Fabrication of a biosimulated nerve repair device for guided axon regeneration and controlled release of bioactives for the treatment of peripheral nerve injury

Abstract

The constant challenges imposed by the clinical procedures in obtaining adequate nerve repair and return of functional recovery to the injured site has buoyed the pursuit for artificial alternatives instead of the utilization of donor tissue grafts for surgical repair. Transected nerve injuries, having tremendous potential to cause permanent paralysis, neuropathies and chronic pain, requires the collective involvement of tissue-engineering and advanced drug delivery applications in combating the perceived challenges associated with its repair. Modern peripheral nerve repair strategies employ the use of polymeric-engineered nerve conduits incorporated with components of a delivery system. This allows for the controlled and sustained release of neurotrophic growth factors and other bioactives for improvements in the innate regenerative capacity of neural tissues. Although currently available synthetic hollow nerve conduits available for clinical use, as an alternative to the nerve autografts, have proven to be successful in the bridging and regeneration of primarily short nerve gap injuries, recent developments entail advancement of nerve conduits to be able to simulate the effectiveness of the autograft which includes, in particular, the ability to deliver growth factors and mimic peripheral nerve structure. A critical factor influencing the level of success of peripheral nerve repair and regeneration is the development of appropriate sustained drug delivery systems for the release of neurotrophic factors and other small molecular weight drugs in addition to the provision of a mechanically stable, biocompatible and preferably biodegradable repair scaffold. This study, therefore, aimed to investigate and establish the proposed concept of pristine polymer particle intercalation as a novel strategy for achieving controlled and sustained release of bioactives from polymeric implants. An initial study involving an investigation into the drug release mechanisms of indomethacin from swellable sodium tripolyphosphatecrosslinked chitospheres laden with pristine pH-responsive polymethylmethacrylate (PMMA) nanoparticles sought to prove the concept of particle intercalation as a modulator for drug release. Swelling and erosion studies, in conjunction with textural profiling, provided an understanding of the dominant and underlying drug release mechanisms of the chitospheres loaded with the pristine PMMA particles. Drug release studies showed that the pristine particle-intercalated chitospheres extended the release of indomethacin over 144 hours in a first-order compared to the 72 hours release achieved with the control. The study further revealed that in situ porogen leaching leading to porous network and polyelectrolyte complex formation were major mechanisms governing drug release from the chitospheres. Having

successfully proved the concept of pristine polymer particle intercalation, this technique was consequentially applied to the development of a nerve conduit for the dual and controlledrelease of proteins and small-molecule drugs. The design and fabrication of a hollow nerve conduit comprised a physically crosslinked interpenetrating network of a gellan-xanthan hydrogel matrix intercalated with pristine PMMA particles for provision sustained and concurrent release of two model compounds: bovine serum albumin (BSA) and diclofenac sodium. Analysis of a Box-Behnken experimental design demonstrated a near zero-order release of BSA and diclofenac sodium over 20 and 30 days, respectively, modulated via a combination of pH-responsive (pH 7.4) dissolution of the intercalated pristine polymer particles and the unique gelling and erosion properties imparted by the graded addition of xanthan gum to the hydrogel blend. Moreover, the concentration-dependent intercalation of PMMA enhanced matrix resilience while alterations in the gellan-xanthan proportions yielded a means for customizing the mechanical characteristics, particularly matrix rigidity and flexibility, for the attainment of neuro-durable conduits. Optimization of the hydrogel conduit provided an effective means for the sustained delivery of the essential but costly neurotrophic growth factor, Nerve Growth Factor (NGF), over 30 days whilst maintaining its bioactivity. Further advancement of the hollow-lumen gellan-xanthan conduits involved implementation of intraluminal guidance scaffolds in an endeavour for simulating the native structural features of peripheral nerve tissue. An array of electrospun aligned nano-fibrous scaffolds formulated for the inclusion of physical, chemical and therapeutic guidance cues was achieved through the blending and incorporation of polyhydroxy butyric-co-valeric acid base polymer with magnesium-oleate and Nacetylcysteine as potential neuro-active agents as indicated by the enhanced proliferation of PC12 neuronal cells. Integration of the two components, the hydrogel conduit and the nanofibrous scaffold, heralded the assembly of the final Biosimulated Nerve Repair Device (BNRD) which proved proficient in an in vivo rat sciatic nerve model in promoting nerve regeneration and re-establishment of functional recovery compared to the autograft gold standard repair as determined by behavioural assessments and morphometric and histological analysis of muscle and nerve tissues. The devised BNRD system for the surgical repair and therapeutic treatment of transected peripheral nerve injuries was successful in providing a long-term neuro-durable scaffold with release kinetics appropriate for the adequate promotion of tissue regeneration and healing bringing about restoration of functional recovery through emulating the positive characteristics of the autograft. The results obtained show promise for the rather challenging repair of peripheral nerves.