Multilayered Films With Biomimetic Stiffness For Regulation Of Cellular Behaviors

All physiological and pathological processes are accompanied by dynamic remodeling of extracellular microenvironment.Dynamic changes in ECM structure and components lead to changes in matrix biophysical properties including stiffness,which have important effects on a variety of cell behaviors and functions.Traditional biomaterials are fabricated by covalent or non-covalent cross-linking polymer,ceramics.They provide a relatively inert and static microenvironment.Although this kind of research has made great progress on study of interaction between cells and materials,it lacks the spatiotemporal dynamics corresponding to complex cellular processes.The emergence and development of stimulus-responsive materials and bio-responsive materials provide new ideas for simulating dynamic extracellular microenvironment,and each has its own advantages.Aiming at combining the dynamics of stimulus response and biological response into the construction of biomaterials,improving understanding of the relationship between material stiffness and cellular behaviors,achieving better regulation of cells,researches were carried out with details shown in the following four parts:1.A growing body of research confirms that autophagy plays a key role in influencing and controlling cellular behavior.Aiming at the effect of substrate stiffness on the autophagy behavior of vascular endothelial cells(VEC)and vascular smooth muscle cells(VSMC),polyelectrolyte multilayer films with different stiffness were constructed in this study.Cell experiment results showed that the level of autophagy was affected by the stiffness of the material,and the effect was cell type dependent.Further,when autophagic level was down-regulated by addition of autophagy inhibitor,both the secretion amount of VEC functional factors and the phenotypic transformation of VSMC changed accordingly.The study confirmed that the material surface stiffness can directly affect cell autophagy and provided a scientific basis for the enhancement of cell microenvironment stiffness which may accelerate the process of cardiovascular disease.2.A number of studies have shown that high surface stiffness is conducive to the growth of cells in the early stage,while low surface stiffness is conducive to the function of cells in the later stage.Aiming at construction of materials with adaptive stiffness,this study introduced matrix metalloproteinases(MMPs)-sensitive peptide into the multilayer films through click reaction.In the process of cell culture,stiffness of the material can be down-regulated dynamically due to the stimulation of MMPs secreted by cells.The results of VEC cell experiments showed that in the early stage,the material had enough stiffness to support the adhesion and proliferation of VEC;after formation of a complete endothelium,the stiffness decreased with the release of MMPs during EC proliferation.Therefore,the endothelium still maintained sound endothelial function and anticoagulant function on the low low-stiffness material.This study demonstrates the unique advantages of adaptive stiffness and has broad application prospects in tissue engineering,regenerative medicine and organ-on-a-chip.3.There is a close correlation between microenvironment hardness and tumor characters.Aiming at the influence of stiffness decrease on human breast cancer cells MDA-MB-231 tumor malignance,this study investigated the adhesion,proliferation,orientation,protein and gene expression of MDA-MB-231 cultured on films with adaptive stiffness.Cell experiment results showed that compared with the static materials with high stiffnessness,MDA-MB-231 cells cultured on the multilayer film with adaptively decreased stiffness had lower tumor malignancy and showed a certain phenomenon of "tumor dormancy/tumor reverse".4.From the perspective of bionics,it will be beneficial for us to further understand the law of the influence of material stiffness on the behavior of vascular cells to build a material system with the characteristics of physiological and pathological vascular stiffness.This study introduced light sensitive azobenzene(AZO)molecule to the design of multilayer film.Through regulating of assembly solution concentration,layer number and ultraviolet illumination time,materials with physiological/pathological vascular stiffness were fabricated.Under the illumination of ultraviolet/visible light,the stiffness of materials can effectively switch between physiological stiffness and pathological stiffness.Results of VSMC cell culture on the material showed that good cellular compatibility.This study provides a new method for constructing biomaterials that mimic the extracellular microenvironment properties.