Silk Scaffolds With Controllable Nanostructure And Secondary Structure

Silk-based scaffolds have been transformed in just the past decade from thecommodity textile world to a growing web of applications in tissue engineering due to theimpressive biocompatibility, biodegradability, minimal inflammatory reactions andexcellent mechanical properties. Although silk-based scaffolds have promising applicationsin tissue repairs, a challenge for scaffold fabrication remains to further improve silkbiocompatibility and inductivity for different tissue regeneration needs. The extracellularmatrix (ECM) provides a framework for scaffold design in which nanoscale andmicroscale dimensions of the physical structures are important, including nanofibrousarchitectures as well as specific porous structures. The nanofibrous structure in ECM ispropitious to the suitable micro-environment for cell culture and tissue regeneration as wellas to achieve high porosity that facilitates cell proliferation and seeding.In the present study, silk fibroin with different nanostructures was firstlyself-assembled in aqueous solution and then used to regulate the porous structure offreeze-dried scaffolds. Viscosity, secondary structure and water interactions were alsostudied to elucidate their influence on the formation and control of porous structures. It isfound that the nanostructure of silk fibroin in aqueous solution is a critical factor inregulating the final features of these porous structures.On the basis of the study, we developed a mild self-assembly approach to preparesilk nanofilaments, and then nanofibrous scaffold having similar nano-fibrillar structurewith extracellular matrix (ECM). Pores of200~250μm diameter and>99%porositywere found in the scaffold. These soluble porous scaffolds were then treated with water ormethanol annealing to achieve water stability and different degradation behaviors to meetthe requirements of different tissue regenerations. It is found that silk fibroin transformedfrom random to Silk I after water-annealing treatment. With the increase of methanolcontent, the crystal structure of silk shifted from Silk I to Silk II, resulting in the improvement of thermal stability as well as degradation stability. Following the dissolutionor degradation of silk fibroin, the nanofibrous structure gruadally emerged, which mightpromote cell adhesion, growth and migration in vitro or in vivo.Finally, in order to clarify the influence of different structures of the scaffolds on cellgrowth and proliferation, human bone marrow mesenchymal stromal cells (BMSC) werecultured, using salt-leaching scaffolds as control. The laser confocal microscope and DNAcontent assay results showed that BMSCs grew much better in the ECM-mimickingnanofibrous scaffolds than in salt-leached scaffolds.Therefore, the study provided an effective way to prepare silk scaffold withcontrollable nanostructures and secondary structures, which would provide suitablebioactive matrices for different tissue engineering or tissue repair applications.