Cell adhesion ability of artificial extracellular matrix proteins containing a long repetitive Arg-Gly-Asp sequence

We generated recombinant artificial extracellular matrix (ECM) proteins containing repetitive Arg-Gly-Asp (RGD) sequences: double (RGD2), 21 (RGD21) and 43 (RGD43) repeats of RGD. RGD43-coated glass slides promoted fibroblast NIH3T3 cell adhesion and spreading on the surface. Since actin stress fibers and focal contacts were also observed in cells adhering on RGD43-coated glass slides, it was suggested that the RGD peptides in RGD43 transmitted an adhesion signal via integrins and promoted cell adhesion. We coated recombinant ECM proteins, each containing repetitive RGD domains, on polystyrene plates and investigated the effects of RGD length on the cell adhesion ability using three different cell lines, namely, fibroblast NIH3T3, HeLa cancer and neuronal PC12 cell lines. The results indicated that RGD43 had a cell adhesion ability superior to those of natural extracellular matrix proteins, fibronectin and laminin, although the effects of RGD repeat length on the cell adhesion ability depended on the cell line. As an artificial three-dimensional scaffold for cell cultivation, we also prepared an RGD43 hydrogel by a cross-linking reaction using glutaraldehyde. On the RGD43 hydrogel scaffold, fibroblast cells also successfully adhered under serum-free conditions.