Construction of Micro/nano Structures,DNA-functionalized Coatings on Biomedical Titanium Surface and Their Biological Response

Author:Jiang Pin Liang

Supervisor:lin chang jian ren lei wang yong


Degree Year:2017





Titanium(Ti)and its alloys have been widely used as hard tissue replacements including artificial bones,joints and dental implants,due to the favorable chemical stability,biocompatibility and mechanical properties.However,the mechanically polished Ti surface is not able to facilitate integration with bone tissue,and it is generally encapsulated by fibrous tissue after placement into the living body,leading to the failure of implantation.In order to improve its biocompatibility,bioactivity and corrosion resistance,it is necessary to modify the structure and composition on Tisurface to enable suitable biological properties.Natural bone is a hierarchical hybrid organization assembled from nanostructure,microstructure and macrostructure.From a biomimetic viewpoint,a hierarchical hybrid structure composed of micro-and nanoscale components may achieve a balanced promotion in multiple cell(stem cell or osteoblast)functions,including adhesion,proliferation,differentiation and mineralization.Recently,many researchers have attempted to prepare hierarchical micro/nanostructures,but in some cases,either a reduced proliferation or a decreased differentiation was observed on the micro/nanostructures.Accordingly,it is essential to optimize the surface structure,composition,crystalline phase and wettability to enhance the bioactivity of the micro/nanostructures.The base-pairing rules endow DNA with specific and precise programmability.The titanium surface modified with DNA strand helps realize the assembly of drug-loading nanogels,and the serum degradation triggers the release of nanogels as well as drug to regulate cell activities.Additionally,the aptamer can bind its target molecules like growth factors with high affinity and specificity,making it a promising candidate for modifying the Ti or mice bone surfaces to achieve the controlled release of growth factors(e.g.VEGF).The controlled release of VEGF is expected to promote angiogenesis and osteogenesis of bone replacements.The thesis focused on the development of functionalized Ti surfaces.Firstly,the octacalcium phosphate(OCP)or hydroxyapatite(HA)was deposited onto the micro/nanostructured TiO2 surface by electrochemical deposition or spin-coating method to prepare a CaP-deposited micro/nanostructured surface.The effects of surface properties,such as structure,composition,crystalline phase and wettability,on the preosteoblast(MC3T3-E1)response were systematically investigated.Secondly,based on the DNA hybridization,it is able to repeatedly realize the assembly and release of drug-loading nanogels and Dox to regulate the growth of smooth muscle cells.Thirdly,the Ti or bone modified with aptamer-functionalized hydrogel helps achieve a controlled release of VEGF.The main results are outlined as follows:1.The micro/nanostructured TiO2 surface was obtained by a combined treatment of acid-etching,anodization and high-temperature calcination.The OCP coating was deposited onto the micro/nanostructured TiO2 surface via electrodeposition.The OCP-deposited micro/nanostructured surface displayed superior corrosion resistance and apatite-forming ability compared to the polished Ti surface and the micro/nanostructured surface without OCP deposition.The biological evaluations also showed that such an OCP-deposited micro/nanostructured surface significantly improved multiple cell functions,such as adhesion,spread,proliferation,differentiation and extracellular matrix mineralization.2.The micro/nanostructured TiO2 surface with an amorphous phase was obtained by a combined treatment of acid-etching and anodization,and it was crystallized into an anatase or a mixed anatase/rutile phase at the calcination temperature of 450 ℃or 550 ℃.The HA was deposited onto the micro/nanostructured TiO2 surface with various crystalline phases via a spin-coating method.All the samples with/without high-temperature calcination or HA deposition were superhydrophilic and superoleophilic.According to the biological evaluations,the TiO2 surface with a mixed anatase/rutile phase exhibited much better proliferation of MC3T3-E1 cells compared to that of the surface with an anatase or amorphous phase,and the amorphous surface showed the inferior cell proliferation.Interestingly,after HA modifications,the existence of HA did not affect the cell proliferation on the surfaces with different crystalline phases,but it obviously improved the cell differentiation and extracellular matrix mineralization.All in all,it is obvious that the HA-deposited micro/nanostructured surface with a mixed anatase/rutile phase obtained the best bioactivity(e.g.cell proliferation,differentiation and extracellular matrix mineralization)among all the samples.3.The polyacrylamide hydrogel with single-stranded DNA(chemically modified cODNi)was prepared on the Ti surface radical polymerization,and the hydrogel surface can bind ODN1-functionalized nanogels by hybridization reaction.It is noteworthy that the nanogels can be released from the hydrogel surface in 5%FBS solution because of the serum degradation of ODNi,whereas the well-preserved cODNi in the hydrogel surface was able to bind nanogels again and the release function was repeatedly achieved.The Dox can be intercalated into a DNA duplex(ODN2 and cODN2)on nanogels to form Dox-loaded nanogels.Hence,the hydrogel surface can repeatedly release the Dox to regulate the smooth muscle cell growth when Dox-loaded nanogels was used.4.The microporous hydrogel composed of gelatin and PEG was prepared on the Ti surface by using PMMA spheres as templates,and such a microporous hydrogel layer facilitated the adhesion and proliferation of endothelial cell(HUVECs)and preosteoblast(MC3T3-E1).In order to obtain the controlled release of VEGF,the anti-VEGF aptamer was incorporated into the hydrogel.However,according to the release profile,the microporous hydrogel cannot retain VEGF well and there was an initial burst release.Based on the above results,another gas formation method was used to prepare an aptamer-functionalized superporous hydrogel on mice bone.It is showed that the aptamer-functionalized superporous hydrogel can obviously decrease the burst release of VEGF.Additionally,the serum did not have a great influence on the short-term release kinetics of VEGF when the bone with superporous hydrogel immersed in 10%FBS medium,indicating that the aptamer-functionalized superporous hydrogel is expected to promote the bioactivity of bone allografts.