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International Journal of Bioprinting 3D bioprinting of nerve guidance conduits

During the bioprinting process, SCs were encapsulated (Figure 9A-iii) compared to the silicone group (Figure 9A-
within the conduit, promoting the longitudinal alignment iv), which could promote nerve regeneration in a rat sciatic
of fine fibrin fibers. The physical guidance cues provided nerve defect model. Although there are some limitations,
by these longitudinally aligned fibrin fibers further the study confirmed that this new purely biological NGC
directed the SCs to align linearly and facilitated linear is effective in promoting nerve regeneration. Alternatively,
neurite elongation along the fibrin factor XIII-hyaluronic Takeuchi et al. explored an approach for larger-gap nerve
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acid chain. Finally, Das et al. formulated cell-carrying defect repair. A 12 mm NGC (Figure 9B-i & ii) was prepared
biobricks with Neuro-2a cell density of 3 × 10 cells/mL into using the Kenzan method and implanted into a 10 mm
6
GelMA/carbon nanofiber/PEGDA/gellan gum hydrogels sciatic nerve defect in the right hind limbs of rats. A silicone
and printed them into two layers of cylindrical conduits tube implantation group was used as a control. Evaluation
(wall thickness of 0.5 mm) using a customized extrusion was performed 8 weeks after surgery, in the distal region
bioprinter. After 5 days of incubation in a differentiation of the suture site, where the number of myelinated axons,
medium, the Neuro-2a cells showed good cell viability myelin sheath diameter, and myelin sheath thickness of
(more than 80%) (Figure 8E). regenerated axons were significantly greater in the NGC
(Figure 9B-iii) than in the silicone (Figure 9B-iv) groups.
4.4. Kenzan It was eventually shown that the NGCs could promote
The Kenzan method uses pre-designed 3D data and peripheral nerve regeneration, even in a 10 mm nerve defect
stainless steel microneedle arrays as temporary scaffolds model. On this basis, Yurie et al. further investigated the
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on which multi-cellular spheres are precisely placed. promotion of nerve regeneration using NGCs generated
Once these spheroids fuse and produce their own ECM, from bone marrow stromal cells. NGCs were fabricated
the microneedles are removed, leaving behind the desired with the Kenzan method and transplanted into Lewis rats
biological structure. This method does not require a to bridge the 5 mm right sciatic nerve gap (Figure 9C-i),
biopolymer solution or hydrogel, relying solely on cells to with two silicone tubes used as control (Figure 9C-ii &
construct fully biological conduits with a certain degree iii). Functional and morphological evaluation of nerve
of biomechanical stability. This approach pioneers a new regeneration was performed 12 weeks after transplantation.
direction in NTE. Needle diameters are typically 100– Electrophysiological studies, kinematic analysis, wet muscle
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200 µm with a needle pitch of 300–400 µm. The size of the weight, and morphological parameters showed that nerve
multi-cellular spheres is determined by the needle spacing. regeneration in the NGC was significantly better than in
The distribution of cells within the sphere undergoes a the silicone tube. Mitsuzawa et al. further investigated
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continuous rearrangement, with cells exhibiting stronger
adhesion migrating toward the core, while those with the effect of this method on nerve defect regeneration
weaker adhesive properties localize to outer concentric promotion in large mammals. Spheres were extracted from
a 96-well plate into a fine nozzle, strung into an array of
layers in decreasing order of adhesion strength. Inside circular needles (Figure 9D-i & ii), and developed into
the sphere, cells move within the available space and a tubular structure according to a predesigned pattern
limits imposed by intercellular adhesions. This process,
combined with ECM deposition, promotes the healing of (Figure 9D-iii). An NGC (Figure 9D-iv–vi) was used to
pinholes. However, since the goal is to print tubes or hollow bridge the ulnar nerve defect in a dog’s 5 mm forelimb, and
structures, this contraction may cause them to disappear nerve regeneration was observed at 10 weeks postoperatively.
prematurely, which would require additional stabilization. Immunohistochemical, histologic, and morphometric
assays confirmed the presence of numerous myelinated
Yurie et al. used the Kenzan method to prepare axons in the NGC. It was shown that the NGC fabricated
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a conduit-free hollow NGC (Figure 9A) from human from canine autologous dermal fibroblasts promoted nerve
normal dermal fibroblasts and verified its restorative regeneration even in a 10 mm nerve defect model. This
effect on sciatic nerve defects in rats. Human dermal technique is therefore feasible for the preclinical treatment
cells were first aggregated to form homogeneous multi- of PNI and segmental nerve defects.
cellular spheres with a diameter of 750 ± 50 μm, and the
spheres were arranged into a 3D shape according to a pre- 5. Clinical translation and commercial
designed 3D model (Figure 9A-i). After about a week of application: moving toward
printing, adjacent spheres were fused together to construct scalable solutions
a single tubular shape in a microneedle array, which
was then removed (Figure 9A-ii). In vivo experiments This review reports the high potential of biofabricating
demonstrated that 3D-bioprinted fibroblast conduits NGCs for nerve tissue repair, focusing on their structure,
showed a significantly higher number of myelinated axons AM-oriented technology, biomaterials, and cells. The

Volume 11 Issue 4 (2025) 54 doi: 10.36922/IJB025140120