Basic Research On Phosphonated Gelatin And Dopa Conjugated Epidermal Growth Factor For Surface Modification

Titanium and titanium alloys are widely used in medical applications such as the replacement of hard tissues including bone and joints. However, there is still a need to further investigate their biocompatibility including the interface between titanium and the biological tissue. Because of a lack of bonding of implants to juxtaposed tissues, current orthopedic implants have a variety of problems including infection, extensive inflammation, and overall poor osseointegration. Therefore, many attempts have been made to modify the surface of titanium with functional or biological components to induce tissue responses to biomaterials and provide a set of powerful signals for cell growth and differentiation. Gelatin can be used for surface modification, but the gelation process occurs at room temperature in gelatin solution. And the adhesion affinity is low between gelatin and the titanium surface. To solve this problem, we intend to take advantage of 3-aminopropylphosphonic acid modified gelatin to obtain adhesion function of gelatin(Phosphonated gelatin, P-gelatin), to better fixed to the titanium surface, to improve the cell compatibility of the titanium surface.How to make the growth factors work better on cell growth, proliferation or differentiation is another hotspot in tissue engineering. In comparison with soluble growth factors, the immobilized growth factors have a longer action time, therefore, the total using amount can be reduced. On the other hand, the concentration regulation and control would be more simple and convenient. In this study, we intend to use the DOPA to design some adhering polypeptide, and introduce to the epidermal growth factor(Epidermal growth factor, EGF) to obtain DOPA conjugated epidermal growth factor(EGF-DOPA). On collagen membrane, the influence of EGF-DOPA for the proliferation of fibroblasts NIH3T3 mouse embryo was reached..Phosphonated gelatin was prepared for surface modification of titanium to stimulate cell functions. The modified gelatin was synthesized by coupling with 3-aminopropylphosphonic acid using water-soluble carbodiimide and characterized by 31P-nuclear magnetic resonance. Circular dichroism revealed no differences in theconformations of unmodified and phosphonated gelatin. However, the gelation temperature was changed by the modification. Even a high concentration of modified gelatin did not form a gel at room temperature. Time-of-flight secondary ion mass spectrometry showed direct bonding between the phosphonated gelatin and titanium surface after binding. The binding behavior of phosphonated gelatin on the titanium surface was quantitatively analyzed by a quartz crystal microbalance. Ellipsometry showed the formation of a several nanometer layer of gelatin on the surface. Contact angle measurement indicated that the modified titanium surface was hydrophobic. Enhancement of the attachment and spreading of MC-3T3L1 osteoblastic cells was observed on the phosphonated gelatin-modified titanium. These effects on cell adhesion also led to growth enhancement.An epidermal growth factor(EGF) derivative with affinity for biomaterial surface was designed using a peptide moiety inspired from mussel adhesive protein. Since the active sequence has some 3,4-dihydroxyphenylalanine(DOPA) residues, the EGF derivative was prepared by organic synthesis, and a 59 residue peptide was successfully prepared using this method. The binding affinity of the modified EGF to collagen membrane was significantly higher than the unmodified EGF. The catechol groups in the affinity sequence contributed to the affinity of modified EGF to the material. Immobilized EGF, engineered to have binding affinity for biomaterial surface, promoted cell growth more efficiently than soluble EGF through prolonged activation of cell signal transduction.Phosphonated gelatin and dopa conjucated epidermal growth factor were successfully preparedfor biomaterial surface modification. These two methods for providing new functional biomaterial that may be suitable for surface modification and the development of medical devices.