Regulation Of Silk Nanofiber Structure And The Influence On Stemness Maintenance Of Stem Cells

Stem cell research has become the core of regenerative medicine and the main obstacle to such research lies in the uncontrollable cell behavior.Smart biomaterials are introduced to address this obstacle.Silk-based materials can be made into a variety of material types with tunable mechanical properties,nanotopography and drug-delivering properties.Therefore,silk-based materials become an ideal candidate to regulate stem cell behavior,such as differentiation,proliferation and sternness maintenance.However,uncontrollable structure and unclear self-assembly mechanism of silk fibroins greatly hinders the preparation and design of materials.Also,integrating multiple signals to mimic the microenvironment of stem cells in vitro makes it difficult to design the structure of silk-based materials.To overcome the limitation,silk nanofibers with more controllable structure prepared by our lab were regulated using physical and chemical methods to gain further insight on the relationships between structure and properties.Moreover,silk-based materials with specific structure were developed to dig their value in sternness maintenance of stem cells.Firstly,we systematically studied the influences of silk fibroin structure on chemical modification and a strategy was provided to improve traditional chemical modification.Silk nanofiber hydrogels with alignment structure(SA)were then prepared in the electric field through mimicking microenvironment of cell mass in blastocyst.These bionic hydrogels integrating multiple signals could maintain sternness of embryonic stem cells(ESCs)in the long term without exogenous leukemia inhibitory factor(LIF)and the mechanism was also discussed.Finally,based on the sternness maintenance results of silk nanofiber hydrogels,we continued to explore the influences of silk fibroin structures on various stem cells.Detailed research contents are as follows:(1)The effect of silk fibroin structure on chemical modification:The structure(conformation and aggregates)of silk fibroin was proved to influence the degree of grafting and reaction rate based on phenol groups of silk fibroin.Amorphous silk short nanofiber(ASS)was found to be a better candidate to achieve more controlled chemical modifications of silk materials after comfirming the influences of changes in nanostructures and conformations on distribution of functional groups and the universality of various functional modifications.Finally,ASS together with stable nanostructure and controllable transformations provided a promising strategy for designing bifunctional materials through various chemical modifications.(2)Regulation of silk nanofiber structure through physical methods and the influence of silk nanofiber structure on sternness of stem cells:SA was prepared in the electric field through mimicking microenvironment of cell mass in blastocyst and its influence on embryonic stem cell behavior was discussed.Results from immunofluorescence analysis,RT-qPCR,differentiation ability in vitro and in vivo confirmed that SA could maintain sternness of mouse ESCs(mESCs)without LIF.The possible mechanism of the effect was also discussed from the perspective of cell adhesion and cell autocrine behavior.Finally,silk nanofiber hydrogels with different structures were prepared-due to the controllable structure.The effects of changes in structures on sternness maintenance of embryonic stem cells,adipose-derived stem cells and bone marrow mesenchymal stem cells were preliminarily studied,providing a reference for the design of materials and the active regulation of different stem cell behaviors.