Improved endothelialization of vascular grafts by local release of growth factor from heparinized collagen matrices

M.J.B. Wissink, R. Beernink, A.A. Poot, T. Beugeling, G.H.M. Engbers, W.G. van Aken, J. Feijen

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Abstract

Endothelial cell seeding, a promising method to improve the performance of small-diameter vascular grafts, requires a suitable substrate, e.g. crosslinked collagen. In addition, the growth of seeded endothelial cells can be improved by local release of a heparin-binding protein, basic fibroblast growth factor (bFGF). In this study, the influence of immobilization of heparin to collagen, crosslinked using N-(3-dimethylaminopropyl)-N′-ethylcarbodiimide (EDC) in combination with N-hydroxysuccinimide (NHS), on the binding and release of bFGF was determined. Heparin was immobilized also using EDC and NHS. Furthermore, the effects of the release of bFGF from (heparinized) EDC/NHS-crosslinked collagen on the proliferation of seeded endothelial cells was studied in vitro. Immobilization of increasing amounts of heparin to EDC/NHS-crosslinked collagen (containing 14 free ε-amino groups per 1000 amino acid residues, E/N14C) resulted in binding of increasing amounts of bFGF to the material. Maximal bFGF binding was observed for E/N14C containing 20–30 mg heparin immobilized per gram of collagen which was obtained using a molar ratio of EDC to heparin-carboxylic acid groups of 0.4 for heparin immobilization (E/N14C-H(0.4)). Up to concentrations of 320 ng bFGF/ml, 10% of the added bFGF bound to E/N14C, while binding of bFGF to E/N14C-H(0.4) was 22%. The initial release rate of bFGF bound to E/N14C was much higher compared to bFGF bound to E/N14C-H(0.4): respectively, 30 vs. 2% in the first 6 h. After 10 days, the bFGF release from E/N14C and E/N14C-H(0.4) amounted to 83 vs. 42%, respectively. Binding of increasing amounts of bFGF resulted in increased growth of human umbilical vein endothelial cells (HUVECs) seeded on both E/N14C and E/N14C-H(0.4). Nevertheless, after 6 and 10 days of proliferation cell numbers on E/N14C-H(0.4) where higher than cell numbers on E/N14C, irrespective of the bFGF concentration used for loading of the matrix. It is concluded that heparinized, EDC/NHS-crosslinked collagen is a good synthetic vascular graft coating for in vivo endothelial cell seeding.
Original languageEnglish
Pages (from-to)103-114
JournalJournal of controlled release
Volume64
Issue number1-3
DOIs
Publication statusPublished - 2000

Fingerprint

Fibroblast Growth Factor 2
Blood Vessels
Intercellular Signaling Peptides and Proteins
Collagen
Transplants
Heparin
Endothelial Cells
Immobilization
Fibroblast Growth Factor 10
Cell Count
Human Umbilical Vein Endothelial Cells
Growth
Carboxylic Acids
N-hydroxysuccinimide

Keywords

  • METIS-106555
  • IR-74371
  • Sustained release
  • Vascular grafts
  • Endothelial cell seeding
  • Heparin immobilization
  • Growth factors

Cite this

Wissink, M.J.B. ; Beernink, R. ; Poot, A.A. ; Beugeling, T. ; Engbers, G.H.M. ; van Aken, W.G. ; Feijen, J. / Improved endothelialization of vascular grafts by local release of growth factor from heparinized collagen matrices. In: Journal of controlled release. 2000 ; Vol. 64, No. 1-3. pp. 103-114.
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abstract = "Endothelial cell seeding, a promising method to improve the performance of small-diameter vascular grafts, requires a suitable substrate, e.g. crosslinked collagen. In addition, the growth of seeded endothelial cells can be improved by local release of a heparin-binding protein, basic fibroblast growth factor (bFGF). In this study, the influence of immobilization of heparin to collagen, crosslinked using N-(3-dimethylaminopropyl)-N′-ethylcarbodiimide (EDC) in combination with N-hydroxysuccinimide (NHS), on the binding and release of bFGF was determined. Heparin was immobilized also using EDC and NHS. Furthermore, the effects of the release of bFGF from (heparinized) EDC/NHS-crosslinked collagen on the proliferation of seeded endothelial cells was studied in vitro. Immobilization of increasing amounts of heparin to EDC/NHS-crosslinked collagen (containing 14 free ε-amino groups per 1000 amino acid residues, E/N14C) resulted in binding of increasing amounts of bFGF to the material. Maximal bFGF binding was observed for E/N14C containing 20–30 mg heparin immobilized per gram of collagen which was obtained using a molar ratio of EDC to heparin-carboxylic acid groups of 0.4 for heparin immobilization (E/N14C-H(0.4)). Up to concentrations of 320 ng bFGF/ml, 10{\%} of the added bFGF bound to E/N14C, while binding of bFGF to E/N14C-H(0.4) was 22{\%}. The initial release rate of bFGF bound to E/N14C was much higher compared to bFGF bound to E/N14C-H(0.4): respectively, 30 vs. 2{\%} in the first 6 h. After 10 days, the bFGF release from E/N14C and E/N14C-H(0.4) amounted to 83 vs. 42{\%}, respectively. Binding of increasing amounts of bFGF resulted in increased growth of human umbilical vein endothelial cells (HUVECs) seeded on both E/N14C and E/N14C-H(0.4). Nevertheless, after 6 and 10 days of proliferation cell numbers on E/N14C-H(0.4) where higher than cell numbers on E/N14C, irrespective of the bFGF concentration used for loading of the matrix. It is concluded that heparinized, EDC/NHS-crosslinked collagen is a good synthetic vascular graft coating for in vivo endothelial cell seeding.",
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author = "M.J.B. Wissink and R. Beernink and A.A. Poot and T. Beugeling and G.H.M. Engbers and {van Aken}, W.G. and J. Feijen",
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volume = "64",
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Improved endothelialization of vascular grafts by local release of growth factor from heparinized collagen matrices. / Wissink, M.J.B.; Beernink, R.; Poot, A.A.; Beugeling, T.; Engbers, G.H.M.; van Aken, W.G.; Feijen, J.

In: Journal of controlled release, Vol. 64, No. 1-3, 2000, p. 103-114.

Research output: Contribution to journalArticleAcademicpeer-review

TY - JOUR

T1 - Improved endothelialization of vascular grafts by local release of growth factor from heparinized collagen matrices

AU - Wissink, M.J.B.

AU - Beernink, R.

AU - Poot, A.A.

AU - Beugeling, T.

AU - Engbers, G.H.M.

AU - van Aken, W.G.

AU - Feijen, J.

PY - 2000

Y1 - 2000

N2 - Endothelial cell seeding, a promising method to improve the performance of small-diameter vascular grafts, requires a suitable substrate, e.g. crosslinked collagen. In addition, the growth of seeded endothelial cells can be improved by local release of a heparin-binding protein, basic fibroblast growth factor (bFGF). In this study, the influence of immobilization of heparin to collagen, crosslinked using N-(3-dimethylaminopropyl)-N′-ethylcarbodiimide (EDC) in combination with N-hydroxysuccinimide (NHS), on the binding and release of bFGF was determined. Heparin was immobilized also using EDC and NHS. Furthermore, the effects of the release of bFGF from (heparinized) EDC/NHS-crosslinked collagen on the proliferation of seeded endothelial cells was studied in vitro. Immobilization of increasing amounts of heparin to EDC/NHS-crosslinked collagen (containing 14 free ε-amino groups per 1000 amino acid residues, E/N14C) resulted in binding of increasing amounts of bFGF to the material. Maximal bFGF binding was observed for E/N14C containing 20–30 mg heparin immobilized per gram of collagen which was obtained using a molar ratio of EDC to heparin-carboxylic acid groups of 0.4 for heparin immobilization (E/N14C-H(0.4)). Up to concentrations of 320 ng bFGF/ml, 10% of the added bFGF bound to E/N14C, while binding of bFGF to E/N14C-H(0.4) was 22%. The initial release rate of bFGF bound to E/N14C was much higher compared to bFGF bound to E/N14C-H(0.4): respectively, 30 vs. 2% in the first 6 h. After 10 days, the bFGF release from E/N14C and E/N14C-H(0.4) amounted to 83 vs. 42%, respectively. Binding of increasing amounts of bFGF resulted in increased growth of human umbilical vein endothelial cells (HUVECs) seeded on both E/N14C and E/N14C-H(0.4). Nevertheless, after 6 and 10 days of proliferation cell numbers on E/N14C-H(0.4) where higher than cell numbers on E/N14C, irrespective of the bFGF concentration used for loading of the matrix. It is concluded that heparinized, EDC/NHS-crosslinked collagen is a good synthetic vascular graft coating for in vivo endothelial cell seeding.

AB - Endothelial cell seeding, a promising method to improve the performance of small-diameter vascular grafts, requires a suitable substrate, e.g. crosslinked collagen. In addition, the growth of seeded endothelial cells can be improved by local release of a heparin-binding protein, basic fibroblast growth factor (bFGF). In this study, the influence of immobilization of heparin to collagen, crosslinked using N-(3-dimethylaminopropyl)-N′-ethylcarbodiimide (EDC) in combination with N-hydroxysuccinimide (NHS), on the binding and release of bFGF was determined. Heparin was immobilized also using EDC and NHS. Furthermore, the effects of the release of bFGF from (heparinized) EDC/NHS-crosslinked collagen on the proliferation of seeded endothelial cells was studied in vitro. Immobilization of increasing amounts of heparin to EDC/NHS-crosslinked collagen (containing 14 free ε-amino groups per 1000 amino acid residues, E/N14C) resulted in binding of increasing amounts of bFGF to the material. Maximal bFGF binding was observed for E/N14C containing 20–30 mg heparin immobilized per gram of collagen which was obtained using a molar ratio of EDC to heparin-carboxylic acid groups of 0.4 for heparin immobilization (E/N14C-H(0.4)). Up to concentrations of 320 ng bFGF/ml, 10% of the added bFGF bound to E/N14C, while binding of bFGF to E/N14C-H(0.4) was 22%. The initial release rate of bFGF bound to E/N14C was much higher compared to bFGF bound to E/N14C-H(0.4): respectively, 30 vs. 2% in the first 6 h. After 10 days, the bFGF release from E/N14C and E/N14C-H(0.4) amounted to 83 vs. 42%, respectively. Binding of increasing amounts of bFGF resulted in increased growth of human umbilical vein endothelial cells (HUVECs) seeded on both E/N14C and E/N14C-H(0.4). Nevertheless, after 6 and 10 days of proliferation cell numbers on E/N14C-H(0.4) where higher than cell numbers on E/N14C, irrespective of the bFGF concentration used for loading of the matrix. It is concluded that heparinized, EDC/NHS-crosslinked collagen is a good synthetic vascular graft coating for in vivo endothelial cell seeding.

KW - METIS-106555

KW - IR-74371

KW - Sustained release

KW - Vascular grafts

KW - Endothelial cell seeding

KW - Heparin immobilization

KW - Growth factors

U2 - 10.1016/S0168-3659(99)00145-5

DO - 10.1016/S0168-3659(99)00145-5

M3 - Article

VL - 64

SP - 103

EP - 114

JO - Journal of controlled release

JF - Journal of controlled release

SN - 0168-3659

IS - 1-3

ER -