Introduction: Cell adhesion on a biomaterial surface is crucial for the long-term success of a titanium implant. Here, a novel concept of combining highly stable and affine titanium adhesive properties with specific cell binding moieties into one molecule is described. Furthermore anti-fouling units and heparin binding segments are included that allow the design of multifunctional surface modification.
Materials and Methods: Titanium oxide surfaces were used for coating. Multifunctional peptides were synthesized by solid phase peptide synthesis including different click chemistry approaches for modification. Immobilization was analyzed by fluorescence methods. Cell adhesion, spreading, viability and proliferation were tested with MG 63 osteoblast cells.
Results and Discussion: A modular peptide containing L-3,4-dihydroxyphenylalanine (DOPA), which was identified as surface binding unit of the mussel foot protein mfp1, for immobilization on titanium surfaces. Short polyethylene glycol (PEG) units were introduced to achieve antifouling properties. Further modification with a cyclic RGD and a heparin binding peptide (HBP) was realized by an efficient on-resin combination of Diels-Alder reaction with inverse electron demand (DARinv) and Cu(I) catalyzed alkyne-azide cycloaddition (CuAAC). Conjugating the cyclic RGD and HBP in one peptide led to a synergistically improved spreading, proliferation, viability and the formation of well-developed actin cytoskeleton and focal contacts of osteoblast-like cells. To additionally modify the coating, different sulfated glycosaminoglycans (GAG), and subsequently the chemokine CXCL12 were added. Heparin showed strong and specific CXCL12 binding. Release experiments showed the typical initial burst with a flattened, but sustained release of the protein after the first 2 days.
Conclusion: Our results clearly demonstrate that multifunctional peptide coating of surfaces is a novel approach to combine different requirements of biomaterials into one specific surface coating. Short peptide sequences containing integrin and heparin binding motifs can efficiently enhance cell surface interactions on titanium and synergistically improve cell spreading and viability. Specific release of chemokines allows a full biomimetic approach that imitates the extracellular membrane.
German Science Foundation, TRR 67 A4
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