Positively and Negatively Modulating Cell Adhesion to Type I Collagen Via Peptide Grafting

Abstract

The biophysical interactions between cells and type I collagen are controlled by the level of cell adhesion, which is dictated primarily by the density of ligands on collagen and the density of integrin receptors on cells. The native adhesivity of collagen was modulated by covalently grafting glycine–arginine–glycine–aspartic acid– serine (GRGDS), which includes the bioactive RGD sequence, or glycine–arginine–aspartic acid–glycine–serine (GRDGS), which includes the scrambled RDG sequence, to collagen with the hetero-bifunctional coupling agent 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide. The peptide-grafted collagen self-assembled into a fibrillar gel with negligible changes in gel structure and rheology. Rat dermal fibroblasts (RDFs) and human smooth muscle cells demonstrated increased levels of adhesion on gels prepared from RGD-grafted collagen, and decreased levels of adhesion on RDG-grafted collagen. Both cell types demonstrated an increased ability to compact freefloating RGD-grafted collagen gels, and an impaired ability to compact RDG-grafted gels. RDF migration on and within collagen was increased with RDG-grafted collagen and decreased with RGD-grafted collagen, and dose–response experiments indicated a biphasic response of RDF migration to adhesion. Smooth muscle cells demonstrated similar, though not statistically significant, trends. The ability to both positively and negatively modulate cell adhesion to collagen increases the versatility of this natural biomaterial for regenerative therapies.

Porous and Nonporous Nerve Conduits: The Effects of a Hydrogel Luminal Filler With and Without a Neurite-Promoting Moiety

“…This percentage equates to 130-160 mg of coupled peptide per milliliter collagen matrix, based on the molar ratio of peptide to monomeric type-I bovine collagen fibers. 36 These values are similar to the effective doses administered in solution in previous in vivo nerve regeneration studies. 29,30 Collagen only and functionalized collagen solutions were injected into Conduits either had a porous outer wall denoted by ''P'' or a nonporous outer wall denoted by ''NP.''…”

“…32 For the functionalized collagen, the peptide mimetic of the m-HNK 33 (FLHTRLFV, MW: 1032.24, synthesized by GenScript) was grafted onto the collagen using EDC chemistry, as described. 32,36 This peptide has been shown to encourage nerve regeneration and axonal targeting. 30,31 Peptide grafting efficiency was measured indirectly by grafting FITC-tagged m-HNK (GenScript) to collagen and comparing the fluorescence intensity of grafted collagen after reconstitution to a standard curve created by admixing fluorescent peptide into collagen solution.…”

Neural cell type-specific responses to glycomimetic functionalized collagen

“…Peptide sequences were covalently conjugated onto oligomeric type I collagen using the heterobifunctional crosslinker 1-Ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) as previously described [24]. Briefly, fetal calf type I collagen (EPC, Owensville, MO) was reconstituted to 3 mg/mL in 0.02N acetic acid to make an oligomeric collagen solution.…”

“…In a previous study using this conjugation technique, we confirmed that the resulting grafting efficiency is between 50–60%. Additionally, the grafting process does not significantly change mechanical properties, fiber diameter, or fiber density of the hydrogel [24]. …”

Osteogenic molecules for clinical applications: improving the BMP-collagen system

“…For the functionalization of titanium implants with RGD-peptides strategies such as simple adsorption (Song et al, 2010) or chemical linking through gold-thiol chemistry (Ferris et al, 1999) or electrodeposited PEG (Oya et al, 2009) have been carried out. In the case of collagen type I, functionalization with this type of biomimetic peptide has been done through chemical procedures such as using thiol or phosphonate anchors (Roessler et al, 2001), periodate activation (Zhang et al, 2005) or hetero-bifunctional coupling agents (Monteiro et al, 2011). To avoid these chemical modifi cations to functionalize ACSs with biomimetic peptides, one approach has been the design of an RGD peptide with the CBD from the vWF (Fig.…”