Controllable fabrication of silk fibroin porous scaffolds and their regulation on cellular behaviours

With the continuous advancement of biomechanics and cell biology, the importance of substrate materials in regulating cell growth, movement, differentiation, apoptosis, gene expression, adhesion, and signal transduction has been increasingly recognized. Silk fibroin (SF) porous scaffolds, owing to their excellent biocompatibility, controllable biodegradability, and ability to effectively simulate the <i>in vivo</i> microenvironment, have been demonstrated to possess broad application prospects in the field of tissue engineering. However, traditional preparation methods for SF porous scaffolds have been found to exhibit poor control over pore size and mechanical properties, and a trade-off between pore size and mechanical performance has often been observed, which has limited their practical application to some extent. A method termed "alcohol addition-freezing method" for preparing SF porous scaffolds was previously developed by our research group, and herein, this method was further extended by adjusting three parameters: the concentration of SF, the concentration of the denaturant <i>n</i>-butanol, and the freezing temperature. Through this approach, controllable preparation of SF porous scaffolds was successfully achieved, resulting in a series of scaffolds with varying pore sizes and compressive moduli. Notably, unidirectional regulation of scaffold pore size and mechanical properties was accomplished, meaning that scaffolds with the same pore size could be designed to exhibit different mechanical properties, and <i>vice versa</i>. Based on this, macrophages, fibroblasts, and bone marrow mesenchymal stem cells (BMSCs), which are frequently involved in tissue engineering scaffold research, were selected to investigate the effects of scaffold pore size and stiffness (represented by compressive modulus) on their biological behaviors. <i>In vitro</i> cell experiments demonstrated that these cells exhibit different biological response in those SF scaffolds with different pore size and stiffness. In summary, the preparation method for SF scaffolds employed in this study has not only addressed the limitations of traditional methods in unidirectionally regulating the physical properties of SF porous scaffolds but has also provided a novel strategy and approach for controlling the microenvironment of cell growth in regenerative medicine, which is considered to hold significant scientific and practical value.