Biopolymer nanofiber membranes are attracting interest as promising biomaterial scaffolds with a remarkable range of structural and functional performances for guided bone regeneration (GBR). In this study, tussah silk nanofiber (TSn) and Bombyx mori silk nanofiber (BSn) membranes were prepared by physical shearing. The diameters of the TSn and BSn membranes were 146.09 ± 63.56 and 120.99 ± 91.32 nm, respectively. TSn showed a Young’s modulus of 3.61 ± 0.64 GPa and a tensile strength of 74.27 ± 5.19 MPa, which were superior to those of BSn, with a Young’s modulus of 0.16 ± 0.03 GPa and a tensile strength of 4.86 ± 0.61 MPa. The potential of TSn and BSn membranes to guide bone regeneration was explored. In vitro, the TSn membrane exhibited significantly higher cell proliferation for MC3T3-E1 cells than the BSn membrane. In a cranial bone defect in a rat model, the TSn and BSn membranes displayed superior bone regeneration compared to the control because the membrane prevented the ingrowth of soft tissue to the defective area. Compared to the BSn membrane, the TSn membrane improved damaged bone regeneration, presumably due to its superior mechanical properties, high osteoconductivity, and increased cell proliferation. The TSn membrane has a bionic structure, excellent mechanical properties, and greater biocompatibility, making it an ideal candidate for GBR.
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