Immobilized amines and basic amino acids as mimetic heparin-binding domains for cell surface proteoglycan-mediated adhesion.
Diamines covalently coupled to glass substrates promoted human foreskin fibroblast adhesion in the absence of serum. These diamine-derivatized substrates were produced by coupling ethylene diamine, N-methylaminoethylamine, and N,N-dimethylaminoethylamine (NNDMAEA), to sulfonyl chloride-activated glass. Electron spectroscopy for chemical analysis demonstrated that the diamines were coupled via their primary amine ends to produce a surface-bound secondary amine linked to a free amino moiety via a two-carbon spacer. NNDMAEA-modified substrates containing free tertiary amines supported the highest degree of cell spreading (73 +/- 7% actively spreading cells) and the most extensive cytoskeletal organization. Both the free tertiary and surface-bound secondary amines were shown to be required for cell spreading. Lysine- and arginine-grafted substrates supported cell spreading and cytoskeletal organization similar to that on NNDMAEA-modified substrates. Although some stress fibers were observed within spread cells on these substrates, focal contacts did not form. Heparinase treatment did not inhibit cell attachment or spreading to the diamine-derivatized substrates, however chondroitinase ABC inhibited cell attachment and spreading on all substrates; heparinase inhibited spreading on lysine- and arginine-derivatized substrates to a lesser extent. These results imply that cell attachment to these substrates was mediated primarily by cell surface chondroitin sulfate proteoglycans. This study demonstrates that covalently grafted NNDMAEA, lysine, and arginine can mimic the adhesion-promoting activity of the glycosaminoglycan-binding domains of cell adhesion proteins. This study also demonstrates that the interaction with these proteoglycans depends in a very sensitive manner on the particular structure of the immobilized amine.
Massia
S P
SP
Department of Chemical Engineering, University of Texas, Austin 78712-1062.
Hubbell
J A
JA
D000588:Amines / Q000494:pharmacology*; D000599:Amino Acids, Diamino / Q000494:pharmacology*; D001665:Binding Sites; D002448:Cell Adhesion / Q000187:drug effects / Q000502:physiology*; D002462:Cell Membrane / Q000378:metabolism / Q000648:ultrastructure; D002478:Cells, Cultured; D003513:Cycloheximide / Q000494:pharmacology; D003959:Diamines / Q000494:pharmacology; D005347:Fibroblasts / Q000187:drug effects / Q000502:physiology / Q000648:ultrastructure; D006493:Heparin / Q000378:metabolism*; D008854:Microscopy, Electron; D011509:Proteoglycans / Q000378:metabolism*; D012879:Skin Physiological Phenomena; D013329:Structure-Activity Relationship
D000588:Amines; D000599:Amino Acids, Diamino; D003959:Diamines; D011509:Proteoglycans; D006493:Heparin; D003513:Cycloheximide