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Three-dimensional reconstruction of nerve fascicle is important in the analysis of biological characteristics in the arm. The topology of fascicle has been used by doctors to investigate the nerve direction and the relationship between the individual nerve fascicle. However, there still does not exist an ideal internal fascicle and 3D model in the human peripheral nerve. Accurate segmentation of fascicle from CT images is a crucial step to obtain reliable 3D nerve fascicle model. Traditional method in the fascicle segmentation is not efficient due to time consuming, manual work and poor generalization capacity. In this study, we proposed an efficient deep segmentation network and then reconstruct 3D nerve fascicle model. The proposed network explores the intra-slice contextual features with convolutional long short-term memory for accurate fascicle segmentation, and model long-range semantic information among image slices. Transfer learning technique is integrated with ResNet34, and the discriminative capability of intermediate features are further improved. The proposed network architecture is efficient, flexible and suitable for separating the adhesive fascicle. Our approach is the first deep learning method for nerves segmentation. The proposed approach achieves state-of-the-art performance on our dataset, where the mean Dice of our method is 95.4% and at least 5% more than other methods.

Biofabricated nanostructured and microstructured scaffolds have exhibited great potential for nerve tissue regeneration and functional restoration, and prevascularization and biotransportation within 3D fascicle structures are critical. Unfortunately, an ideal internal fascicle and microvascular model of human peripheral nerves is lacking. In this study, we used microcomputed tomography (microCT) to acquire high-resolution images of the human sciatic nerve. We propose a novel deep-learning network technique, called ResNetH3D-Unet, to segment fascicles and microvascular structures. We reconstructed 3D intraneural fascicles and microvascular topography to quantify the fascicle volume ratio (FVR), microvascular volume ratio (MVR), microvascular to fascicle volume ratio (MFVR), fascicle surface area to volume ratio (FSAVR), and microvascular surface area to volume ratio (MSAVR) of human samples. The frequency distributions of the fascicle diameter, microvascular diameter, and fascicle-to-microvasculature distance were analyzed. The obtained microCT analysis and reconstruction provided high-resolution microstructures of human peripheral nerves. Our proposed ResNetH3D-Unet method for fascicle and microvasculature segmentation yielded a mean intersection over union (IOU) of 92.1% (approximately 5% higher than the U-net IOU). The 3D reconstructed model showed that the internal microvasculature runs longitudinally within the internal epineurium and connects to the external vasculature at some points. Analysis of the 3D data indicated a 48.2 ± 3% FVR, 23.7 ± 1.8% MVR, 4.9 ± 0.5% MFVR, 7.26 ± 2.58 mm-1 FSAVR, and 1.52 ± 0.52 mm-1 MSAVR. A fascicle diameter of 0.98 mm, microvascular diameter of 0.125 mm, and microvasculature-to-fascicle distance of 0.196 mm were most frequent. This study provides fundamental data and structural references for designing bionic scaffolding constructs with 3D microvascular and fascicle distributions.

Objective To investigate whether lymphoedema in a Chinese family showed the hereditary and clinical characteristics of Milroy disease, an autosomal dominant form of congenital lymphoedema, typically characterized by chronic lower limb tissue swelling due to abnormal lymphatic vasculature development, and to perform mutational analyses of vascular endothelial growth factor receptor ( VEGFR)3. Methods Individuals from a three-generation family affected by congenital lymphoedema were clinically assessed for Milroy disease. Mutation analysis of VEGFR3 was performed using DNA from family members and healthy controls. Results Out of 20 family members, eight were diagnosed with hereditary lymphoedema. Mutation analyses revealed a novel mutation site for c.3163 G>A, resulting in a p.1055D>N mutation in the second tyrosine kinase domain of VEGFR3, which was present in affected individuals only (absent in all unaffected family members and 130 healthy controls). Computed functional analyses showed the mutation may lead to structural alterations with a probability of 0.99999 of being disease causing. Conclusion A novel mutation associated with Milroy disease was identified in a Chinese family, expanding our knowledge of VEGFR3 gene function and providing a potential molecular target for treating hereditary lymphoedema.

Decellularized matrix hydrogels derived from tissues or organs have been used for tissue repair due to their biocompatibility, tunability, and tissue-specific extracellular matrix (ECM) components. However, the preparation of decellularized peripheral nerve matrix hydrogels and their use to repair nerve defects have not been reported. Here, we developed a hydrogel from porcine decellularized nerve matrix (pDNM-G), which was confirmed to have minimal DNA content and retain collagen and glycosaminoglycans content, thereby allowing gelatinization. The pDNM-G exhibited a nanofibrous structure similar to that of natural ECM, and a ∼280-Pa storage modulus at 10 mg/mL similar to that of native neural tissues. Western blot and liquid chromatography tandem mass spectrometry analysis revealed that the pDNM-G consisted mostly of ECM proteins and contained primary ECM-related proteins, including fibronectin and collagen I and IV). In vitro experiments showed that pDNM-G supported Schwann cell proliferation and preserved cell morphology. Additionally, in a 15-mm rat sciatic nerve defect model, pDNM-G was combined with electrospun poly(lactic-acid)-co-poly(trimethylene-carbonate) conduits to bridge the defect, which did not elicit an adverse immune response and promoted the activation of M2 macrophages associated with a constructive remodeling response. Morphological analyses and electrophysiological and functional examinations revealed that the regenerative outcomes achieved by pDNM-G were superior to those by empty conduits and closed to those using rat decellularized nerve matrix allograft scaffolds. These findings indicated that pDNM-G, with its preserved ECM composition and nanofibrous structure, represents a promising biomaterial for peripheral nerve regeneration. Statement of Significance Decellularized nerve allografts have been widely used to treat peripheral nerve injury. However, given their limited availability and lack of bioactive factors, efforts have been made to improve the efficacy of decellularized nerve allograft for nerve regeneration, with limited success. Xenogeneic decellularized tissue matrices or hydrogels have been widely used for surgical applications owing to their ease of harvesting and low immunogenicity. Moreover, decellularized tissue matrix hydrogels show good biocompatibility and are highly tunable. In this study, we prepared a porcine decellularized nerve matrix (pDNM-G) and evaluated its potential for promoting nerve regeneration. Our results demonstrate that pDNM-G can support Schwann cell proliferation and peripheral nerve regeneration by means of residual primary extracellular matrix components and nano-fibrous structure features.

Background The C5-C6 nerve roots are usually spared from avulsion after brachial plexus injury (BPI) and thus can be used as donors for nerve grafting. To date, there are no appropriate animal models to evaluate spared nerve root stumps. Hence, the aim of this study was to establish and evaluate a rat model with spared nerve root stumps in BPI. New method In rupture group, the proximal parts of C5-T1 nerve roots were held with the surrounding muscles and the distal parts were pulled by a sudden force after the brachial plexus was fully exposed, and the results were compared with those of sham group. To validate the model, the lengths of C5-T1 spared nerve root stumps were measured and the histologies of the shortest one and the corresponding spinal cord were evaluated. Results C5 nerve root stump was found to be the shortest. Histology findings demonstrated that the nerve fibers became more irregular and the continuity decreased; numbers and diameters of myelinated axons and thickness of myelin sheaths significantly decreased over time. The survival of motoneurons was reduced, and the death of motoneurons may be related to the apoptotic process. Comparison with existing method(s) Our model could successfully create BPI model with nerve root stumps by traction, which could simulate injury mechanisms. While other models involve root avulsion or rupturing by distal nerve transection. Conclusions This model would be suitable for evaluating nerve root stumps and testing new therapeutic strategies for neuroprotection through nerve root stumps in the future.

Background/Objectives Complex extremity trauma commonly involves both soft tissue and vascular injuries. Traditional two-stage surgical repair may delay rehabilitation and functional recovery, as well as increase the risk of infections. We report a single-stage reconstructive surgical method that repairs soft tissue defects and vascular injuries with flow-through free flaps to improve functional outcomes. Methods Between March 2010 and December 2016 in our hospital, 5 patients with severe upper extremity trauma received single-stage reconstructive surgery, in which a flow-through anterolateral thigh free flap was applied to repair soft tissue defects and vascular injuries simultaneously. Cases of injured artery were reconstructed with the distal trunk of the descending branch of the lateral circumflex femoral artery. A segment of adjacent vein was used if there was a second artery injury. Patients were followed to evaluate their functional recoveries, and received computed tomography angiography examinations to assess peripheral circulation. Results Two patients had post-operative thumb necrosis; one required amputation, and the other was healed after debridement and abdominal pedicle flap repair. The other 3 patients had no major complications (infection, necrosis) to the recipient or donor sites after surgery. All the patients had achieved satisfactory functional recovery by the end of the follow-up period. Computed tomography angiography showed adequate circulation in the peripheral vessels. Conclusions The success of these cases shows that one-step reconstructive surgery with flow-through anterolateral thigh free flaps can be a safe and effective treatment option for patients with complex upper extremity trauma with soft tissue defects and vascular injuries.

Background The precise annotation and accurate identification of the topography of fascicles to the end organs are prerequisites for studying human peripheral nerves. New method In this study, we present a feasible imaging method that acquires 3D high-resolution (HR) topography of peripheral nerve fascicles using an iodine and freeze-drying (IFD) micro-computed tomography (microCT) method to greatly increase the contrast of fascicle images. Results The enhanced microCT imaging method can facilitate the reconstruction of high-contrast HR fascicle images, fascicle segmentation and extraction, feature analysis, and the tracing of fascicle topography to end organs, which define fascicle functions. Comparison with existing methods The complex intraneural aggregation and distribution of fascicles is typically assessed using histological techniques or MR imaging to acquire coarse axial three-dimensional (3D) maps. However, the disadvantages of histological techniques (static, axial manual registration, and data instability) and MR imaging (low-resolution) limit these applications in reconstructing the topography of nerve fascicles. Conclusions Thus, enhanced microCT is a new technique for acquiring 3D intraneural topography of the human peripheral nerve fascicles both to improve our understanding of neurobiological principles and to guide accurate repair in the clinic. Additionally, 3D microstructure data can be used as a biofabrication model, which in turn can be used to fabricate scaffolds to repair long nerve gaps.

The exact inner 3D microstructure of the human peripheral nerve has been a mystery for decades. Therefore, it has been difficult to solve several problems regarding peripheral nerve injury and repair. We used high-resolution X-ray computed microtomography (microCT) to scan a freeze-dried human acellular nerve allograft (hANA). The microCT images were then used to reconstruct a 3D digital model, which was used to print a 3D resin model of the nerve graft. The 3D digital model of the hANA allowed visualization of all planes. The magnified 3D resin model clearly showed the nerve bundles and basement membrane tubes of the hANA. Scanning electron microscopy (SEM) was used to analyse the microstructure of the hANA. Compared to the SEM images, the microCT image clearly demonstrated the microstructure of the hANA cross section at a resolution of up to 1.2 μm. The 3D digital model of the hANA facilitates a clear and easy understanding of peripheral nerve microstructure. Furthermore, the enlarged 3D resin model duplicates the unique inner structure of each individual hANA. This is a crucial step towards achieving 3D printing of a hANA or nerve that can be used as a nerve graft.