Bioavailability of immobilized epidermal growth factor: Covalent versus noncovalent grafting.
In an effort to rationalize and optimize an antiapoptotic coating combining chondroitin sulfate (CS) and epidermal growth factor (EGF) for vascular applications, the authors here report the comparison of two grafting strategies aiming to display EGF in an oriented fashion on CS. For that purpose, the authors produced, purified, and characterized a chimeric protein corresponding to EGF that was N-terminally fused to a cysteine and a coil peptide. The chimera was covalently immobilized via its free thiol group or captured via coiled-coil interactions at the surface of a biosensor or on a chondroitin sulfate coating in multiwell plates, mimicking the coating that was previously developed by them for stent-graft surfaces. The interactions of grafted EGF with the soluble domain of its receptor or the impact of grafted EGF upon vascular smooth muscle survival in proapoptotic conditions indicated that the coiled-coil based tethering was the best approach to display EGF. These results, combined to direct enzyme-linked immunosorbent assay measurements, indicated that the coiled-coil tethering approach allowed increasing the amount of bioavailable EGF when compared to covalent coupling, rather than the total amount of grafted EGF, while using much lower concentrations of tagged EGF during incubation.
Department of Chemical Engineering, Groupe de Recherche en Sciences et Technologies Biomédicales (GRSTB), Bio-PResearch Unit, École Polytechnique de Montréal, P.O. Box 6079, succ. Centre-Ville, Montreal, Quebec H3C 3A7, Canada.
Nesrine
N
Riahi
Department of Chemical Engineering, Groupe de Recherche en Sciences et Technologies Biomédicales (GRSTB), Bio-PResearch Unit, École Polytechnique de Montréal, P.O. Box 6079, succ. Centre-Ville, Montreal, Quebec H3C 3A7, Canada.
Frederic
F
Murschel
Department of Mechanical Engineering, Ecole de technologie supérieure, Montreal, Quebec H3C 1K3 & Research Centre, Centre Hospitalier de l'Université de Montreal (CRCHUM), Montreal, Quebec H2X 0A9, Canada.
Sophie
S
Lerouge
Life Sciences|NRC Human Health Therapeutics Portfolio, Building Montreal-Royalmount, National Research Council Canada, Montreal, Quebec H4P 2R2, Canada.
Yves
Y
Durocher
Department of Chemical Engineering, Groupe de Recherche en Sciences et Technologies Biomédicales (GRSTB), Bio-PResearch Unit, École Polytechnique de Montréal, P.O. Box 6079, succ. Centre-Ville, Montreal, Quebec H3C 3A7, Canada.
Olivier
O
Henry
Department of Chemical Engineering, Groupe de Recherche en Sciences et Technologies Biomédicales (GRSTB), Bio-PResearch Unit, École Polytechnique de Montréal, P.O. Box 6079, succ. Centre-Ville, Montreal, Quebec H3C 3A7, Canada.
Gregory
G
De Crescenzo
D000818:Animals; D001682:Biological Availability; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D002809:Chondroitin Sulfates / Q000378:metabolism*; D004815:Epidermal Growth Factor / Q000378:metabolism* / Q000493:pharmacokinetics*; D006801:Humans; D055767:Immobilized Proteins / Q000378:metabolism* / Q000493:pharmacokinetics*; D032389:Myocytes, Smooth Muscle / Q000187:drug effects / Q000502:physiology; D011485:Protein Binding; D051381:Rats
D055767:Immobilized Proteins; D004815:Epidermal Growth Factor; D002809:Chondroitin Sulfates