3-D Fibrin Scaffold Improves Stemness of Human Peripheral Blood Endothelial Progenitor Cells (Abstract/Comunicazione in rivista)

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  • 3-D Fibrin Scaffold Improves Stemness of Human Peripheral Blood Endothelial Progenitor Cells (Abstract/Comunicazione in rivista) (literal)
Anno
  • 2008-01-01T00:00:00+01:00 (literal)
Alternative label
  • Rossella Di Stefano (a,b), Monica Lemmi (a), Chiara Armani (a), Angela Magera (a), Maria Chiara Barsotti (a), Mariacarla Iorio (c), Elisa Simonetti (a), Roberta Arici (a), Dinuccio Dinucci (d), Federica Chiellini (d), Antonio Minnocci (e), Michele Alderighi (f), Roberto Solaro (f), Giorgio Soldani (g), Alberto Balbarini (b) (2008)
    3-D Fibrin Scaffold Improves Stemness of Human Peripheral Blood Endothelial Progenitor Cells
    (literal)
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  • Rossella Di Stefano (a,b), Monica Lemmi (a), Chiara Armani (a), Angela Magera (a), Maria Chiara Barsotti (a), Mariacarla Iorio (c), Elisa Simonetti (a), Roberta Arici (a), Dinuccio Dinucci (d), Federica Chiellini (d), Antonio Minnocci (e), Michele Alderighi (f), Roberto Solaro (f), Giorgio Soldani (g), Alberto Balbarini (b) (literal)
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  • Lavoro presentato al 69°Congresso Nazionale della Società Italiana di Cardiologia, 13-16 dicembre 2008, Roma (comunicazione orale). (literal)
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  • In: XV Congresso Nazionale della Società Italiana di Ricerche Cardiovascolari (SIRC) (Imola (Bologna), 9 - 11 ottobre 2008). Abstract, pp. 1 - 1. Società Italiana di Ricerche Cardiovascolari (SIRC), 2008. (literal)
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  • Aims Fibrin is a natural biopolymer appealing for cell-based regenerative therapies, because it can support growth, migration and differentiation of different cell types. Endothelial progenitor cells (EPC) represent a very interesting alternative cell source for mature endothelial cells; the fact that can easily isolated from the peripheral blood, thereby eliminating donor morbidity, makes them ideal in applications in the field of regenerative medicine. We have demonstrated that fibrin can support EPC viability and growth. Aim of this study was to evaluate if fibrin can affect EPC differentiation and stem cell markers expression. Methods Fibrin was prepared mixing commercially available (Kedrion S.p.A. Lucca, Italy) fibrinogen (9 mg/ml) and thrombin (25 U/ml). Clot ultrastructure was investigated by scanning electron microscopy (SEM) and cryogenic SEM (CRYO-SEM) to measure fibre diameter and density. Clot elasticity was evaluated by atomic force microscopy (AFM), measuring the tip-sample force by cantilever displacement. EPC were obtained from peripheral blood and cultured on fibrin at the concentration of 1x106cell/cm2. Fibronectin coating was used as a control. Metabolic activity was assessed after 7 and 14 days by WST1 assay and viability by confocal microscopy (calcein incorporation). The expression of both endothelial (CD31, KDR, vWF, Ve-Cadherin) and stem cell markers (nanog, oct-4) was assessed by flow cytometry, confocal microscopy and Real Time RT-PCR. Results SEM analysis revealed a nanometric fibrous structure, with mean fiber diameter of 165±4 nm and mean density of 95.9±0.2 %. CRYO-SEM suggested a reticulate structure with mesh-size up to 10 µm. Fibrin clot elasticity was 1.78 MPa, as in literature. WST1 assay showed that fibrin increased EPC metabolic activity as compared to fibronectin (fibrin: 0.606±0.056 a.u. vs. fibronectin: 0.311±0.067). Calcein staining demonstrated that EPC were still viable at 14 days. Flow cytometry showed the expression of endothelial markers (CD31=41.8±8.4%; vWF=32.3±3.0%; KDR=89.3±3.7%; VE-Cadherin=41.2±3.8%), confirmed also by confocal microscopy and Real Time RT-PCR. Interestingly, nanog and oct-4 (embryonic stem cell markers) expression was significantly greater on fibrin (p0.001) as compared to fibronectin. Conclusions These findings suggest that fibrin it is not only a suitable scaffold for EPC growth and viability but also induces EPC differentiation. The observation that Nanog, known as the most important marker of stemness, is maintained longer than on fibronectin, may offer a surplus value to stem cell-based therapies. (literal)
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  • Comunicazione (literal)
  • Abstract (literal)
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  • (a) Cardiovascular Research Lab.-Cardiac, Thoracic and Vascular Dept.-University of Pisa-Pisa-Italy, (b) Angiology Unit-Cardiac, Thoracic and Vascular Dept.-University of Pisa-Pisa-Italy, (c) Flow Cytometry Section-Immunohaematology II-AOUP-Pisa-Italy, (d) BIOLab-Dept. of Chemistry and Industrial Chemistry-University of Pisa-Pisa-Italy, (e) BIO Labs-Polo S. Anna Valdera-SSSUP S.Anna-Pisa-Italy, (f) Dept. of Chemistry and Industrial Chemistry-University of Pisa-Pisa-Italy, (g) Laboratory for Biomaterials and Graft Technology-Inst. of Clinical Physiology-CNR-Massa-Italy (literal)
Titolo
  • 3-D Fibrin Scaffold Improves Stemness of Human Peripheral Blood Endothelial Progenitor Cells (literal)
Abstract
  • Aims. Fibrin is a natural biopolymer appealing for cell-based regenerative therapies, because it can support growth, migration and differentiation of different cell types. Endothelial progenitor cells (EPC) represent a very interesting alternative cell source for mature endothelial cells; the fact that can easily isolated from the peripheral blood, thereby eliminating donor morbidity, makes them ideal in applications in the field of regenerative medicine.We have demonstrated that fibrin can support EPC viability and growth. Aim of this study was to evaluate if fibrin can affect EPC differentiation and stem cell markers expression. Methods. Fibrin was prepared mixing commercially available (Kedrion S.p.A. Lucca, Italy) fibrinogen (9 mg/ml) and thrombin (25 U/ml). Clot ultrastructure was investigated by scanning electron microscopy (SEM) and cryogenic SEM (CRYO-SEM) to measure fibre diameter and density. Clot elasticity was evaluated by atomic force microscopy (AFM), measuring the tip-sample force by cantilever displacement. EPC were obtained from peripheral blood and cultured on fibrin at the concentration of 1x106cell/cm2. Fibronectin coating was used as a control.Metabolic activity was assessed after 7 and 14 days by WST1 assay and viability by confocal microscopy (calcein incorporation). The expression of both endothelial (CD31, KDR, vWF, Ve-Cadherin) and stem cell markers (nanog, oct-4) was assessed by flow cytometry, confocal microscopy and Real Time RT-PCR. Results. SEM analysis revealed a nanometric fibrous structure, with mean fiber diameter of 165±4 nm and mean density of 95.9±0.2 %. CRYO-SEM suggested a reticulate structure with mesh-size up to 10 ?m. Fibrin clot elasticity was 1.78 MPa, as in literature. WST1 assay showed that fibrin increased EPC metabolic activity as compared to fibronectin (fibrin: 0.606±0.056 a.u. vs. fibronectin: 0.311±0.067). Calcein staining demonstrated that EPC were still viable at 14 days. Flow cytometry showed the expression of endothelial markers (CD31=41.8±8.4%; vWF=32.3±3.0%; KDR=89.3±3.7%; VE-Cadherin=41.2±3.8%), confirmed also by confocal microscopy and Real Time RT-PCR. Interestingly, nanog and oct-4 (embryonic stem cell markers) expression was significantly greater on fibrin (p<0.001) as compared to fibronectin. Conclusions. These findings suggest that fibrin it is not only a suitable scaffold for EPC growth and viability but also induces EPC differentiation. The observation that Nanog, known as the most important marker of stemness, is maintained longer than on fibronectin, may offer a surplus value to stem cell-based therapies. (literal)
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