Adhesion of MC3T3-E1 cells to RGD peptides of different flanking residues: detachment strength and correlation with...

We synthesized a series of RGD peptides and immobilized them to an amine-functional self-assembled monolayer using a modified maleimide-based conjugate technique that minimizes nonspecific interactions. Using a spinning disc apparatus, a trend in the detachment strength (tau(50)) of RGD peptides of different flanking residues was found: RGDSPK > RGDSVVYGLR approximately RGDS > RGES. Using blocking monoclonal antibodies, cellular adhesion to the peptides was shown to be primarily alpha(v)-integrin-mediated. In contrast, the tau(50) value of the cells on fibronectin (Fn)-coated substrates of similar surface density was 6-7 times higher and involved both alpha(5)beta(1) and alpha(v)beta(3) integrins. Cellular spreading was enhanced on RGD peptides after 1 h when compared to RGE and unmodified substrates. However, no significant differences were observed between the different RGD peptides. Long-term function of MC3T3-E1 cells was also evaluated by measuring alkaline phosphatase (ALP) activity and mineral deposition. Among the four peptides, RGDSPK exhibited the highest level of ALP activity after 11 days and mineralization after 15 days and reached comparable levels as Fn substrates after 15 and 24 days, respectively. These findings collectively illustrate both the advantages and limitations of enhancing cellular adhesion and function by the design of RGD peptides.