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Print this pageVEGF-C Enhances Fibroblast Dependent EPC Migration
Stephen M Bauer1, Zhao-Jun Liu2, Meenhard Herlyn2, Omaida C Velazquez1
1Hospital of the Univeristy of Pennsylvania, Philadelphia, PA;2Wistar Institute, Philadelphia, PA Vasculogenesis, the de novo formation of vascular networks, is dependent upon bone marrow derived endothelial progenitor cells (EPCs) migration into the extracellular matrix (ECM). Generally, cellular migration into the ECM relies on interacting with ECM components, resident cells, and cytokines. Our objectives were to (1) determine if fibroblasts are necessary for EPC migration into ECM and (2) if endothelial growth factor-C (VEGF-C) enhances this effect. We employed a three-dimensional model whereby bone marrow derived mononuclear cells (BMMCs) cultured as a monolayer on fibronectin for five days are overlaid with two layers of type I collagen (CN). The first layer (1mm-thick) consisted of acellular CN. The second layer (3mm-thick) was either another acellular CN layer or cellular CN layer with embedded dermal fibroblasts. BMMCs were obtained from LacZ-Tie-2 transgenic mice in which every endothelial lineage cell expresses beta-galactosidase (beta-gal). Fibroblasts were untransduced (no vector) or transduced with VEGF-C/Ad5 at 100-plaque forming units/ cell. Collagen gels were harvested after seven days. EPCs were quantified by beta-gal staining in serial cross sections. Quantification was performed in 10 random high power fields (HPF); results are expressed as percent beta-gal positive cells of the total nucleii; data is presented as means +/- SEM; two-tail t-test was used for statistical analysis. Acellular CN only did not support EPC migration into the gels. There was significant migration of EPCs into the CN in the fibroblast-embedded gels. When fibroblasts were introduced with the gene to over-express VEGF-C, there was significantly enhanced EPC migration into the CN compared to untransduced fibroblasts (Figure 1). Fibroblasts are necessary for EPC migration into gels composed of collagen type I, in this in vitro model. The over expression of VEGF-C enhances this effect demonstrating VEGF-C's role as a vasculogenic cytokine. This data offers insight into cellular and molecular mechanisms for vasculogenesis.
1Hospital of the Univeristy of Pennsylvania, Philadelphia, PA;2Wistar Institute, Philadelphia, PA Vasculogenesis, the de novo formation of vascular networks, is dependent upon bone marrow derived endothelial progenitor cells (EPCs) migration into the extracellular matrix (ECM). Generally, cellular migration into the ECM relies on interacting with ECM components, resident cells, and cytokines. Our objectives were to (1) determine if fibroblasts are necessary for EPC migration into ECM and (2) if endothelial growth factor-C (VEGF-C) enhances this effect. We employed a three-dimensional model whereby bone marrow derived mononuclear cells (BMMCs) cultured as a monolayer on fibronectin for five days are overlaid with two layers of type I collagen (CN). The first layer (1mm-thick) consisted of acellular CN. The second layer (3mm-thick) was either another acellular CN layer or cellular CN layer with embedded dermal fibroblasts. BMMCs were obtained from LacZ-Tie-2 transgenic mice in which every endothelial lineage cell expresses beta-galactosidase (beta-gal). Fibroblasts were untransduced (no vector) or transduced with VEGF-C/Ad5 at 100-plaque forming units/ cell. Collagen gels were harvested after seven days. EPCs were quantified by beta-gal staining in serial cross sections. Quantification was performed in 10 random high power fields (HPF); results are expressed as percent beta-gal positive cells of the total nucleii; data is presented as means +/- SEM; two-tail t-test was used for statistical analysis. Acellular CN only did not support EPC migration into the gels. There was significant migration of EPCs into the CN in the fibroblast-embedded gels. When fibroblasts were introduced with the gene to over-express VEGF-C, there was significantly enhanced EPC migration into the CN compared to untransduced fibroblasts (Figure 1). Fibroblasts are necessary for EPC migration into gels composed of collagen type I, in this in vitro model. The over expression of VEGF-C enhances this effect demonstrating VEGF-C's role as a vasculogenic cytokine. This data offers insight into cellular and molecular mechanisms for vasculogenesis.
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