The addition of a polyglutamate domain to the angiogenic QK peptide improves peptide coupling to bone graft materials leading to enhanced endothelial cell activation.

Abstract

Vascularization of bone grafts is vital for graft integration and bone repair, however non-autologous graft sources have limited potential to induce angiogenesis. Accordingly, intensive research has focused on functionalizing non-autologous materials with angiogenic factors. In the current study we evaluated a method for coupling an angiogenic peptide to the surface of two clinically-relevant graft materials, anorganic bovine bone (ABB) and synthetic hydroxyapatite (HA). Specifically, the VEGF-derived "QK" peptide was synthesized with a heptaglutamate (E7) domain, a motif that has strong affinity for calcium phosphate graft materials. Compared with unmodified QK, a 4-6 fold enrichment was observed in the binding of E7-modified QK (E7-QK) to ABB and HA. The E7-QK peptide was then assessed for its capacity to stimulate angiogenic cell behaviors. Human umbilical vein endothelial cells (HUVECs) were treated with solutions of either QK or E7-QK, and it was found that QK and E7-QK elicited equivalent levels of cell migration, tubule formation and activation of the Akt and ERK signaling pathways. These data confirmed that the inherent bioactivity of the QK sequence was not diminished by the addition of the E7 domain. We further verified that the activity of E7-QK was retained following peptide binding to the graft surface. HA disks were coated with QK or E7-QK, and then HUVECs were seeded onto the disks. Consistent with the increased amount of E7-QK bound to HA, relative to QK, markedly greater activation of Akt and ERK 1/2 was observed in cells exposed to the E7-QK-coated disks. Taken together, these results suggest that the E7 domain can be leveraged to concentrate angiogenic peptides on graft materials, facilitating delivery of higher peptide concentrations within the graft site. The ability to endow diverse graft materials with angiogenic potential holds promise for augmenting the regenerative capacity of non-autologous bone grafts.