Fabrication Of Novel Biomaterials Based On Biomimetic Strategies And Their Applications For Dermal Papilla Cell Culture And Hair Follicle Reconstruction

Background and objectiveThe development of regenerative medicine has led to the hope of constructing tissue engineering hair follicles for the treatment of androgenetic alopecia.At present,there are two major challenges in this field:(1)dermal papilla cell(DPC)is one of the fundamental seed cell for hair follicle reconstruction,however,currently there is a lack of cell culture strategy that can bridge the gap between number expansion and the gradually loss of hair-inductive capacity with passage;(2)There is still a lack of hair follicle reconstruction models which can recapitulate the relationship between cells and the microenvironment.To solve these problems,the novel biomaterials are indispensable.In recent years,the emergence of the biomimetic concept has greatly enriched the practicality and functionality of biomaterials.Biomimetic design strategies can be practiced in a variety of ways,such as by directly using the functional components of the target when synthesizing the material,or by adjusting biomaterials to simulate the specific structural or physicalchemical properties of the target tissue.In short,this experiment will be studied from two aspects,(1)modification of electrospun nanofibers by cell membrane coating technology,and to verify its feasibility as a new culture platform for dermal papilla cells;(2)using cell loaded hydrogel microspheres and functional cells as building block to establish a novel hair follicle reconstruction model from the bottom-up.The above research is expected to propose a new comprehensive solution for the research field of hair follicle tissue engineering.Methods1.Cell membrane pellets were derived from mouse DPCs.Biomimetic DPCs culture scaffold were prepared by using electrospinning and cell membrane coaking techniques.Cell membrane wrapping results were identified by contact angle measurement,immunoblotting,and fluorescent expression.The evaluation of biocompatibility of materials and proliferation performance of DPCs was performed using Live and Dead Kit and CCK-8 Kit,respectively.2.Morphological observation of DPCs was performed by SEM and confocal microscope.The effects of intercellular interaction simulation provided by culture model on the biological characteristics of DPCs were investigated by gene expression,protein qualitative and quantitative expression analysis of the hair-inducing markers of DPCs.3.Gelatin methacrylate(GelMA),which is excellent in biocompatibility,is used as a raw material for preparing hydrogel microspheres.First,optimize the diameter of the microspheres,the proportion of microspheres and other parameters.Subsequently,the dermal papilla cells were prepared by hanging drop method.According to the structural characteristics of the hair follicle and its microenvironment in the transwell chamber,the spatial distribution of the carrier microspheres,dermal papilla cells and neonatal mouse epidermal cells was arranged in order.The biomimetic reconstruction model was performed and the in vivo hair follicle reconstruction was verified.Results1.The cell culture model of biomimetic nanofiber scaffolds was successfully constructed by electrospun and cell membrane coating techniques.It was confirmed that the cell culture model has good biocompatibility and supports the proliferation of DPCs.2.High passage dermal papilla cells cultured with the biomimetic scaffolds can expressspeciflic markers ALP,P-catenin and Versican and have the ability to induce hair follicle regeneration in vivo.3.Successfully constructed a large-scale biomimetic hair reconstruction model by using hydrogel microspheres,dermal papilla spheroids and keratinocyte as building blocks and regenerated hair follicle in vivo.Conclusion'1.Cell membrane coated nanofiber scaffold is a new type of DPCs culture platform2.The synergistic interaction between structure and cell surface profile of the scaffold materials can induce the expression of specific markers of non-aggregated DPCs,which may be related to N-cadherin-mediated intercellular interaction pathway.3.The hair reconstruction model assembled via building blocks effectively restore the positional relationship between cells and their environment thus to achieve the goal of saving cells.