Construction Of Aligned Silk Fibroin Nanofiber Hydrogels For The Manipulation Of Cell Behavior

The interaction between cells and extracellular environment has a decisive effect on cell behavior.The cellular microenvironment includes interface structure,mechanical properties,chemical properties,etc.Active design of multiple factors will achieve desired cell behaviors.Here,silk fibroin nanofibers as basic elements were used to form hydrogel with oriented structures under an AC electric field.More regulating signals such as graphene and gradient structure were then introduced to optimize the control of cell behaviors.Firstly,silk fibroin hydrogels with tunable orientation structure was derived from silk fibroin nanofiber solutions with different concentrations,providing a suitable platform for evaluating the influence of oriented structure on cell behaviors.Besides better cell growth and proliferation,in vitro cell cultures also showed oriented cell morphology and preferred migration along the orientation direction.The exfoliated graphene nanosheet as another factor was then introduced in the aligned hydrogel system.The graphene nanosheet was exfoliated using silk nanofiber as a stabilizer and directly added to silk nanofiber solutions for hydrogel formation.The structure,mechanical properties,orientation morphology and conductive properties of the hydrogels were investigated.Neural related cells including Schwann cells,PC 12 cells and embryonic stem cells were cultured on these hydrogels to assess the effects of different factors.The results indicated that the oriented composite gels was beneficial to the adhesion and proliferation of Schwann cells and PC 12 cells,and promoted the growth and extension of Schwann cytoskeleton and PC 12 cell neurites along the orientation direction.The addition of graphene enhanced the differentiation capacity of stem cells into neurons and increased the proportion and growth of synapses formed by PC 12 cells.Finally,the gradient signal was introduced into the oriented hydrogel.Mechanical gradients,drug concentration gradients,and growth factor concentration gradients formed in the oriented hydrogels for better control cell behavior.The introduction of gradient signal didn't influence the regulation of orientation structure on cell behavior.Further active control of different cell behaviors was achieved after the gradient structure formation.The gradient mechanical signals maintained the orientation morphology of cells.The increased inhibition of cancer cell proliferation appeared on the anticancer drug-loaded hydrogels following the gradient increase of drug density.Changeable differentiation behaviors also happened for the cells cultured on the BMP-2 loaded hydrogels along the density gradient direction.The above results suggested the possibility of studying the synergistic effects of different signals through the active design of gradient signals on the aligned hydrogels.In summary,our present study provides a feasible strategy of developing aligned hydrogels containing multiple regulatory factors.Better cell behavior control was achieved on these novel hydrogels.Further insight into the relation of cell and biomaterials opens up new possibility of designing bioactive biomaterials with optimized microenvironment.