Construction Of Highly Ordered Porous Hydroxyapatite Micropattern And Its Regulation On Cell Growth Behavior

Micropattern is an interesting product of nature.A variety of micro/nano ordered micropatterns in nature give the living body unique biological functions.Inspired by nature,Micropatterning technology can be used to precisely design nanoscale or microscale topology of material surface,allowing the cells to be placed in defined locations and to promote cell alignment for exhibiting specific functions.These artificial micro/nano structures can give biomaterials excellent mechanical properties and many unique biomimetic functions,which already has become a core research topic and direction in the field of biomaterials and tissue engineering.The development of micropatterning technology could provide an effective structural template for investigating the cell response mechanism to the growth microenvironment,and also provide an effective experimental method for precise control of cell growth behavior.In this dissertation,combined with lithography technology,microsphere self-assembly technology,electrochemical deposition technology and high temperature calcination technology,a hydroxyapatite(HAp)micropattern coating with multi-level ordered porous structure was constructed on the surface of titanium substrate.The adsorption behavior of bovine serum albumin(BSA)on the HAp micropattern surface was qualitatively analyzed.The adhesion morphology,adhesion region,proliferation and directional migration behavior of pre-osteoblasts(MC3T3-E1)on the micropattern surface were studied in depth.And the intrinsic relationship between the micro-level structure of the coating and its biological properties are discussed.It provides important scientific basis and technical support for the application in the fields of basic cell biology,tissue engineering,and biosensors.The main results of this dissertation are outlined as following:(1)A easy method for fabricating a complex multi-level micropattern structure of biomaterial was proposed,and a HAp micropattern coating with a highly ordered porous structure was controllably constructed on a titanium metal substrate.The microsphere interface coating technology was used to construct the ordered structure of large-sized microspheres(10 μm,50 μm),and the fine microstructure of the HAp micropattern coating was effectively adjusted by controlling the size of the microspheres.(2)The effects of each steps in the preparation process on the microstructure,chemical composition and crystal structure of the samples were investigated systematically.The key factors for the successful construction of highly ordered multi-level structures of HAp micropatterns were studied.The OCP micropattem is converted to a HAp micropattern while the photoresist/microsphere template is removed by high temperature(600℃)calcination treatment.(3)The results of in vitro protein adsorption test indicate that BSA can be preferentially adsorbed in the HAp microdomain to form a BSA micropattern which consistent with the shape of the micropatterned substrate.The ribbon-like HAp crystal structure of the prepared micropattern sample can control the early stage of BSA adsorption behavior(0～7 Days).And the multi-level fine microstructure of samples M10/HAp and M50/HAp can immobilize BSA in the HAp micro-region for up to 21 days.The construction of multi-level fine microstructure can significantly enhance controlling protein adsorption behavior.(4)The results of in vitro cell culture experiment indicate that the prepared HAp micropattern samples can effectively promote cell proliferation,which possess good cell compatibility.The HAp micropattern can promote cell growth in the direction of the microstripe at the initial stage of cell adhesion.The construction of the multi-level ordered porous structure can significantly enhance the long-term guidance and control of cell elongation and orientation.The cells can maintain a high degree of orientation and stretch on the surface of the sample M50/HAp for up to 21 days.(5)The results of the wound healing model experiment indicate that HAp striped micropattern can promote the directional migration of cells along the stripe direction.The ordered porous fine structure constructed in the HAp micro-region can enhance the guiding and controlling effects on the directional migration behavior of cells.The cell directional migration rate on the M50/HAp sample is up to 500 μm/d,which is more than four times that of the Ti substrate.It’s worth noting that the healing of the wound model on the M50/HAp sample can be completed in 3 days.