Enzyme-responsive release of Insulin-like Growth Factor-1 from recombinant spider-silk protein nanofibrous membrane...

The application of biomaterial scaffolds capable of controllable growth factor release plays a pivotal role in nerve tissue repair. In this study, a recombinant fusion protein RSSP@IGF was designed and fabricated, yielding a novel recombinant spider silk protein nanofiber membrane. By integrating insulin-like growth factor-1 (IGF-1) with recombinant spider silk protein (RSSP) and incorporating specific cleavage sites responsive to thrombin and TEV protease, the construct enabled the controlled and sustained delivery of IGF-1. It was also expressed and purified using the Escherichia coli system to obtain a purity exceeding 90% RSSP@IGF. Subsequently, uniform nanofiber membranes were prepared by electrospinning. The membrane delivered excellent biocompatibility, with a hemolytic activity of 0.3%, and could significantly promote the growth of PC12 cells. Its degradation kinetics could be precisely adjusted by thrombin concentration, and 20-90% degradation achieved within 30 min at concentrations ranging from 0.1 to 4.0 U/mL. The IGF-1 release profile triggered by TEV protease showed that following its initial rapid release phase (0-24 h), its biological activity level could be maintained for up to 120 h. Notably, compared with all control groups (tissue culture plate, RSSP alone, and RSSP with soluble IGF-1), the membrane significantly enhanced PC12 cell proliferation within 10 days, demonstrating the advantage of the local delivery system. Mechanism studies revealed that RSSP@IGF may promote the proliferation of PC12 cells via the PI3K/Akt/mTOR signaling pathway. In summary, the controllable degradation spider silk protein nanofiber membrane constructed in this article provides a highly biocompatible and efficient platform for the sustained delivery of IGF-1. Its controllable degradation and sustained release characteristics hold enormous potential for applications in nerve regeneration and other fields warranting precise spatiotemporal control of growth factor presentation.