Silk fibroin (SF) has garnered significant interest in tissue engineering due to its excellent biocompatibility and bioactivity. Notably, SF exhibits piezoelectric properties that can be harnessed to electrically stimulate cells, enhancing tissue morphogenesis. However, a systematic approach to control SF's piezoelectricity and biodegradation, and its application in neural morphogenesis, has not been fully explored. In this study, SF was electrospun into nanofibers of various sizes and subjected to a post-electrospinning chemical treatment to examine the effects of phase composition, especially the β-sheet formation, on the piezoelectricity and biostability of SF. The optimized SF scaffolds having a nanofiber diameter of 750 nm and a scaffold thickness of 250 µm allowed for the maintenance of nanofibrous structure for at least 6 weeks in aqueous environments while maintaining piezoelectric potential outputs of at least 200 mVp-p. This enabled cell membrane depolarization over an extended cell culture duration, facilitating the functional maturation of human neural stem cells (hNSCs). Specifically, acoustic piezo-activation of the SF scaffolds, resulting in mechano-electrical stimulation (MES) of the cells, accelerated their differentiation into neurons, astrocytes, and especially oligodendrocytes, leading to robust axon myelination within two weeks. Furthermore, MES enhanced neural connectivity and functional maturity, as evidenced by significant increases in excitatory and inhibitory synapse formation (46 % and 58 %, respectively), action potential amplitude (252 %), and velocity (210 %) compared to static control conditions. These findings demonstrate the potential of biodegradable electrospun SF nanofibers with balanced piezoelectricity and biostability for advancing neural tissue engineering.
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