Effect Of Bead-on-string And Shish-kebab Structures On Fibroblast Behavior Under Dynamic/Static Conditions

Scar/fibrosis is a common complication of wound healing and tissue injury that imposes a severephysical,psychological and economic burden on patients.The development of scar/fibrosis is due to persistence and functionality of inflammatory cells and fibroblastmyofibroblast differentiation beyond the normal range.Fibroblasts are the main cells of extracellular matrix(ECM)production and maintenance,which can be transformed into myofibroblasts with strong contractility,affecting collagen deposition and wound healing process and quality.It is therefore particularly important to understand the effects of various biochemical and physical signals on fibroblast behavior during repair.Electrospinning is a simple and effective micro/nano fiber preparation technique.In addition,cells are subjected to various mechanical stimuli in vivo,such as stretching and compression,so introducing appropriate mechanical stimuli for cell-implant culture in vitro can better simulate the natural microenvironment in vivo.Based on this,poly-caprolactone(PCL)fiber scaffolds with different surface topologies were prepared by electrospinning combined with solution crystallization method to study their effects on fibroblast adhesion,proliferation and differentiation,under dynamic/static loading.Then,the rationality of in vitro experiments and in vivo induced regenerative effects scaffolds of different surface topologies were verified through in vivo animal tests,which provided a reference for the design of fiber scaffolds to accelerate tissue healing and reduce scarring.The main studies and conclusions are as follows.(1)Preparation of fiber scaffolds with different surface topologies.Smooth and beaded structured fibers,P12 and P6,were prepared by adjusting the electrospinning parameters,and shish-kebab structured(SK)and beaded shish-kebab structured(BSK)fiber scaffolds with different kebab densities were prepared by solution crystallization method.With the increase of PCL quality fractions in crystallization dilute solution,the crystallinity,roughness,fracture strength and elastic modulus of SK and BSK increase.In particular,BSK improved the mechanical properties of the P6 defective crystalline structure,and the elastic modulus of BSK with proper kebab density(P6-SK1)and overdense(P6-SK2)were two and three times higher than that of P6(1.04±0.22MPa),respectively.(2)The effects of different surface topologies on fibroblast proliferation and differentiation under in dynamic/static culture conditions were explored.Live/dead staining and cell proliferation results showed that all scaffolds have good biocompatibility.Immunofluorescence staining results also showed that different surface topologies scaffolds possess certain effect on protein/gene expression.P6-SK1 and proper kebab density(P12-SK1)could decrease integrin ?1 expression and have lower transforming growth factor-?1(TGF-?1)and ?-smooth muscle actin(?-SMA)expression as well as reasonable collagen deposition.(3)A rat abdominal subcutaneous embedding test was constructed to verify the vitro test correlation and the in vivo induced regeneration of the fiber scaffolds.The results of H?E staining,Masson staining and q PCR showed that all samples had good histocompatibility,no obvious inflammation,and show good inward growth.Compared with P6 and P12,the inflammatory cell layer of SK and BSK fiber scaffolds is thin,and ECM deposition is less.Especially,P6-SK1 and P12-SK1 barely have inflammatory cells,loose collagen tissue,and low expression of TGF-?1 and ?-SMA,indicating that P6-SK1 and P12-SK1 have certain effects on wound healing and scar inhibition.In summary,electrospun fiber scaffolds with different topologies were prepared,which have significant effects in regulating fibroblast behavior,such as proliferation,adhesion and differentiation.Moreover,the dynamic culture conditions can better simulate the microenvironment in vivo,and provide guidance for the screening of more suitable implants in vivo.At the same time,it also provides guidance and reference for the study of the mechanism of structure-mechanics-cell and subsequent research on reducing scar and accelerating wound healing in tissue/organ regeneration and repair.