Leibniz Research Laboratory for Biotechnology and Artificial Organs

Hannover, Germany

Leibniz Research Laboratory for Biotechnology and Artificial Organs

Hannover, Germany
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Bar A.,Leibniz Research Laboratory for Biotechnology and Artificial Organs | Dorfman S.E.,Leibniz Research Laboratory for Biotechnology and Artificial Organs | Fischer P.,Hannover Medical School | Hilfiker-Kleiner D.,Hannover Medical School | And 5 more authors.
Cardiovascular Research | Year: 2010

AimsA problem in generating artificial tissues is supplying nutrients to cells within 3D constructs. The use of a decellularized biological vascularized matrix with preserved pedicles (BioVaM), as a scaffold, appears to aid the generation of perfusable tissue constructs in vitro. To prevent vessel occlusion upon implantation, a functional endothelium must line the graft vessel bed. Here we tested whether the pro-angiogenic factor CCN1 could improve the re-endothelialization of BioVaM in vitro.Methods and resultsBioVaM vessel beds were incubated with 100 ng/mL recombinant human CCN1. Human cord blood endothelial cells (hCBEC) were analysed with respect to adhesion behaviour upon CCN1 exposure and seeded onto vessel structures of CCN1 exposed BioVaM (cBioVaM). BioVaMs were fixed in a bioreactor and perfusion cultured for 4 and 14 days (d). BioVaM without CCN1 treatment served as controls. Initial seeding success and endothelialization progression were monitored by fluorescencelabelled hCBEC. During construct cultivation, pH and lactate production were measured. Degree of endothelialization and characterization of seeded cells, with respect to endothelial markers, were investigated histologically. BioVaM vessel structures showed a 78 ± 17 increase of attached cells when pre-treated with CCN1. Evaluation of re-endothelialization (arbitrary units) was 4.0 ± 0.8 and 2.6 ± 0.8 after 4 d, and 5.0 ± 0.0 and 3.0 ± 0.5 after 14 d in cBioVaM vs. BioVaM, respectively. On day 14, lactate concentration, an indicator of metabolic activity, was increased 12-fold in cBioVaM relative to BioVaM. A preserved endothelial phenotype of seeded cells was verified in all cultures by acetylated low density lipoprotein uptake and positive immunohistochemistry against von Willebrand factor, endothelial nitric oxide synthase, and CD31.ConclusionCoating of decellularized vessel structures with CCN1 supports adhesion of hCBEC and enhances re-endothelialization of BioVaM. Perfusable, endothelialized constructs may aid in solving the problem of nourishing cells inside 3D tissueengineered constructs.


PubMed | Leibniz Research Laboratory for Biotechnology and Artificial Organs
Type: Journal Article | Journal: Cardiovascular research | Year: 2010

A problem in generating artificial tissues is supplying nutrients to cells within 3D constructs. The use of a decellularized biological vascularized matrix with preserved pedicles (BioVaM), as a scaffold, appears to aid the generation of perfusable tissue constructs in vitro. To prevent vessel occlusion upon implantation, a functional endothelium must line the graft vessel bed. Here we tested whether the pro-angiogenic factor CCN1 could improve the re-endothelialization of BioVaM in vitro.BioVaM vessel beds were incubated with 100 ng/mL recombinant human CCN1. Human cord blood endothelial cells (hCBEC) were analysed with respect to adhesion behaviour upon CCN1 exposure and seeded onto vessel structures of CCN1 exposed BioVaM (cBioVaM). BioVaMs were fixed in a bioreactor and perfusion cultured for 4 and 14 days (d). BioVaM without CCN1 treatment served as controls. Initial seeding success and endothelialization progression were monitored by fluorescence-labelled hCBEC. During construct cultivation, pH and lactate production were measured. Degree of endothelialization and characterization of seeded cells, with respect to endothelial markers, were investigated histologically. BioVaM vessel structures showed a 78 +/- 17% increase of attached cells when pre-treated with CCN1. Evaluation of re-endothelialization (arbitrary units) was 4.0 +/- 0.8 and 2.6 +/- 0.8 after 4 d, and 5.0 +/- 0.0 and 3.0 +/- 0.5 after 14 d in cBioVaM vs. BioVaM, respectively. On day 14, lactate concentration, an indicator of metabolic activity, was increased 12-fold in cBioVaM relative to BioVaM. A preserved endothelial phenotype of seeded cells was verified in all cultures by acetylated low density lipoprotein uptake and positive immunohistochemistry against von Willebrand factor, endothelial nitric oxide synthase, and CD31.Coating of decellularized vessel structures with CCN1 supports adhesion of hCBEC and enhances re-endothelialization of BioVaM. Perfusable, endothelialized constructs may aid in solving the problem of nourishing cells inside 3D tissue-engineered constructs.

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