Polyesters With Physical Guidance And Biochemical Signals For Peripheral Nerve Regeneration

Large defect gaps(generally>5 mm)in peripheral nerve system(PNS)caused by traffic accident,diseases as well as natural disasters lead to enormous healthcare expenditure and poor life quality of numerous patients,which becomes an increasingly serious challenge.Up to present,the functional recovery of injuried nerves still cannot be completely realized by neither autografts nor artificial nerve guidance conduit(NGC)in all instances of PNS injury because of the diverse clinical complications such as donor site morbidity and nerve site mismatch.It is suggested that specific topographical,chemical and biological cues,which can mimic the beneficial signals from native nerve tissue,should be integrated into the surface of biomaterials platforms.These characteristics are vital to directly regulate Schwann cell(SCs)behaviors,to guide the orientation of axonal fibers,and to construct the microenvironment and ultimately to facilitate the PNS regeneration.Firstly,stripe micropattems having ridges/grooves of 20/20 and 20/40 ?m in width were fabricated on the surface of maleimide-functionalized biodegradable poly(ester carbonate)(P(LLA-MTMC))films by a polydimethylsiloxane mould-pressing method,respectively.The laminin-derived CQAASIKVAV peptide end-capped with a—SH group was then grafted via thiol-ene click reaction at mild conditions to obtain the micropatterned and peptides-grafted films.SCs cultured on these films,especially on the 20/40 ?m film,displayed faster and aligned adhesion,and a larger number of elongated cells with a higher length to width ratio along the stripe direction compared to those on the flat-pep film.The migration rate of SCs was significantly enhanced in parallel to the stripe direction with a large net displacement.The micropatterned and peptides-grafted films,especially the 20/40 ?m film,could similarly promote SCs proliferation and NGF secretion as the peptides-grafted planar film.Moreover,the neurites of PC 12 cells sprouted along the ridges with a longer average length on the micropattemed and peptides-grafted films.The synergistic effect of physical patterns and biological cues was evaluated by taking into account the results of cell adhesion force,immunofluorescence staining of vinculin,fluorescence staining of F-actin and nucleus,and reletive gene expression of neural cadherin(NCAD),neurocan(NCAN)and myelin protein zero(PO).As it is not easy to modify the artificial polyester with physical morphology,a simple method was developed.Stripe micropatterns with ridges/grooves width of 3/3 and 10/10 ?m were respectively fabricated on the surface of PLCL films by the polydimethylsiloxane(PDMS)templates thermo-pressing procedure.Surface entrapment,a facile and convenient method,was adopted to immobilize two different densities of gelatin on the micropatterned PLCL films during the swelling-shrinking process.SCs cultured on the higher density of gelatin being entrapped on micropatterned films,especially on 3/3 ?m film,displayed a larger number of elongated cells with a higher length to width ratio along the stripe direction compared to those on the other films.Moreover,the migration rate of SCs was significantly enhanced in parallel to the stripe direction with a large net displacement.The synergistic role played by physical patterns and bioactive gelatin was further investigated considering the results of cell adhesion force,immunofluorescence staining of vinculin,fluorescence staining of F-actin and nucleus,and genes expression of integrin ?1 Rac1 RhoA and Cdc42.The SCs could form "Bunger bands" after the PNS injury,and collective cells would migrate to fix the gap.The SCs spheres were obtained to mimic the "Bunger bands".Substrates with micropatterns and chemical signals were fabricated to investigate the collective SCs migration and the SCs migration from the spheres.PLCL films were harvested by a solvent evaporation method.Stripe micropattems with ridges/grooves width of 3/3,5/5,10/10 and 30/30 ?m were respectively fabricated on the surface of PLCL films by the polydimethylsiloxane(PDMS)templates thermo-pressing procedure.The graphene oxide(GO)was coated on the surface of PLCL films after aminolysis.On the micropatterned PLCL with GO films,the patterns of a smaller size lead to a larger number of elongated cells with a higher length to width ratio along the stripe direction,and more extensive directional migration of cells.Moreover,the wound healing percentage was about 75%,and the distance that cells migrated from the cell spheres was up to 678?m on 3/3?m-PLCL/GO film.In this study,the alignment,elongation,single cell migration and collective cell migration were controlled by stripe micropatterns with bioactive molecules introduced on the biomaterial surface via imitating the in vivo tissue structure.This study offers some novel thoughts to design tissue engineering materials with organized structures such as tendon,urethra and nerve.