A Rabbit Model for Peripheral Nerve Reconstruction Studies Avoiding Automutilation Behavior.

Background The rabbit sciatic nerve injury model may represent a valuable alternative for critical gap distance seen in humans but often leads to automutilation. In this study, we modified the complete sciatic nerve injury model for avoiding autophagy. Materials and Methods In 20 adult female New Zealand White rabbits, instead of transecting the complete sciatic nerve, we unilaterally transected the tibial portion and preserved the peroneal portion. Thereby loss of sensation in the dorsal aspect of the paw was avoided. The tibial portion was repaired in a reversed autograft approach in a length of 2.6 cm. In an alternative repair approach, a gap of 2.6 cm in length was repaired with a chitosan-based nerve guide. Results During the 6-month follow-up period, there were no incidents of autotomy. Nerve regeneration of the tibial portion of the sciatic nerve was evaluated histologically and morphometrically. A clear difference between the distal segments of the healthy contralateral and the repaired tibial portion of the sciatic nerve was detectable, validating the model. Conclusion By transecting the isolated tibial portion of the rabbit sciatic nerve and leaving the peroneal portion intact, it was possible to eliminate automutilation behavior.

In Vivo and In Vitro Evaluation of a Novel Hyaluronic Acid-Laminin Hydrogel as Luminal Filler and Carrier System for Genetically Engineered Schwann Cells in Critical Gap Length Tubular Peripheral Nerve Graft in Rats.

In the current study we investigated the suitability of a novel hyaluronic acid-laminin hydrogel (HAL) as luminal filler and carrier system for co-transplanted cells within a composite chitosan-based nerve graft (CNG) in a rat critical nerve defect model. The HAL was meant to improve the performance of our artificial nerve guides by giving additional structural and molecular support to regrowing axons. We filled hollow CNGs or two-chambered nerve guides with an inserted longitudinal chitosan film (CNG[F]s), with cell-free HAL or cell-free HA or additionally suspended either naïve Schwann cells (SCs) or fibroblast growth factor 2-overexpressing Schwann cells (FGF2-SCs) within the gels. We subjected female Lewis rats to immediate 15 mm sciatic nerve gap reconstruction and comprehensively compared axonal and functional regeneration parameters with the gold standard autologous nerve graft (ANG) repair. Motor recovery was surveyed by means of electrodiagnostic measurements at 60, 90, and 120 days post-reconstruction. Upon explantation after 120 days, lower limb target muscles were harvested for calculation of muscle-weight ratios. Semi-thin cross-sections of nerve segments distal to the grafts were evaluated histomorphometrically. After 120 days of recovery, only ANG treatment led to full motor recovery. Surprisingly, regeneration outcomes revealed no regeneration-supportive effect of HAL alone and even an impairment of peripheral nerve regeneration when combined with SCs and FGF2-SCs. Furthermore, complementary in vitro studies, conducted to elucidate the reason for this unexpected negative result, revealed that SCs and FGF2-SCs suspended within the hydrogel relatively downregulated gene expression of regeneration-supporting neurotrophic factors. In conclusion, cell-free HAL in its current formulation did not qualify for optimizing regeneration outcome through CNG[F]s. In addition, we demonstrate that our HAL, when used as a carrier system for co-transplanted SCs, changed their gene expression profile and deteriorated the pro-regenerative milieu within the nerve guides.

Modification of tubular chitosan-based peripheral nerve implants: applications for simple or more complex approaches.

Surgical treatment of peripheral nerve injuries is still a major challenge in human clinic. Up to now, none of the well-developed microsurgical treatment options is able to guarantee a complete restoration of nerve function. This restriction is also effective for novel clinically approved artificial nerve guides. In this review, we compare surgical repair techniques primarily for digital nerve injuries reported with relatively high prevalence to be valuable attempts in clinical digital nerve repair and point out their advantages and shortcomings. We furthermore discuss the use of artificial nerve grafts with a focus on chitosan-based nerve guides, for which our own studies contributed to their approval for clinical use. In the second part of this review, very recent future perspectives for the enhancement of tubular (commonly hollow) nerve guides are discussed in terms of their clinical translatability and ability to form three-dimensional constructs that biomimick the natural nerve structure. This includes materials that have already shown their beneficial potential in in vivo studies like fibrous intraluminal guidance structures, hydrogels, growth factors, and approaches of cell transplantation. Additionally, we highlight upcoming future perspectives comprising co-application of stem cell secretome. From our overview, we conclude that already simple attempts are highly effective to increase the regeneration supporting properties of nerve guides in experimental studies. But for bringing nerve repair with bioartificial nerve grafts to the next level, e.g. repair of defects > 3 cm in human patients, more complex intraluminal guidance structures such as innovatively manufactured hydrogels and likely supplementation of stem cells or their secretome for therapeutic purposes may represent promising future perspectives.

Two-Chambered Chitosan Nerve Guides With Increased Bendability Support Recovery of Skilled Forelimb Reaching Similar to Autologous Nerve Grafts in the Rat 10 mm Median Nerve Injury and Repair Model.

Tension-free surgical reconstruction of transected digital nerves in humans is regularly performed using autologous nerve grafts (ANGs) or bioartificial nerve grafts. Nerve grafts with increased bendability are needed to protect regenerating nerves in highly mobile extremity parts. We have recently demonstrated increased bendability and regeneration supporting properties of chitosan nerve guides with a corrugated outer wall (corrCNGs) in the common rat sciatic nerve model (model of low mobility). Here, we further modified the hollow corrCNGs into two-chambered nerve guides by inserting a perforated longitudinal chitosan-film (corrCNG[F]s) and comprehensively monitored functional recovery in the advanced rat median nerve model. In 16 adult female Lewis rats, we bilaterally reconstructed 10 mm median nerve gaps with either ANGs, standard chitosan nerve guides (CNGs), CNGs (CNG[F]s), or corrCNG[F]s (n = 8, per group). Over 16 weeks, functional recovery of each forelimb was separately surveyed using the grasping test (reflex-based motor task), the staircase test (skilled forelimb reaching task), and non-invasive electrophysiological recordings from the thenar muscles. Finally, regenerated tissue harvested from the distal part of the nerve grafts was paraffin-embedded and cross-sections were analyzed regarding the number of Neurofilament 200-immunopositive axons and the area of newly formed blood vessels. Nerve tissue harvested distal to the grafts was epon-embedded and semi-thin cross-sections underwent morphometrical analyses (e.g., number of myelinated axons, axon and fiber diameters, and myelin thicknesses). Functional recovery was fastest and most complete in the ANG group (100% recovery rate regarding all parameters), but corrCNG[F]s accelerated the recovery of all functions evaluated in comparison to the other nerve guides investigated. Furthermore, corrCNG[F]s supported recovery of reflex-based grasping (87.5%) and skilled forelimb reaching (100%) to eventually significantly higher rates than the other nerve guides (grasping test: CNGs: 75%, CNG[F]s: 62.5%; staircase test: CNGs: 66.7%, CNG[F]s: 83.3%). Histological and nerve morphometrical evaluations, in accordance to the functional results, demonstrated best outcome in the ANG group and highest myelin thicknesses in the corrCNG[F] group compared to the CNG and CNG[F] groups. We thus clearly demonstrate that corrCNG[F]s represent promising innovative nerve grafts for nerve repair in mobile body parts such as digits.