TY - JOUR
T1 - Reduction of non-endothelial cell contamination of microvascular endothelial cell seeded grafts decreases thrombogenicity and intimal hyperplasia
AU - Arts, C. H.P.
AU - Blankensteijn, J. D.
AU - Heijnen-Snyder, G. J.
AU - Verhagen, H. J.M.
AU - Joosten, P. Ph A.Hedeman
AU - Sixma, J. J.
AU - Eikelboom, B. C.
AU - De Groot, P. G.
PY - 2002/5
Y1 - 2002/5
N2 - Introduction: fat derived microvascular endothelial cells (MVEC) seeded on prosthetic vascular grafts, improve patency in animals. Results in humans were disappointing, due to thrombogenicity and progressive intimal hyperplasia. Also in animals intimal hyperplasia was found. We postulate that contaminating cells present in the transplant are involved in the intimal hyperplasia. We developed a method to further purify human MVEC from 40-90%. Here we tested the effects of enrichment upon thrombogenicity and seeding-related intimal hyperplasia. Methods: liposuction fat was enzymatically digested and centrifuged. To enrich MVEC, contaminating macrophages and fibroblasts were removed with dynabeads coated with macrophage- and fibroblast-specific antibodies. Thrombogenicity was assessed by measuring tissue factor and thrombomodulin activity, presence of endothelial nitric oxide synthase and via perfusion of the cells with whole blood. To investigate seeding-related intimal hyperplasia, PTFE grafts were seeded with the cells and cultured for 3 weeks. Results: tissue factor activity of purified cells was reduced compared to nonpurified cells. Purified cells showed thrombomodulin activity and eNOS expression. Fragment 1+2 and Fibrinopeptide A generation after perfusion of purified cells were significantly lower than after perfusion of nonpurified cells, and only nonpurified cells were covered with platelets and fibrin. Prostheses seeded with nonpurified cells showed an EC monolayer above a multilayer of myofibroblasts, prostheses seeded with purified cells only showed a single EC monolayer. Mixing experiments with human umbilical cord EC (HUVEC) and fibroblasts showed that when more than 25% HUVEC were present a confluent EC layer was formed. When the amount of fibroblasts was 25% or less, no development of a subendothelial multilayer of myofibroblasts was found within 3 weeks. Conclusion: reduction of non-endothelial cell contamination of microvascular endothelial cell seeded grafts decreases thrombogenicity and might prevent seeding-related intimal hyperplasia.
AB - Introduction: fat derived microvascular endothelial cells (MVEC) seeded on prosthetic vascular grafts, improve patency in animals. Results in humans were disappointing, due to thrombogenicity and progressive intimal hyperplasia. Also in animals intimal hyperplasia was found. We postulate that contaminating cells present in the transplant are involved in the intimal hyperplasia. We developed a method to further purify human MVEC from 40-90%. Here we tested the effects of enrichment upon thrombogenicity and seeding-related intimal hyperplasia. Methods: liposuction fat was enzymatically digested and centrifuged. To enrich MVEC, contaminating macrophages and fibroblasts were removed with dynabeads coated with macrophage- and fibroblast-specific antibodies. Thrombogenicity was assessed by measuring tissue factor and thrombomodulin activity, presence of endothelial nitric oxide synthase and via perfusion of the cells with whole blood. To investigate seeding-related intimal hyperplasia, PTFE grafts were seeded with the cells and cultured for 3 weeks. Results: tissue factor activity of purified cells was reduced compared to nonpurified cells. Purified cells showed thrombomodulin activity and eNOS expression. Fragment 1+2 and Fibrinopeptide A generation after perfusion of purified cells were significantly lower than after perfusion of nonpurified cells, and only nonpurified cells were covered with platelets and fibrin. Prostheses seeded with nonpurified cells showed an EC monolayer above a multilayer of myofibroblasts, prostheses seeded with purified cells only showed a single EC monolayer. Mixing experiments with human umbilical cord EC (HUVEC) and fibroblasts showed that when more than 25% HUVEC were present a confluent EC layer was formed. When the amount of fibroblasts was 25% or less, no development of a subendothelial multilayer of myofibroblasts was found within 3 weeks. Conclusion: reduction of non-endothelial cell contamination of microvascular endothelial cell seeded grafts decreases thrombogenicity and might prevent seeding-related intimal hyperplasia.
UR - http://www.scopus.com/inward/record.url?scp=0036584429&partnerID=8YFLogxK
U2 - 10.1053/ejvs.2002.1604
DO - 10.1053/ejvs.2002.1604
M3 - Article
C2 - 12027467
AN - SCOPUS:0036584429
SN - 1078-5884
VL - 23
SP - 404
EP - 412
JO - European Journal of Vascular and Endovascular Surgery
JF - European Journal of Vascular and Endovascular Surgery
IS - 5
ER -