Characterisation of cell–substrate interactions between Schwann cells and three‐dimensional fibrin hydrogels containing orientated nanofibre topographical cues

Abstract

The generation of complex three-dimensional bioengineered scaffolds that are capable of mimicking the molecular and topographical cues of the extracellular matrix found in native tissues is a field of expanding research. The systematic development of such scaffolds requires the characterisation of cell behaviour in response to the individual components of the scaffold. In the present investigation, we studied cell-substrate interactions between purified populations of Schwann cells and three-dimensional fibrin hydrogel scaffolds, in the presence or absence of multiple layers of highly orientated electrospun polycaprolactone nanofibres. Embedded Schwann cells remained viable within the fibrin hydrogel for up to 7 days (the longest time studied); however, cell behaviour in the hydrogel was somewhat different to that observed on the two-dimensional fibrin substrate: Schwann cells failed to proliferate in the fibrin hydrogel, whereas cell numbers increased steadily on the two-dimensional fibrin substrate. Schwann cells within the fibrin hydrogel developed complex process branching patterns, but, when presented with orientated nanofibres, showed a strong tendency to redistribute themselves onto the nanofibres, where they extended long processes that followed the longitudinal orientation of the nanofibres. The process length along nanofibre-containing fibrin hydrogel reached near-maximal levels (for the present experimental conditions) as early as 1 day after culturing. The ability of this three-dimensional, extracellular matrix-mimicking scaffold to support Schwann cell survival and provide topographical cues for rapid process extension suggest that it may be an appropriate device design for the bridging of experimental lesions of the peripheral nervous system.

Cell-free artificial implants of electrospun fibres in a three-dimensional gelatin matrix support sciatic nerve regeneration in vivo

“…On the other hand, the potential for successful regeneration diminishes with time after injury (Xu et al , ), implying that the presence of a growth substrate may be an important factor for functional recovery within a critical window of opportunity. Previous experiments with dorsal root ganglia and Schwann cell explants in 3D gels (Hodde et al , ; Kriebel et al , ) demonstrated the significance of the electrospun fibres for cell migration and axonal elongation. However, an effect of the gel alone cannot be dismissed entirely because no gelatin‐containing conduits without fibres were tested.…”

“…This effect could be enhanced when fibres included biological signals, for example by blending PCL with collagen (c/PCL; Schnell et al , ) or by covalent binding of integrin‐activating peptides (Bockelmann et al , ). This was first shown on two‐dimensional (2D) substrates in vitro and recently transferred to three‐dimensional (3D) substrates with fibres embedded in hydrogels (Hodde et al , ; Kriebel et al , ). The procedure developed in this study is based on 3D spinning of aligned microfibres, which are subsequently embedded in a hydrogel.…”

A bench‐top molding method for the production of cell‐laden fibrin micro‐fibers with longitudinal topography

“…For the estimation of cell purity, sample cultures that were seeded onto PLL/laminin‐coated 12 mm glass coverslips were fixed for 30 min in cold 4% paraformaldehyde (in 0.1 M PBS). The cells were processed for double immunofluorescence as previously described (Hodde et al, ) and incubated with a mouse monoclonal antibody against vimentin (V6630, Sigma‐Aldrich) diluted 1:1000, and polyclonal rabbit antibodies against S100β (T8578, Dako, Z0311, Hamburg, Germany) diluted 1:1000.…”

Spatially and temporally controlled hydrogels for tissue engineering

“…Hodde et al reported another approach to provide topographical cues to hydrogel systems [18]. Individual frames with aligned polycaprolactone (PCL) electrospun nanofibers were prepared, which were then stacked together (with parallel fiber orientation).…”