Numerous nanostructured synthetic scaffolds mimicking the architecture of the natural extracellular matrix (ECM) have been described, but the polymeric nanofibers comprising the scaffold were substantially thicker than the natural collagen nanofibers of neural ECM. Here, we report neuron growth on electrospun scaffolds of nylon-4,6 fibers with an average diameter of 60 nm, which closely matches the diameter of collagen nanofibers of neural ECM, and compare their properties with the scaffolds of thicker 300 nm nanofibers. Previously unmodified nylon was not regarded as an independent nanostructured matrix for guided growth of neural cells; however, it is particularly useful for ultrathin nanofiber production. We demonstrate that, while both types of fibers stimulate directed growth of neuronal processes, ultrathin fibers are more efficient in promoting and accelerating neurite elongation. Both types of scaffolds also improved synaptogenesis and the formation of connections between hippocampal neurons; however, the mechanisms of interaction of neurites with the scaffolds were substantially different. While ultrathin fibers formed numerous weak immature β1-integrin-positive focal contacts localized over the entire cell surface, scaffolds of submicron fibers formed β1-integrin focal adhesions only on the cell soma. This indicates that the scaffold nanotopology can influence focal adhesion assembly involving various integrin subunits. The fabricated nanostructured scaffolds demonstrated high stability and resistance to biodegradation, as well as absence of toxic compound release after 1 month of incubation with live cells in vitro. Our results demonstrate the high potential of this novel type of nanofibers for clinical application as substrates facilitating regeneration of nervous tissue.
Keywords: focal adhesions; nanofibers; neurites growth guidance; nylon scaffolds.