Recombinant Spider Silk and Collagen-Based Nerve Guidance Conduits Support Neuronal Cell Differentiation and Functionality in Vitro

Abstract

Biomaterial scaffolds are under investigation as therapeutic tools to bridge nerve endings following traumatic peripheral nerve injury. The goal is to develop biocompatible nerve guidance conduits (NGCs) with internal guiding structures that promote longitudinally oriented cell migration and regeneration. In the present study, a nonwoven mesh (NWM) made of a recombinant spider silk protein was processed into a tubular structure, ensuring structural integrity of enclosed microfluidics-produced collagen fibers for cell and neurite guidance. The differentiated type of the neuroblastoma X glioma hybrid cell line NG108-15 was used as a model for studying neuronal differentiation on the individual components and on the complete NGC. Differentiated NG108-15 cells grown on recombinant spider silk NWM and collagen fibers formed neuronal networks and synapses. Additionally, whole-cell patch clamp recordings confirmed that all components supported the differentiation of NG108-15 cells into functional neurons. Our NGC demonstrated that tubes made of recombinant spider silk NWM filled with microfluidicsproduced collagen fibers are well suited for peripheral nerve repair.

Defining the regenerative effects of native spider silk fibers on primary Schwann cells, sensory neurons, and nerve‐associated fibroblasts

“…Recombinant silk shall not only possess the unique material characteristics, but also show the same (or superior) effect on cells. Previous studies suggest different methods to analyze the behavior of cells to silk fibers 57,63,[65][66][67] or silk-based materials 68,69 in vitro. However, quantitative and qualitative read-outs designed to assess the biological response of cells to silk fibers concerning nerve tissue regeneration are scarce.…”

Spider Silk for Tissue Engineering Applications

“…Pawar et al [86,87] and Aigner et al [88] reported the development of NGCs using recombinant spider silk eADF4(C16) materials. Pawar et al [86,87] showed the use of fibers made of eADF4(C16) nonwoven-electrospun meshes filled with microfluidics-produced collagen fibers [87]. In this study, neuroblastoma X glioma hybrid cells NG108-15 were used and differentiated to study the effect of the individual components of the NGC on cell behavior.…”

“…PC-12 cells cultured in these constructs showed differentiation and formation of aligned neurites within the microchannels, as well as along the aligned fibers of spider silk. Pawar et al [86,87] and Aigner et al [88] reported the development of NGCs using recombinant spider silk eADF4(C16) materials. Pawar et al [86,87] showed the use of fibers made of eADF4(C16) nonwoven-electrospun meshes filled with microfluidics-produced collagen fibers [87].…”

“…(D) Whole-cell patch clamp recordings from cells grown on NGCs, demonstrating the maturation of cells and generation of action potentials in response to current injections (100 pA, 300 ms). Adopted and modified with permission [87]. Copyright 2019, American Chemical Society.…”

Silkworm and spider silk electrospinning: a review

“…The peripheric tissues have shown excellent compatibility with cell growth and differentiation, the cells being able to achieve functional neuronal activity. Recombinant spider silk provided the anti-inflammatory barrier to the collagen compounds, while allowing the passage of nutrients and gas exchanges (Pawar et al 2019 ; Salehi et al 2020 ).…”