nestin expression is known to increase in areas of ischemic tissue damage. Therefore, we also examined the effects of hypoxia on expression of nestin in human MSCs and examined a role for hypoxia inducible factor 1-alpha (HIF-1α) and vascular endothelial growth factor (VEGF) in the response. Additionally, we quantified the temporal expression of nestin in the fracture callus during bone regeneration, a site that has been characterized as hypoxic.ResultsThere were no significant changes in nestin expression in MSCs during osteogenic differentiation. There was a significant increase in expression of nestin mRNA and protein in human MSCs in response to hypoxia (1% O2) or the chemical hypoxia mimetic desferroxamine. This may be due to upregulation of VEGF under hypoxia, as treatment of cells with the VEGF receptor antagonist CPO-P11 attenuated hypoxia-induced nestin expression. A significant increase in nestin mRNA expression was observed in the fracture callus of mice three and seven days post fracture.ConclusionsNestin was not a selective marker for MSCs, as its expression was maintained during osteogenic differentiation, in all species examined. Furthermore our data suggest that nestin expression can be induced by hypoxia, and that this increase in nestin is partially regulated by HIF-1α and VEGF. Interestingly, nestin levels were significantly upregulated at the fracture site. Further studies are required to understand the role of nestin in bone cell biology and ultimately bone regeneration.

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Interferon gamma and T cells inhibit osteogenesis induced by allogeneic mesenchymal stromal cells.

Publications

Interferon gamma and T cells inhibit osteogenesis induced by allogeneic mesenchymal stromal cells.

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