Using Mixed Self-Assembled Monolayers Presenting RGD and (EG)₃OH Groups To Characterize Long-Term Attachment of Bovine Capillary Endothelial Cells to Surfaces

Abstract

This paper describes surfaces that promote the ligand-directed binding of cells and resist the cellular deposition of adhesive proteins. These surfaces are based on self-assembled monolayers (SAMs) of alkanethiolates on gold that present mixtures of arginine-glycine-aspartate (RGD), a tripeptide that promotes cell adhesion by binding to cell surface integrin receptors, and oligo(ethyleneglycol) moieties, groups that resist nonbiospecific adsorption of proteins and cells. Surface plasmon resonance (SPR) spectroscopy was used to measure the adsorption of carbonic anhydrase and fibrinogen to mixed SAMs comprising RGD groups ((EG)6OGRGD) and tri(ethylene glycol) groups ((EG)3OH); SAMs having values of the mole fraction of RGD (χRGD) ≤ 0.05 adsorbed nearly undetectable levels of carbonic anhydrase or fibrinogen. Bovine capillary endothelial cells attached and spread on SAMs at χRGD ≥ 0.00001, with spreading of cells reaching a maximum at χRGD ≥ 0.001. These mixed SAMs reduced the deposition of proteins by attached cells relative to both fibronectin adsorbed on SAMs of hexadecanethiolate on gold and RGD peptide coated on glass. After allowing cells to attach for 2 or 4 h to any of these surfaces presenting RGD groups, addition of soluble GRGDSP to the medium contacting the adherent cells rapidly released them from the surfaces. However, if cells were allowed to attach to surfaces for 24 h, only those cells attached to the mixed SAM presenting (EG)6OGRGD and (EG)3OH groups could be released using the soluble GRGDSP at a rate comparable to cells attached to fibronectin for 2 h. These results demonstrate that RGD alone is sufficient for adhesion and survival of cells over 24 h.

Biology on a Chip: Microfabrication for Studying the Behavior of Cultured Cells

“…235 A range of techniques have been used to micropattern endothelial cells. [236][237][238][239][240][241][242][243] Chen et al 244 found that cell spreading acted as a regulator of proliferation or death in human and bovine capillary endothelial (CE) cells by tightly controlling cell spreading with adhesive microislands of fibronectin; fibronectin was adsorbed on a methyl-terminated alkanethiol SAM, whereas the nonadhesive background consisted of a hexaethylene-glycolterminated alkanethiol SAM (HEG-SAM) that resisted protein adsorption and cell attachment. Cells that were free to spread over large microislands displayed growth, as expected, but the cells that were constrained to limit their spreading to a small island displayed apoptosis.…”

Attachment of hyaluronan to metallic surfaces

“…Cell integrins are transmembrane proteins on the cell surface that adhere to the proteins of the ECM such as collagen, fibronectin, or vitronectin. These and other adhesive proteins contain short peptide sequences such as RGD, REDV, YIGSR, PHSRN, KNEED, and PRRARV, [37][38][39][40][41][42][43][44][45][46][47][48][49] which are bound by the integrins to promote cell adhesion, spreading, and proliferation. Although significant work has been done to attach proteins and peptides to metal oxide particles 21 and silicon oxide on Si wafers, 26 relatively little research has been reported on attachment of polypeptides to macroscopic metallic surfaces.…”

Protein Adhesion on SAM Coated Semiconductor Wafers: Hydrophobic versus Hydrophilic Surfaces

“…Grafted polymers and self-assembled monolayers are methods used to introduce functionality to surfaces, as well as establishing a packing density on the surface and specific orientation of the functionality. Polyethylene glycol (PEG), or polyethyleneoxide (PEO), functionalized surfaces are examples of hydrophilic coatings that offer excellent resistance to protein and cellular adhesion as both grafted polymer and SAM films (19,20,21,22,23,24). In contrast, hydrophobic surfaces, such as octadecyltrichlorosilane SAMs, tend to adsorb proteins as a monolayer thick biofilm (25,26,27).…”