Dataset Information

Association between in vivo bone formation and ex vivo migratory capacity of human bone marrow stromal cells.

ABSTRACT: There is a clinical need for developing systemic transplantation protocols for use of human skeletal stem cells (also known bone marrow stromal stem cells) (hBMSC) in tissue regeneration. In systemic transplantation studies, only a limited number of hBMSC home to injured tissues suggesting that only a subpopulation of hBMSC possesses "homing" capacity. Thus, we tested the hypothesis that a subpopulation of hBMSC defined by ability to form heterotopic bone in vivo, is capable of homing to injured bone.We tested ex vivo and in vivo homing capacity of a number of clonal cell populations derived from telomerized hBMSC (hBMSC-TERT) with variable ability to form heterotopic bone when implanted subcutaneously in immune deficient mice. In vitro transwell migration assay was used and the in vivo homing ability of transplanted hBMSC to bone fractures in mice was visualized by bioluminescence imaging (BLI). In order to identify the molecular phenotype associated with enhanced migration, we carried out comparative DNA microarray analysis of gene expression of hBMSC-derived high bone forming (HBF) clones versus low bone forming (LBF) clones.HBF clones were exhibited higher ex vivo transwell migration and following intravenous injection, better in vivo homing ability to bone fracture when compared to LBF clones. Comparative microarray analysis of HBF versus LBF clones identified enrichment of gene categories of chemo-attraction, adhesion and migration associated genes. Among these, platelet-derived growth factor receptor (PDGFR) ? and ? were highly expressed in HBF clones. Follow up studies showed that the chemoattractant effects of PDGF in vitro was more enhanced in HBF compared to LBF clones and this effect was reduced in presence of a PDGFR?-specific inhibitor: SU-16 f. Also, PDGF exerted greater chemoattractant effect on PDGFR?(+) cells sorted from LBF clones compared to PDGFR?(-) cells.Our data demonstrate phenotypic and molecular association between in vivo bone forming ability and migratory capacity of hBMSC. PDGFR? can be used as a potential marker for the prospective selection of hBMSC populations with high migration and bone formation capacities suitable for clinical trials for enhancing bone regeneration.

SUBMITTER: Andersen RK

PROVIDER: S-EPMC4599318 | biostudies-literature | 2015

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Association between in vivo bone formation and ex vivo migratory capacity of human bone marrow stromal cells.

Andersen Rikke K RK Zaher Walid W Larsen Kenneth H KH Ditzel Nicholas N Drews Katharina K Wruck Wasco W Adjaye James J Abdallah Basem M BM Kassem Moustapha M

Stem cell research & therapy 20151008

<h4>Introduction</h4>There is a clinical need for developing systemic transplantation protocols for use of human skeletal stem cells (also known bone marrow stromal stem cells) (hBMSC) in tissue regeneration. In systemic transplantation studies, only a limited number of hBMSC home to injured tissues suggesting that only a subpopulation of hBMSC possesses "homing" capacity. Thus, we tested the hypothesis that a subpopulation of hBMSC defined by ability to form heterotopic bone in vivo, is capable ...[more]

PMID: 26450135

Similar Datasets

Project description:Introduction: There is a clinical need for developing systemic transplantation protocols for use of human skeletal stem cells (also known bone marrow stromal stem cells) (hBMSC) in tissue regeneration. In systemic transplantation studies, only a limited number of hBMSC home to injured tissues suggesting that only a subpopulation of hBMSC possess âhomingâ capacity. Thus, we tested the hypothesis that a subpopulation of hBMSC defined by ability to form heterotopic bone in vivo, is capable of homing to injured bone. Methods: We tested ex vivo and in vivo homing capacity of a number of clonal cell populations derived from telomerized hBMSC (hBMSC-TERT) with variable ability to form heterotopic bone when implanted subcutaneously in immune deficient mice. In vitro transwell migration assay was used and in vivo homing ability to bone fractures in mice was visualized by bioluminescence imaging (BLI). In order to identify the molecular phenotype associated with enhanced migration, we carried out comparative DNA microarray analysis of gene expression of hBMSC-derived high bone forming (HBF) clones versus low bone forming (LBF) clones. Results: In this study, clonal cell populations forming in vivo bone were shown to exhibit higher ex vivo transwell migration and following intravenous infusion, enhanced in vivo homing ability to bone fractures. Comparative microarray analysis of LBF versus HBF clones identified a significant enrichment of gene categories of cell chemo-attraction, adhesion and migration. Among these genes, platelet-derived growth factor receptor (PDGF-R) Î± and Î² were highly expressed in HBF clones. Further studies showed that the chemoattractive effects of PDGF in vitro was more enhanced in HBF clones compared to LBF clones and this effect was abolished in presence of a PDGFR-Î²-specific inhibitor: SU-16f. Sorted PDGFR-Î²+ cells from LBF clones showed that the PDGFR-Î²+ enriched cell population exhibited strong chemoattractance towards PDGF. Conclusion: Our data demonstrate phenotypic and molecular association between in vivo bone formation and migratory capacity of hBMSC. PDGFR-Î² can be used as a potential marker for the prospective selection of hBMSC populations with high migration and bone formation capacities suitable for clinical trials for enhancing bone regeneration. Total RNA obtained from telomerized human bone marrow derived mesenchymal stromal cells (T4P38a) with high in vivo bone forming capacity and (T4P74a) with low bone forming capacity and three single cell clones with high bone forming capacities (DD8, AD10 and BB10) and three clones with low bone forming capacities (CF1, CB4 and CD4). All cells were cultured under standard conditions and RNAs were isolated at baseline with no induction. each sample was represented in doublicate as a and b.

Project description:Introduction: There is a clinical need for developing systemic transplantation protocols for use of human skeletal stem cells (also known bone marrow stromal stem cells) (hBMSC) in tissue regeneration. In systemic transplantation studies, only a limited number of hBMSC home to injured tissues suggesting that only a subpopulation of hBMSC possess “homing” capacity. Thus, we tested the hypothesis that a subpopulation of hBMSC defined by ability to form heterotopic bone in vivo, is capable of homing to injured bone. Methods: We tested ex vivo and in vivo homing capacity of a number of clonal cell populations derived from telomerized hBMSC (hBMSC-TERT) with variable ability to form heterotopic bone when implanted subcutaneously in immune deficient mice. In vitro transwell migration assay was used and in vivo homing ability to bone fractures in mice was visualized by bioluminescence imaging (BLI). In order to identify the molecular phenotype associated with enhanced migration, we carried out comparative DNA microarray analysis of gene expression of hBMSC-derived high bone forming (HBF) clones versus low bone forming (LBF) clones. Results: In this study, clonal cell populations forming in vivo bone were shown to exhibit higher ex vivo transwell migration and following intravenous infusion, enhanced in vivo homing ability to bone fractures. Comparative microarray analysis of LBF versus HBF clones identified a significant enrichment of gene categories of cell chemo-attraction, adhesion and migration. Among these genes, platelet-derived growth factor receptor (PDGF-R) α and β were highly expressed in HBF clones. Further studies showed that the chemoattractive effects of PDGF in vitro was more enhanced in HBF clones compared to LBF clones and this effect was abolished in presence of a PDGFR-β-specific inhibitor: SU-16f. Sorted PDGFR-β+ cells from LBF clones showed that the PDGFR-β+ enriched cell population exhibited strong chemoattractance towards PDGF. Conclusion: Our data demonstrate phenotypic and molecular association between in vivo bone formation and migratory capacity of hBMSC. PDGFR-β can be used as a potential marker for the prospective selection of hBMSC populations with high migration and bone formation capacities suitable for clinical trials for enhancing bone regeneration.

Project description:Mesenchymal stem cells (MSCs) are highly responsive to cues in the microenvironment (niche) that must be recapitulated ex vivo to study their authentic behavior. In this study, we hypothesized that native bone marrow (BM)- and adipose (AD)-derived extracellular matrices (ECM) were unique in their ability to control MSC behavior. To test this, we compared proliferation and differentiation of bone marrow (BM)-derived MSCs when maintained on native decellularized ECM produced by BM versus AD stromal cells (i.e. BM- versus AD-ECM). We found that both ECMs contained similar types of collagens but differed in the relative abundance of each. Type VI collagen was the most abundant (≈60% of the total collagen present), while type I was the next most abundant at ≈30%. These two types of collagen were found in nearly equal proportions in both ECMs. In contrast, type XII collagen was almost exclusively found in AD-ECM, while types IV and V were only found in BM-ECM. Physically and mechanically, BM-ECM was rougher and stiffer, but less adhesive, than AD-ECM. During 14 days in culture, both ECMs supported BM-MSC proliferation better than tissue culture plastic (TCP), although MSC-related surface marker expression remained relatively high on all three culture surfaces. BM-MSCs cultured in osteogenic (OS) differentiation media on BM-ECM displayed a significant increase in calcium deposition in the matrix, indicative of osteogenesis, while BM-MSCs cultured on AD-ECM in the presence of adipogenic (AP) differentiation media showed a significant increase in Oil Red O staining, indicative of adipogenesis. Further, culture on BM-ECM significantly increased BM-MSC-responsiveness to rhBMP-2 (an osteogenic inducer), while culture on AD-ECM enhanced responsiveness to rosiglitazone (an adipogenic inducer). These findings support our hypothesis and indicate that BM- and AD-ECMs retain unique elements, characteristic of their tissue-specific microenvironment (niche), which promote retention of MSC differentiation state (i.e. "stemness") during expansion and direct cell response to lineage-specific inducers. This study provides a new paradigm for precisely controlling MSC fate to a desired cell lineage for tissue-specific cell-based therapies.

Project description:BACKGROUND:The human bone marrow (BM) niche contains a population of mesenchymal stromal cells (MSCs) that provide physical support and regulate hematopoietic stem cell (HSC) homeostasis. β-Thalassemia (BT) is a hereditary disorder characterized by altered hemoglobin beta-chain synthesis amenable to allogeneic HSC transplantation and HSC gene therapy. Iron overload (IO) is a common complication in BT patients affecting several organs. However, data on the BM stromal compartment are scarce. METHODS:MSCs were isolated and characterized from BM aspirates of healthy donors (HDs) and BT patients. The state of IO was assessed and correlated with the presence of primitive MSCs in vitro and in vivo. Hematopoietic supportive capacity of MSCs was evaluated by transwell migration assay and 2D coculture of MSCs with human CD34+ HSCs. In vivo, the ability of MSCs to facilitate HSC engraftment was tested in a xenogenic transplant model, whereas the capacity to sustain human hematopoiesis was evaluated in humanized ossicle models. RESULTS:We report that, despite iron chelation, BT BM contains high levels of iron and ferritin, indicative of iron accumulation in the BM niche. We found a pauperization of the most primitive MSC pool caused by increased ROS production in vitro which impaired MSC stemness properties. We confirmed a reduced frequency of primitive MSCs in vivo in BT patients. We also discovered a weakened antioxidative response and diminished expression of BM niche-associated genes in BT-MSCs. This caused a functional impairment in MSC hematopoietic supportive capacity in vitro and in cotransplantation models. In addition, BT-MSCs failed to form a proper BM niche in humanized ossicle models. CONCLUSION:Our results suggest an impairment in the mesenchymal compartment of BT BM niche and highlight the need for novel strategies to target the niche to reduce IO and oxidative stress before transplantation. FUNDING:This work was supported by the SR-TIGET Core grant from Fondazione Telethon and by Ricerca Corrente.

Dataset Information

Association between in vivo bone formation and ex vivo migratory capacity of human bone marrow stromal cells.

Publications

Association between in vivo bone formation and ex vivo migratory capacity of human bone marrow stromal cells.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets