Fabrication Of Heparin And Fibronectin Micropattern On Tianium Surface And Its Effects On The Behavior Of Platelet And Endothelial Cells

For cardiovascular implants, neointimal hyperplasia and thrombus are the major problems in their application. Promoting endothelial cell adhesion or reducing platelet adhesion through surface modification technology can effectively improve the biocompatibility of the materials, and surface patterning technology has becoming popular.In this paper, microtransfer molding technology (μTM), one method of soft lithography was used on titanium (Ti) surface after pretreatment to fabricate bionic biological molecules micropatterns. The pre-modified of Ti surface including NaOH alkali activation and3-aminopropyl triethoxysilane (APTE) treatment, and its surface characteristics observed by scanning electron microscopy (SEM), contact angle measurement, infrared spectrum analysis (FTIR) and amino quantitative method. Polydimethylsiloxane (PDMS) was prepared by the casting method, and then used it to imprint a-Methoxy-poly(ethylene glycol)-ω-succinimidyl carbonate (mPEG-SC) on the pre-modified Ti surface, which could resist proteins and cells ahhesion. After that the mixture of heparin and fibronectin, which have the properties of anticoagulation and promotion of endothelial cell adhesion, respectively, was assembled to fabricate the micropattern of mPEG-SC/Hep-Fn. SEM, contact angle measurement, FTIR, Hep and Fn staining method were used to examine the surface morphology and chemical composition of the mPEG-SC/Hep-Fn micropattem. On this basis, platelet adhesion and endothelial cell culture experiments in vitro were carried out to study the effect of mPEG-SC/Hep-Fn micropattem on the behavior of platelets and endothelial cells.SEM, contact angle, FTIR, light microscopy and fluorescence microscopy results showed that: on alkali activated titanium surface, the roughness and surface hydrophilic were increased,-OH absorption peak was detected. On silane coupling Ti surface, the roughness was further increased, hydrophobicity was increased, CH2absorption peak was detected and the amino density of NH2was about51nmol/cn2. Clear stripes could be observed on the micropattem sample surface. FTIR showed the absorption peaks of mPEG-SC, Hep and Fn. And the Hep and Fn staining further confirmed the biological inertness of mPEG-SC, Hep and Fn was only distributed in the regions not coated with mPEG-SC.Platelet adhesion experiments showed that:on Ti and Hep-Fn surface, platelets covered the entire surface with extended pseudopods, but did not aggregation. As the anticoagulant effect of heparin, the numbers of platelets adherent on Hep-Fn surface was less than Ti. On mPEG-SC surface, there was almost no platelet adhesion; on micropaterned sample surface, platelet selective distributed on Hep-Fn stripe region, further reduce platelet adhesion amount compared to nonpatterned Hep-Fn surface. Different number of platelet adhesion due to different sizes of Hep-Fn on micropatterned sample. The amount and distribution of activated platelets were similar with the results of total platelet adhesion. APTT results showed that both nonpatterned and micropatterned Hep-Fn did not significant prolongation of APTT time, but they also did not aggravate clotting time, which may had anticoagulant effect.EC adhesion results showed that: nonpatterned Hep-Fn surface could promote EC adhesion, spreading and proliferation compared to titanium. The mPEG-SC surface almost had no endothelial cell adhesion. ECs distributed along with the Hep-Fn direction on the patterned sample surfaces. The different sizes of micropatterns had different effects on the orientation angle, ratio of length to width as well as shape index of EC:there could be two or more ECs parallel arranged on the micropatterns with greater size than EC, which could increase cell spreading and proliferation; and there could only be a single cell distribution on the micropatterns with similar size to EC, which could elongate and has guiding function to EC.The above results suggest that:the biological functions of mPEG-SC/Hep-Fn micropattern could regulate the distribution of platelet and endothelial cell, at the same time reduce platelet adhesion and regulate the adhesion, growth and proliferation of EC. And this may effectively improve the biocompatibility of Ti.