title: Magnetic beads (Dynabead) toxicity to endothelial cells at high bead concentration: implication for tissue engineering of vascular prosthesis.
abstract: Magnetic beads (Dynabeads) have been used for the purification of endothelial cells. One application for this procedure may be for single-stage seeding of bypass grafts. The number of endothelial cells (EC) isolated is crucial and therefore to increase the number of cells extracted, a higher number of Dynabeads per cell may need to be used. The effect of large numbers of CD31 Dynabeads on cell proliferation/metabolism is unknown. We undertook this study using CD31-coated Dynabeads and EC from human umbilical vein. EC were coated at concentrations of 4, 10, or 50 beads per cell. The cells were cultured for 6 days with control being normal EC. Cellular proliferation was assessed by trypsinization of cells and metabolism assessed with an Alamar blue viability assay. In a further experiment a compliant polyurethane graft was single-stage seeded with both coated Dynabeads and normal EC. The results showed that using a higher number of beads per cell resulted in a reduction in cell proliferation and a reduction in cell metabolism. The total number of Dynabeads-coated cells in culture compared to controls (%) by day 6 were 30.7 +/- 2.56, 41.3 +/- 9.8 and 59.2 +/- 7.3 for 50, 10, and 4 beads per cell, respectively. The corresponding results for Alamar blue were 43.7 +/- 1.2, 61.8 +/- 1.4, and 72.1 +/- 4.3. The seeded grafts showed reduced metabolism with the Dynabeads-coated EC. In conclusion, high numbers of beads per cell have a late detrimental effect on cell proliferation and metabolism. Therefore for single-stage seeding lower numbers of Dynabeads will need to be used with resultant reduction in the number of available EC.
authors.affiliation: Tissue Engineering Centre, University Department of Surgery, Royal Free and University College Medical School, University College London, Royal Free Hospital, London, United Kingdom.
authors.forename: A
authors.initials: A
authors.lastname: Tiwari
authors.forename: G
authors.initials: G
authors.lastname: Punshon
authors.forename: A
authors.initials: A
authors.lastname: Kidane
authors.forename: G
authors.initials: G
authors.lastname: Hamilton
authors.forename: A M
authors.initials: AM
authors.lastname: Seifalian
mesh_terms: D001672:Biocompatible Materials; D001807:Blood Vessel Prosthesis; D002455:Cell Division; D002478:Cells, Cultured; D004396:Coloring Agents / Q000494:pharmacology; D064420:Drug-Related Side Effects and Adverse Reactions; D042783:Endothelial Cells / Q000166:cytology*; D004730:Endothelium, Vascular / Q000166:cytology; D006801:Humans; D018189:Immunomagnetic Separation / Q000379:methods*; D008280:Magnetics; D008855:Microscopy, Electron, Scanning; D010078:Oxazines; D019408:Platelet Endothelial Cell Adhesion Molecule-1 / Q000096:biosynthesis; D011140:Polyurethanes / Q000737:chemistry; D023822:Tissue Engineering / Q000379:methods*; D014471:Umbilical Veins / Q000166:cytology; D014966:Xanthenes
chemical_list: D001672:Biocompatible Materials; D004396:Coloring Agents; D010078:Oxazines; D019408:Platelet Endothelial Cell Adhesion Molecule-1; D011140:Polyurethanes; D014966:Xanthenes; C005843:resazurin