Photoactive Poly(3-hexylthiophene) Nanoweb for Optoelectrical Stimulation to Enhance Neurogenesis of Human Stem Cells

Optoelectrical manipulation has recently gained attention for cellular engineering; however, few material platforms can be used to efficiently regulate stem cell behaviors via optoelectrical stimulation. In this study, we developed nanoweb substrates composed of photoactive polymer poly(3-hexylthiophene) (P3HT) to enhance the neurogenesis of human fetal neural stem cells (hfNSCs) through photo-induced electrical stimulation. Methods: The photoactive nanoweb substrates were fabricated by self-assembled one-dimensional (1D) P3HT nanostructures (nanofibrils and nanorods). The hfNSCs cultured on the P3HT nanoweb substrates were optically stimulated with a green light (539 nm) and then differentiation of hfNSCs on the substrates with light stimulation was examined. The utility of the nanoweb substrates for optogenetic application was tested with photo-responsive hfNSCs engineered by polymer nanoparticle-mediated transfection of an engineered chimeric opsin variant (C1V1)-encoding gene. Results: The nanoweb substrates provided not only topographical stimulation for activating focal adhesion signaling of hfNSCs, but also generated optoelectrical stimulation via photochemical and charge-transfer reactions upon exposure to 539 nm wavelength light, leading to significantly enhanced neuronal differentiation of hfNSCs. The optoelectrically stimulated hfNSCs exhibited mature neuronal phenotypes with highly extended neurite formation and functional neuron-like electrophysiological features of sodium currents and action potentials. Optoelectrical stimulation with 539 nm light simultaneously activated both C1V1-modified hfNSCs and nanoweb substrates, which upregulated the expression and activation of voltage-gated ion channels in hfNSCs and further increased the effect of photoactive substrates on neuronal differentiation of hfNSCs. Conclusion: The photoactive nanoweb substrates developed in this study may serve as platforms for producing stem cell therapeutics with enhanced neurogenesis and neuromodulation via optoelectrical control of stem cells.