title: Bone marrow-derived mesenchymal stem cells from early diffuse systemic sclerosis exhibit a paracrine machinery and stimulate angiogenesis in vitro.
abstract: OBJECTIVE
  To characterise bone marrow-derived mesenchymal stem cells (MSCs) from patients with systemic sclerosis (SSc) for the expression of factors implicated in MSC recruitment at sites of injury, angiogenesis and fibrosis. The study also analysed whether the production/release of bioactive mediators by MSCs were affected by stimulation with cytokines found upregulated in SSc serum and tissues, and whether MSCs could modulate dermal microvascular endothelial cell (MVEC) angiogenesis.
  
  
  METHODS
  MSCs obtained from five patients with early severe diffuse SSc (SSc-MSCs) and five healthy donors (H-MSCs) were stimulated with vascular endothelial growth factor (VEGF), transforming growth factor β (TGFβ) or stromal cell-derived factor-1 (SDF-1). Transcript and protein levels of SDF-1 and its receptor CXCR4, VEGF, TGFβ(1) and receptors TβRI and TβRII were evaluated by quantitative real-time PCR, western blotting and confocal microscopy. VEGF, SDF-1 and TGFβ(1) secretion in culture supernatant was measured by ELISA. MVEC capillary morphogenesis was performed on Matrigel with the addition of MSC-conditioned medium.
  
  
  RESULTS
  In SSc-MSCs the basal expression of proangiogenic SDF-1/CXCR4 and VEGF was significantly increased compared with H-MSCs. SSc-MSCs constitutively released higher levels of SDF-1 and VEGF. SDF-1/CXCR4 were upregulated after VEGF stimulation and CXCR4 redistributed from the cytoplasm to the cell surface. VEGF was increased by SDF-1 challenge. VEGF, TGFβ and SDF-1 stimulation upregulated TGFβ(1), TβRI and TβRII in SSc-MSCs. TβRII redistributed from the cytoplasm to focal adhesion contacts. SSc-MSC-conditioned medium showed a greater proangiogenic effect on MVECs than H-MSCs. Experiments with blocking antibodies showed that MSC-derived cytokines were responsible for this potent proangiogenic effect.
  
  
  CONCLUSIONS
  SSc-MSCs constitutively overexpress and release bioactive mediators/proangiogenic factors and potentiate dermal MVEC angiogenesis.
authors.affiliation: Department of Biomedicine, Division of Rheumatology, AOUC, Excellence Centre for Research, Transfer and High Education DENOthe, University of Florence, Florence, Italy.
authors.forename: Serena
authors.initials: S
authors.lastname: Guiducci
authors.forename: Mirko
authors.initials: M
authors.lastname: Manetti
authors.forename: Eloisa
authors.initials: E
authors.lastname: Romano
authors.forename: Benedetta
authors.initials: B
authors.lastname: Mazzanti
authors.forename: Claudia
authors.initials: C
authors.lastname: Ceccarelli
authors.forename: Simone
authors.initials: S
authors.lastname: Dal Pozzo
authors.forename: Anna Franca
authors.initials: AF
authors.lastname: Milia
authors.forename: Silvia
authors.initials: S
authors.lastname: Bellando-Randone
authors.forename: Ginevra
authors.initials: G
authors.lastname: Fiori
authors.forename: Maria Letizia
authors.initials: ML
authors.lastname: Conforti
authors.forename: Riccardo
authors.initials: R
authors.lastname: Saccardi
authors.forename: Lidia
authors.initials: L
authors.lastname: Ibba-Manneschi
authors.forename: Marco
authors.initials: M
authors.lastname: Matucci-Cerinic
mesh_terms: D000293:Adolescent; D000328:Adult; D001854:Bone Marrow Cells / Q000502:physiology; D002454:Cell Differentiation / Q000502:physiology; D002455:Cell Division / Q000502:physiology; D002478:Cells, Cultured; D054377:Chemokine CXCL12 / Q000378:metabolism; D003114:Colony-Forming Units Assay; D017077:Culture Media, Conditioned; D042783:Endothelial Cells / Q000502:physiology; D005260:Female; D006801:Humans; D016130:Immunophenotyping; D059630:Mesenchymal Stem Cells / Q000276:immunology / Q000378:metabolism / Q000502:physiology*; D009389:Neovascularization, Pathologic / Q000473:pathology*; D019899:Paracrine Communication / Q000502:physiology*; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D000077293:Receptor, Transforming Growth Factor-beta Type I; D000077294:Receptor, Transforming Growth Factor-beta Type II; D019718:Receptors, CXCR4 / Q000378:metabolism; D018125:Receptors, Transforming Growth Factor beta / Q000378:metabolism; D045743:Scleroderma, Diffuse / Q000378:metabolism / Q000473:pathology*; D012867:Skin / Q000098:blood supply*; D042461:Vascular Endothelial Growth Factor A / Q000378:metabolism; D055815:Young Adult
chemical_list: C527276:CXCR4 protein, human; D054377:Chemokine CXCL12; D017077:Culture Media, Conditioned; D019718:Receptors, CXCR4; D018125:Receptors, Transforming Growth Factor beta; C467484:VEGFA protein, human; D042461:Vascular Endothelial Growth Factor A; D017346:Protein Serine-Threonine Kinases; D000077293:Receptor, Transforming Growth Factor-beta Type I; D000077294:Receptor, Transforming Growth Factor-beta Type II