Study On Patterned Gelatin-polycaprolactone/silk Fibroin Composite Membranes For Repairing Full-thickness Skin Defects In Rats

Cell-substrate interactions have always been a hotspot in cell biology,and play a pivotal role in the development of biomaterials.Studies have shown that physical properties of substrate,including surface morphology,stiffness,roughness,etc.,will affect vinculin expression and cytoskeleton activity,thus stimulating downstream signaling pathways to modulate cell fate.Recently,micropatterning technology has gradually matured,which could accurately prepare material substrates with specific geometries,making it possible to control cell fate through material surface topology structure.Because large-scale skin wounds regeneration is time-consuming and wounds are prone to recur,resulting in serious infections,it is important to use wound dressings to provide a protective environment,which can accelerate wound regeneration.Therefore,the development of wound dressings that promote rapid healing of skin wounds performs high practical value.In this project,we developed a gelatin-polycaprolactone(PCL)/ silk fibroin(SF)composite membrane with specific micron-level patterns on the surface and used it to treat the full-thickness skin defect in rats.First,we successfully prepared polydimethylsiloxane(PDMS)stamp and gelatin membrane with stripe pattern and grid pattern on the surface through photolithography and soft-etching techniques.Subsequently,we used PCL and SF as raw materials to prepare PCL/SF blended film by electrospinning,and attached it to the unpatterned surface of the gelatin membrane to form a composite membrane.We used in vitro cell experiments to evaluate the effect of micropatterns on cell behavior.Then,we used a rat full-thickness skin defect model with a diameter of 13 mm to evaluate the ability of the micro-patterned gelatin composite membrane to promote skin regeneration.Through optical microscopy,scanning electron microscopy(SEM)and other characterization methods,it was confirmed that the surface of the gelatin layer in composite membrane had highly accurate micropatterns.Additionally,the PCL/SF film and gelatin were successfully closely adhered,which increased the mechanical properties of the composite membrane.In vitro cell experiments showed that the micro-patterned gelatin composite membrane presented good cell adhesion and played an active role in proliferation,which laid a solid foundation for in vivo implantation.In in vivo studies,we evaluated the ability of patterned gelatin composite membranes to promote skin healing by the relative area of the wound,HE staining,Masson staining,and immunofluorescence staining.The results demonstrated that the micro-patterned gelatin-PCL/SF composite membrane could promote the formation of blood vessels at the wound area and promote skin wound healing.Also,the micro-patterned composite membrane could reduce the expression of α-smooth muscle actin(α-SMA)to some extent,thereby reducing the generation of scars.On the other hand,stripe pattern composite membrane exhibited the best effect on skin wound regeneration in comparison to grid pattern and unpatterned composite membrane.