Gelatin fibre diameter influences Schwann cell behaviour and axonal outgrowth

For tissue engineering application, the distribution and growth of cells on a scaffold are key requirements. A number of studies demonstrated that micro-to-nano-scale topography plays an important role in controlling cell adhesion, proliferation and survival. Schwann cells (SC), forming bands of Büngner, support and promote axonal outgrowth leading to target reinnervation during nerve regeneration. In order to enhance SC adhesion, proliferation, migration and axonal outgrowth, a number of bio-mimetic materials were studied. Appropriate fibrous substrates, functioning as a temporary extracellular-matrix, can be easily prepared by electrospinning technique, which allows the obtainment of fibrous matrices suitable as internal filler for nerve guidance channels to be applied in peripheral nerve repair. Gelatin micro or nano-fibres were prepared by electrospinning technique by tuning gelatin concentration and solution flow rate. The influence of gelatine fibre diameter on cell adhesion and proliferation, was tested in vitro by using SC and Dorsal Root Ganglia (DRG) explant cultures. Results demonstrated that gelatin fibres tested were biocompatible. Cell adhesion was evaluated by quantifying cell spreading area, actin cytoskeleton organization and focal adhesion complex formation. Fibre diameter influences SC behaviour and morphology. Nano-fibres have been shown to promote cell spreading and actin cytoskeleton organization, resulting in higher cellular adhesion and proliferation rate in comparison to micro-fibres. Cell migration and motility were quantified by transwell and time lapse assays respectively. Cells cultured on micro-fibres displayed higher motility and migration rate in comparison to nano-fibres. Finally, axonal outgrowth evaluated by culturing DRG explants on the different fibres resulted in higher axonal outgrowth on micro-fibres in comparison to nano-fibres. These data provide a better understanding about glial cell and neuritis viability and organization on gelatin electrospun nano- and micro-fibres suggesting that micro-fibres can be a better filler to be used in the design of new devices for peripheral nerve repair applications.