Preparation Of Micropatterned Collagen Films And Their Effect On Corneal Cell Behaviors

Surface topography has ability to regulate cell behaviors,including spreading,proliferation,migration differentiation and so on,playing an important role in tissue regeneration.Abundant topographies appear in basement membrane and stroma of cornea,which are fundamental to corneal function.Therefore,application of surface topography in corneal tissue engineering and studying responses of corneal cells to engineering topography not only helps deepen the understanding of interaction between corneal cells and their growth microenvironment but also benefits for development of corneal equivalent similar with native cornea.A method combined soft lithography and solution casting was used for preparation of microgrooved collagen films with different groove size were prepared.The groove sizes were25,50 and 100?m,respectively.Their physiochemical properties of were characterized using scanning electron microscope,contact angle measurement and so on,and the impacts on morphology and behaviors of corneal epithelial cells and keratocytes was studied.Results showed that the microgrooves increased hydrophilicity of collagen films,decreasing their contact angle from 53°to 30°⁴0°.The ion permeability of collagen films was also promoted with the microgrooves,and the diffusion coefficient of collagen films with 100-?m-wide grooves reached 1.3×10^-6 cm²/s.Both corneal epithelial cells and keratocytes showed significant orientation on microgrooved collagen films,and the percentage of aligned cells was up to 80%on microgrooved collagen films.The microgrooved collagen film could promote directional migration of epithelial cells,and the migration speed increased with decrease of groove width.In addition,the transformation of keratocytes into myofibroblasts were also inhibited on microgrooves,especially when the groove width is 100?m.Collagen films with four kind of micropillars were fabricated,three of which was organized micropillar array and the diameter of micropillars were 2?m,5?m and 10?m.The other was disorganized micropillar array consisted of the three micropillars.Their physiochemical properties and effect on morphology and behavior of corneal epithelial cells and keratocytes were investigated using atomic force microscope and so on.Results showed that the micropillar arrays increased water contact angle and roughness of collagen films.The tensile strength was also increased with the micropillars,and the collagen films with 10-?m-wide micropillar has tensile strength with 1.8 MPa.On micropillared collagen films,corneal epithelial cells and keratocytes tended to shift from a rectangular or diamond shape to a triangle or star shape,and their cell area and circularity also decreased,while the aspect ratio increased.Interestingly,the micropillars with a diameter of 10?m induced alignment of corneal epithelial cells.,and the orientation index reached 40%.With the morphology changes,the adhesion and proliferation of the cells on the micropillared collagen films were significant attenuated.These discoveries shed lights on cell-topography interaction and have reference value for the development of corneal tissue engineering scaffolds.In conclusion,to investigate the interaction between surface topography and corneal cells,collagen films with microgrooves and micropillars was fabricated using micropatterning method,and their physiochemical properties and effects on corneal cell behaviors was measured.These discoveries shed lights on cell-topography interaction and have reference value for the development of corneal tissue engineering scaffolds.