Interactions of Peripheral Nerve Cells with Soft Micropillar Interfaces

Abstract

Regenerative peripheral nerve interfaces are particularly invasive implant devices. They rely on the growth of axons from transected nerve stumps through electrode-bearing structures in order to establish highly efficient communication channels with the nervous system. The functioning and lifetime of these devices is challenged by the host response from the nervous tissue, which complicates their use in long-term applications. The elicited foreign body reaction is characterized by chronic inflammation and excessive deposition of collagen. The difference in mechanical properties between implant material and nervous tissue, which are among the softest in the body, is thought to be an important contributor to these adverse effects. With the motivation of improving the bio-integration of implanted nerve interfaces, this thesis evaluate micropillar arrays as a texture to modulate the behaviour of peripheral nerve cells on polydimethylsiloxane (PDMS) silicone surfaces. Thanks to their flexibility, micropillars can reduce the effective stiffness perceived by cells, while providing topographic cues at the microscopic level. Importantly, these mechanical and topographic cues can be easily tailored by modification of their geometrical dimensions. Micropillar arrays with varying dimensions were fabricated by soft lithography process. Their mechanical behaviour under bending was simulated by finite element method (FEM) analysis to approximate their spring constants. The softest micropillar configuration had an effective surface stiffness of 0.7 kPa, representing a reduction of 3 orders of magnitude compared to bulk PDMS. The effect of micropillar diameter (from 1.2 to 4.2 um), modulating the surface stiffness, and the effect of interpillar spacing (from 1.0 to 7.6 um), modifying micropillars density, were investigated in vitro. Dissociated cell cultures from dorsal root ganglion (DRG), comprising neurons and glial cells, were probed for different parameters after 7 days of incubation. While neurons spread and established neurite networks on all configurations, fewer glial cells were found on the softest and lowest density of micropillars. Neurites explored this pseudo-3D environment and preferentially anchored to the top of pillar shafts, well above the bottom surface. The matrix of pillars provided physical cues for growing neurites, which strongly aligned with the densest arrays. The morphology of neuronal bodies was affected by the most flexible pillars, which induced smaller and rounder somas. Macrophages and adipose-derived stem cells showed significantly increased attachment to micropillar topographies compared to flat surface. The effect of micropillars was investigated on nerve regeneration in vivo. Nerve guidance conduits patterned with micropillars on their lumen surface were sutured to the transected sciatic nerve of rats. While the axonal regeneration was slightly affected by the micropillar texture, macrophages were massively recruited to the micropillar surface in comparison to the flat control. Their phenotype was also affected, with predominantly classically activated macrophage in contact with the micropillars. Altogether, these results demonstrate the possibility to modulate the behaviour of a variety of cell types in vitro and in vivo, through the patterning of micropillars on the surface. This approach is amenable to implementation on silicone neural implants for mitigating tissue-material interactions.