Preparation of PLCL/ECM nerve conduits by electrostatic spinning technique and evaluationin vitroandin vivo.

Objective. Artificial nerve scaffolds composed of polymers have attracted great attention as an alternative for autologous nerve grafts recently. Due to their poor bioactivity, satisfactory nerve repair could not be achieved. To solve this problem, we introduced extracellular matrix (ECM) to optimize the materials.Approach.In this study, the ECM extracted from porcine nerves was mixed with Poly(L-Lactide-co-ϵ-caprolactone) (PLCL), and the innovative PLCL/ECM nerve repair conduits were prepared by electrostatic spinning technology. The novel conduits were characterized by scanning electron microscopy (SEM), tensile properties, and suture retention strength test for micromorphology and mechanical strength. The biosafety and biocompatibility of PLCL/ECM nerve conduits were evaluated by cytotoxicity assay with Mouse fibroblast cells and cell adhesion assay with RSC 96 cells, and the effects of PLCL/ECM nerve conduits on the gene expression in Schwann cells was analyzed by real-time polymerase chain reaction (RT-PCR). Moreover, a 10 mm rat (Male Wistar rat) sciatic defect was bridged with a PLCL/ECM nerve conduit, and nerve regeneration was evaluated by walking track, mid-shank circumference, electrophysiology, and histomorphology analyses.Main results.The results showed that PLCL/ECM conduits have similar microstructure and mechanical strength compared with PLCL conduits. The cytotoxicity assay demonstrates better biosafety and biocompatibility of PLCL/ECM nerve conduits. And the cell adhesion assay further verifies that the addition of ECM is more beneficial to cell adhesion and proliferation. RT-PCR showed that the PLCL/ECM nerve conduit was more favorable to the gene expression of functional proteins of Schwann cells. Thein vivoresults indicated that PLCL/ECM nerve conduits possess excellent biocompatibility and exhibit a superior capacity to promote peripheral nerve repair.Significance.The addition of ECM significantly improved the biocompatibility and bioactivity of PLCL, while the PLCL/ECM nerve conduit gained the appropriate mechanical strength from PLCL, which has great potential for clinical repair of peripheral nerve injuries.

The response of Dual-leucine zipper kinase (DLK) to nocodazole: Evidence for a homeostatic cytoskeletal repair mechanism.

Genetic and pharmacological perturbation of the cytoskeleton enhances the regenerative potential of neurons. This response requires Dual-leucine Zipper Kinase (DLK), a neuronal stress sensor that is a central regulator of axon regeneration and degeneration. The damage and repair aspects of this response are reminiscent of other cellular homeostatic systems, suggesting that a cytoskeletal homeostatic response exists. In this study, we propose a framework for understanding DLK mediated neuronal cytoskeletal homeostasis. We demonstrate that low dose nocodazole treatment activates DLK signaling. Activation of DLK signaling results in a DLK-dependent transcriptional signature, which we identify through RNA-seq. This signature includes genes likely to attenuate DLK signaling while simultaneously inducing actin regulating genes. We identify alterations to the cytoskeleton including actin-based morphological changes to the axon. These results are consistent with the model that cytoskeletal disruption in the neuron induces a DLK-dependent homeostatic mechanism, which we term the Cytoskeletal Stress Response (CSR) pathway.

Strategies to enhance the ability of nerve guidance conduits to promote directional nerve growth.

Severely damaged peripheral nerves will regenerate incompletely due to lack of directionality in their regeneration, leading to loss of nerve function. To address this problem, various nerve guidance conduits (NGCs) have been developed to provide guidance for nerve repair. However, their clinical application is still limited, mainly because its effect in promoting nerve repair is not as good as autologous nerve transplantation. Therefore, it is necessary to enhance the ability of NGCs to promote directional nerve growth. Strategies include preparing various directional structures on NGCs to provide contact guidance, and loading various substances on them to provide electrical stimulation or neurotrophic factor concentration gradient to provide directional physical or biological signals.

Oriented Graphene Oxide Scaffold Promotes Nerve Regeneration in vitro and in vivo.

Background: Treating peripheral nerve injuries (PNI) with defects remains challenging in clinical practice. The commercial conduits have shown suboptimal nerve regeneration and functional recovery due to their basic tubular design without electroactive and oriented topographical cues. Purpose: To develop a new scaffold with oriented microstructure and electroactive Graphene oxide (GO) and investigate its' therapeutic effect on nerve regeneration in vitro and in vivo. Methods: This study employed a straightforward approach to co-spin PCL and GO, yielding an oriented hybrid nanofibrous scaffold known as the O-GO/PCL scaffold. The physical and chemical properties of nanofibrous scaffold were tested by scanning electron microscopy (SEM), transmission electron microscope (TEM), tensile test and so on. Primary Schwann cells (SCs) and dorsal root ganglia (DRG) were used to investigate the impact of the newly developed scaffolds on the biological behavior of neural cells in vitro. Transcriptome sequencing (mRNA-seq) was employed to probe the underlying mechanisms of the synergistic effect of electroactive GO and longitudinal topographic guidance on nerve regeneration. Furthermore, the developed O-GO/PCL scaffold was utilized to bridge a 10-mm sciatic nerve defect in rat, aiming to investigate its therapeutic potential for peripheral nerve regeneration in vivo. Results and discussion: The SEM and TEM revealed that the newly developed O-GO/PCL scaffold showed longitudinally oriented microstructure and GO particles were homogenously and uniformly distributed inside the nanofibers. Primary SCs were utilized to assess the biocompatibility of the GO-based scaffold, revealing that negligible cytotoxicity when GO concentration does not exceed 0.5%. In vitro analysis of nerve regeneration demonstrated that axons in the O-GO/PCL group exhibited an average length of 1054.88 ± 161.32 µm, significant longer than those in the other groups (P < 0.05). Moreover, mRNA sequencing results suggested that the O-GO/PCL scaffold could enhance nerve regeneration by upregulating genes associated with neural regeneration, encompassing ion transport, axon guidance and cell-cell interactions. Most importantly, we employed the O-GO/PCL scaffold to repair a 10-mm sciatic nerve defect in rat, resulting in augmented nerve regeneration, myelination, and functional recovery. Conclusion: The O-GO/PCL scaffold with oriented microstructure and electroactive GO represents a promising heral nerve reconstruction.

The Protein Acetylation after Traumatic Spinal Cord Injury: Mechanisms and Therapeutic Opportunities.

Spinal cord injury (SCI) leads to deficits of various normal functions and is difficult to return to a normal state. Histone and non-histone protein acetylation after SCI is well documented and regulates spinal cord plasticity, axonal growth, and sensory axon regeneration. However, our understanding of protein acetylation after SCI is still limited. In this review, we summarize current research on the role of acetylation of histone and non-histone proteins in regulating neuron growth and axonal regeneration in SCI. Furthermore, we discuss inhibitors and activators targeting acetylation-related enzymes, such as α-tubulin acetyltransferase 1 (αTAT1), histone deacetylase 6 (HDAC6), and sirtuin 2 (SIRT2), to provide promising opportunities for recovery from SCI. In conclusion, a comprehensive understanding of protein acetylation and deacetylation in SCI may contribute to the development of SCI treatment.

TREM2 deficiency impairs the energy metabolism of Schwann cells and exacerbates peripheral neurological deficits.

Triggering receptor expressed on myeloid cells-2 (TREM2) has been implicated in susceptibility to neurodegenerative disease. Schwann cells (SCs), the predominant glial cell type in the peripheral nervous system (PNS), play a crucial role in myelination, providing trophic support for neurons and nerve regeneration. However, the function of TREM2 in SCs has not been fully elucidated. Here, we found that TREM2 is expressed in SCs but not in neurons in the PNS. TREM2 deficiency leads to disruption of glycolytic flux and oxidative metabolism in SCs, impairing cell proliferation. The energy crisis caused by TREM2 deficiency triggers mitochondrial damage and autophagy by activating AMPK and impairing PI3K-AKT-mTOR signaling. Combined metabolomic analysis demonstrated that energic substrates and energy metabolic pathways were significantly impaired in TREM2-deficient SCs. Moreover, TREM2 deficiency impairs energy metabolism and axonal growth in sciatic nerve, accompanied by exacerbation of neurological deficits and suppression of nerve regeneration in a mouse model of acute motor axonal neuropathy. These results indicate that TREM2 is a critical regulator of energy metabolism in SCs and exerts neuroprotective effects on peripheral neuropathy. TREM2 deficiency impairs glycolysis and oxidative metabolism in Schwann cells, resulting in compromised cell proliferation. The energy crisis caused by TREM2 deficiency induces mitochondrial damage and autophagy by activating AMPK and impairing PI3K-AKT-mTOR signaling. Moreover, TREM2 deficiency disrupts the energy metabolism of the sciatic nerve and impairs support for axonal regeneration, accompanied by exacerbation of neurological deficits and suppression of nerve regeneration in a mouse model of acute motor axonal neuropathy (by FigDraw).

Reduced Graphene Oxide Fibers Combined with Electrical Stimulation Promote Peripheral Nerve Regeneration.

Background: The treatment of long-gap peripheral nerve injury (PNI) is still a substantial clinical problem. Graphene-based scaffolds possess extracellular matrix (ECM) characteristic and can conduct electrical signals, therefore have been investigated for repairing PNI. Combined with electrical stimulation (ES), a well performance should be expected. We aimed to determine the effects of reduced graphene oxide fibers (rGOFs) combined with ES on PNI repair in vivo. Methods: rGOFs were prepared by one-step dimensionally confined hydrothermal strategy (DCH). Surface characteristics, chemical compositions, electrical and mechanical properties of the samples were characterized. The biocompatibility of the rGOFs were systematically explored both in vitro and in vivo. Total of 54 Sprague-Dawley (SD) rats were randomized into 6 experimental groups: a silicone conduit (S), S+ES, S+rGOFs-filled conduit (SGC), SGC+ES, nerve autograft, and sham groups for a 10-mm sciatic defect. Functional and histological recovery of the regenerated sciatic nerve at 12 weeks after surgery in each group of SD rats were evaluated. Results: rGOFs exhibited aligned micro- and nano-channels with excellent mechanical and electrical properties. They are biocompatible in vitro and in vivo. All 6 groups exhibited PNI repair outcomes in view of neurological and morphological recovery. The SGC+ES group achieved similar therapeutic effects as nerve autograft group (P > 0.05), significantly outperformed other treatment groups. Immunohistochemical analysis showed that the expression of proteins related to axonal regeneration and angiogenesis were relatively higher in the SGC+ES. Conclusion: The rGOFs had good biocompatibility combined with excellent electrical and mechanical properties. Combined with ES, the rGOFs provided superior motor nerve recovery for a 10-mm nerve gap in a murine acute transection injury model, indicating its excellent repairing ability. That the similar therapeutic effects as autologous nerve transplantation make us believe this method is a promising way to treat peripheral nerve defects, which is expected to guide clinical practice in the future.

Differential Protein Expression between the Motor and Sensory Fascicles in Rat Femoral Nerve Injury.

BACKGROUND AND AIMS: It is important to distinguish between motor and sensory fascicles of the peripheral nerves for nerve alignment in surgery. However, there are no biomarkers currently available for effective identification of motor or sensory fascicles. The objective of this study was to identify differentially expressed proteins between motor and sensory fascicles of rats in response to injury. SETTINGS AND DESIGN: The study was carried out using a rat femoral nerve injury model. MATERIALS: A proteomic analysis was performed to detect differential protein expression using samples of bilateral motor and sensory branches of intact and injured rat femoral nerves through fluorescent two-dimensional difference gel electrophoresis and matrix-assisted laser desorption ionization-time of flight mass spectrometry. STATISTICAL ANALYSIS: Chi-square tests and t-tests were performed for comparison between motor or sensory nerve groups. RESULTS: The data identified six proteins that were differentially expressed between motor and sensory fascicles (>1.5-fold, P < 0.05), including apolipoprotein E, neurofilament light polypepticle, TEC kinase, serine protease inhibitor A3N, peroxiredoxin-2, and TPM1. The proteomic results were consistent with the mRNA expression levels of these genes as determined by quantitative reverse transcription polymerase chain reaction. CONCLUSIONS: Our study suggests that these proteins may play roles in nerve regeneration and repair. Importantly, apolipoprotein E and Serpina3n may serve as specific biomarkers for distinguishing motor and sensory fascicles of the peripheral nerves for nerve alignment in surgery.

Skin Reinnervation by Collateral Sprouting Following Spared Nerve Injury in Mice.

Following peripheral nerve injury, denervated tissues can be reinnervated via regeneration of injured neurons or collateral sprouting of neighboring uninjured afferents into denervated territory. While there has been substantial focus on mechanisms underlying regeneration, collateral sprouting has received less attention. Here, we used immunohistochemistry and genetic neuronal labeling to define the subtype specificity of sprouting-mediated reinnervation of plantar hindpaw skin in the mouse spared nerve injury (SNI) model, in which productive regeneration cannot occur. Following initial loss of cutaneous afferents in the tibial nerve territory, we observed progressive centripetal reinnervation by multiple subtypes of neighboring uninjured fibers into denervated glabrous and hairy plantar skin of male mice. In addition to dermal reinnervation, CGRP-expressing peptidergic fibers slowly but continuously repopulated denervated epidermis, Interestingly, GFRα2-expressing nonpeptidergic fibers exhibited a transient burst of epidermal reinnervation, followed by a trend towards regression. Presumptive sympathetic nerve fibers also sprouted into denervated territory, as did a population of myelinated TrkC lineage fibers, though the latter did so inefficiently. Conversely, rapidly adapting Aß fiber and C fiber low threshold mechanoreceptor (LTMR) subtypes failed to exhibit convincing sprouting up to 8 weeks after nerve injury in males or females. Optogenetics and behavioral assays in male mice further demonstrated the functionality of collaterally sprouted fibers in hairy plantar skin with restoration of punctate mechanosensation without hypersensitivity. Our findings advance understanding of differential collateral sprouting among sensory neuron subpopulations and may guide strategies to promote the progression of sensory recovery or limit maladaptive sensory phenomena after peripheral nerve injury.

[Research progress in functional regeneration methods and mechanisms of taste buds].

Gustation is one of the most important human senses. Taste dysfunctions, which may be due to aging, tongue cancer surgery, radiotherapy and chemotherapy, affect life quality. That is why the need for taste bud regeneration has received more attention. At present, research on development and renewal of taste cells provides a basis for taste bud regeneration; molecular mechanisms related to taste bud regeneration are being continuously uncoverd, aiding in the identification of more accurate targets for therapy. New methods such as nerve regeneration, tissue engineering, and cytokine therapy have emerged. The author reviews the mechanism and the latest methods of taste bud regeneration of lingual epithelium, aiming to open new horizions for the prevention and treatment of gustatory diseases, and provide theoretical references for its regeneration.

Development of new nanofibrous nerve conduits by PCL-Chitosan-Hyaluronic acid containing Piracetam-Vitamin B12 for sciatic nerve: A rat model.

Peripheral nerve injury is a critical condition that can disrupt nerve functions. Despite the progress in engineering artificial nerve guidance conduits (NGCs), nerve regeneration remains challenging. Here, we developed new nanofibrous NGCs using polycaprolactone (PCL) and chitosan (CH) containing piracetam (PIR)/vitamin B12(VITB12) with an electrospinning method. The lumen of NGCs was coated by hyaluronic acid (HA) to promote regeneration in sciatic nerve injury. The NGCs were characterized via Scanning Electron Microscopy (SEM), Fourier transform infrared (FTIR), tensile, swelling, contact angle, degradation, and drug release tests. Neuronal precursor cell line (PCL12 cell) and rat mesenchymal stem cells derived from bone marrow (MSCs) were seeded on the nanofibrous conduits. After that, the biocompatibility of the NGCs was evaluated by the 2,5-diphenyl-2H-tetrazolium bromide (MTT) assay, 4',6-diamidino-2-phenylindole (DAPI) staining, and SEM images. The SEM demonstrated that PCL/CH/PIR/VITB12 NGCs had nonaligned, interconnected, smooth fibers. The mechanical properties of these NGCs were similar to rat sciatic nerve. These conduits had an appropriate swelling and degradation rate. The In Vitro studies exhibited favorable biocompatibility of the PCL/CH/PIR/VITB12 NGCs towards PC12 cells and MSCs. The in vitro studies exhibited favorable biocompatibility of the PCL/CH/PIR/VIT B12 NGCs towards MSCs and PC12 cells. To analyze functional efficacy, NGCs were implanted into a 10 mm Wistar rat sciatic nerve gap and bridged the proximal and distal stump of the defect. After three months, the results of sciatic functional index (55.3 ± 1.8), hot plate latency test (5.6 ± 0.5 s), gastrocnemius muscle wet weight-loss (38.57 ± 1.6 %) and histopathological examination using hematoxylin-eosin (H&E) /toluidine blue/ Anti-Neurofilament (NF200) staining demonstrated that the produced conduit recovered motor and sensory functions and had comparable nerve regeneration compared to the autograft that can be as the gold standard to bridge the nerve gaps.

Targeting Vasohibins to Promote Axon Regeneration.

Treatments accelerating axon regeneration in the nervous system are still clinically unavailable. However, parthenolide promotes adult sensory neurons' axon growth in culture by inhibiting microtubule detyrosination. Here, we show that overexpression of vasohibins increases microtubule detyrosination in growth cones and compromises growth in culture and in vivo. Moreover, overexpression of these proteins increases the required parthenolide concentrations to promote axon regeneration. At the same time, the partial knockdown of endogenous vasohibins or their enhancer SVBP in neurons facilitates axon growth, verifying them as pharmacological targets for promoting axon growth. In vivo, repeated intravenous application of parthenolide or its prodrug di-methyl-amino-parthenolide (DMAPT) markedly facilitates the regeneration of sensory, motor, and sympathetic axons in injured murine and rat nerves, leading to acceleration of functional recovery. Moreover, orally applied DMAPT was similarly effective in promoting nerve regeneration. Thus, pharmacological inhibition of vasohibins facilitates axon regeneration in different species and nerves, making parthenolide and DMAPT the first promising drugs for curing nerve injury.

Boundary cap neural crest stem cells promote angiogenesis after transplantation to avulsed dorsal roots in mice and induce migration of endothelial cells in 3D printed scaffolds.

Dorsal root avulsion injuries lead to loss of sensation and to reorganization of blood vessels (BVs) in the injured area. The inability of injured sensory axons to re-enter the spinal cord results in permanent loss of sensation, and often also leads to the development of neuropathic pain. Approaches that restore connection between peripheral sensory axons and their CNS targets are thus urgently need. Previous research has shown that sensory axons from peripherally grafted human sensory neurons are able to enter the spinal cord by growing along BVs which penetrate the CNS from the spinal cord surface. In this study we analysed the distribution of BVs after avulsion injury and how their pattern is affected by implantation at the injury site of boundary cap neural crest stem cells (bNCSCs), a transient cluster of cells, which are located at the boundary between the spinal cord and peripheral nervous system and assist the growth of sensory axons from periphery into the spinal cord during development. The superficial dorsal spinal cord vasculature was examined using intravital microscopy and intravascular BV labelling. bNCSC transplantation increased vascular volume in a non-dose responsive manner, whereas dorsal root avulsion alone did not decrease the vascular volume. To determine whether bNCSC are endowed with angiogenic properties we prepared 3D printed scaffolds, containing bNCSCs together with rings prepared from mouse aorta. We show that bNCSC do induce migration and assembly of endothelial cells in this system. These findings suggest that bNCSC transplant can promote vascularization in vivo and contribute to BV formation in 3D printed scaffolds.

Neutrophil-inflicted vasculature damage suppresses immune-mediated optic nerve regeneration.

In adult mammals, injured retinal ganglion cells (RGCs) fail to spontaneously regrow severed axons, resulting in permanent visual deficits. Robust axon growth, however, is observed after intra-ocular injection of particulate ß-glucan isolated from yeast. Blood-borne myeloid cells rapidly respond to ß-glucan, releasing numerous pro-regenerative factors. Unfortunately, the pro-regenerative effects are undermined by retinal damage inflicted by an overactive immune system. Here, we demonstrate that protection of the inflamed vasculature promotes immune-mediated RGC regeneration. In the absence of microglia, leakiness of the blood-retina barrier increases, pro-inflammatory neutrophils are elevated, and RGC regeneration is reduced. Functional ablation of the complement receptor 3 (CD11b/integrin-αM), but not the complement components C1q-/- or C3-/-, reduces ocular inflammation, protects the blood-retina barrier, and enhances RGC regeneration. Selective targeting of neutrophils with anti-Ly6G does not increase axogenic neutrophils but protects the blood-retina barrier and enhances RGC regeneration. Together, these findings reveal that protection of the inflamed vasculature promotes neuronal regeneration.

Ca2+/Calmodulin-Dependent Protein Kinase II Enhances Retinal Ganglion Cell Survival But Suppresses Axon Regeneration after Optic Nerve Injury.

Neuroprotection after injury or in neurodegenerative disease remains a major goal for basic and translational neuroscience. Retinal ganglion cells (RGCs), the projection neurons of the eye, degenerate in optic neuropathies after axon injury, and there are no clinical therapies to prevent their loss or restore their connectivity to targets in the brain. Here we demonstrate a profound neuroprotective effect of the exogenous expression of various Ca2+/calmodulin-dependent protein kinase II (CaMKII) isoforms in mice. A dramatic increase in RGC survival following the optic nerve trauma was elicited by the expression of constitutively active variants of multiple CaMKII isoforms in RGCs using adeno-associated viral (AAV) vectors across a 100-fold range of AAV dosing in vivo. Despite this neuroprotection, however, short-distance RGC axon sprouting was suppressed by CaMKII, and long-distance axon regeneration elicited by several pro-axon growth treatments was likewise inhibited even as CaMKII further enhanced RGC survival. Notably, in a dose-escalation study, AAV-expressed CaMKII was more potent for axon growth suppression than the promotion of survival. That diffuse overexpression of constitutively active CaMKII strongly promotes RGC survival after axon injury may be clinically valuable for neuroprotection per se. However, the associated strong suppression of the optic nerve axon regeneration demonstrates the need for understanding the intracellular domain- and target-specific CaMKII activities to the development of CaMKII signaling pathway-directed strategies for the treatment of optic neuropathies.

Cell Heterogeneity and Variability in Peripheral Nerve after Injury.

With the development of single-cell sequencing technology, the cellular composition of more and more tissues is being elucidated. As the whole nervous system has been extensively studied, the cellular composition of the peripheral nerve has gradually been revealed. By summarizing the current sequencing data, we compile the heterogeneities of cells that have been reported in the peripheral nerves, mainly the sciatic nerve. The cellular variability of Schwann cells, fibroblasts, immune cells, and endothelial cells during development and disease has been discussed in this review. The discovery of the architecture of peripheral nerves after injury benefits the understanding of cellular complexity in the nervous system, as well as the construction of tissue engineering nerves for nerve repair and axon regeneration.

Long-Acting Heterodimeric Paclitaxel-Idebenone Prodrug-Based Nanomedicine Promotes Functional Recovery after Spinal Cord Injury.

After spinal cord injury (SCI), successive systemic administration of microtubule-stabilizing agents has been shown to promote axon regeneration. However, this approach is limited by poor drug bioavailability, especially given the rapid restoration of the blood-spinal cord barrier. There is a pressing need for long-acting formulations of microtubule-stabilizing agents in treating SCI. Here, we conjugated the antioxidant idebenone with microtubule-stabilizing paclitaxel to create a heterodimeric paclitaxel-idebenone prodrug via an acid-activatable, self-immolative ketal linker and then fabricated it into chondroitin sulfate proteoglycan-binding nanomedicine, enabling drug retention within the spinal cord for at least 2 weeks and notable enhancement in hindlimb motor function and axon regeneration after a single intraspinal administration. Additional investigations uncovered that idebenone can suppress the activation of microglia and neuronal ferroptosis, thereby amplifying the therapeutic effect of paclitaxel. This prodrug-based nanomedicine simultaneously accomplishes neuroprotection and axon regeneration, offering a promising therapeutic strategy for SCI.

Effect of Long-term Peripheral Nerve Stimulation on Neuro-Muscular Complex Morphologic Recovery in Experiment / Efecto de la Estimulación del Nervio Periférico a Largo Plazo sobre la Recuperación Morfológica del Complejo Neuromuscular en Experimento

SUMMARY: Peripheral nerve injury is an extremely important medical and socio-economic problem. It is far from a solution, despite on rapid development of technologies. To study the effect of long-term electrical stimulation of peripheral nerves, we used a domestically produced electrical stimulation system, which is approved for clinical use. The study was performed on 28 rabbits. Control of regeneration was carried out after 3 month with morphologic techniques. The use of long-term electrostimulation technology leads to an improvement in the results of the recovery of the nerve trunk after an injury, both directly at the site of damage, when stimulation begins in the early period, and indirectly, after the nerve fibers reach the effector muscle.

La lesión de los nervios periféricos es un problema médico y socioeconómico extremadamente importante. Sin embargo, y a pesar del rápido desarrollo de las tecnologías, aún no tiene solución. Para estudiar el efecto de la estimulación eléctrica a largo plazo de los nervios periféricos, utilizamos un sistema de estimulación eléctrica de producción nacional, que está aprobado para uso clínico. El estudio se realizó en 28 conejos. El control de la regeneración se realizó a los 3 meses con técnicas morfológicas. El uso de tecnología de electro estimulación a largo plazo conduce a una mejora en los resultados de la recuperación del tronco nervioso después de una lesión, tanto directamente en el lugar del daño, cuando la estimulación comienza en el período temprano, como indirectamente, después de que las fibras nerviosas alcanzan el músculo efector.

[Experimental study of M2 microglia transplantation promoting spinal cord injury repair in mice].

Objective: To investigate the effect of M2 microglia (M2-MG) transplantation on spinal cord injury (SCI) repair in mice. Methods: Primary MG were obtained from the cerebral cortex of 15 C57BL/6 mice born 2-3 days old by pancreatic enzyme digestion and identified by immunofluorescence staining of Iba1. Then the primary MG were co-cultured with interleukin 4 for 48 hours (experimental group) to induce into M2 phenotype and identified by immunofluorescence staining of Arginase 1 (Arg-1) and Iba1. The normal MG were harvested as control (control group). The dorsal root ganglion (DRG) of 5 C57BL/6 mice born 1 week old were co-cultured with M2-MG for 5 days to observe the axon length, the DRG alone was used as control. Forty-two 6-week-old female C57BL/6 mice were randomly divided into sham group ( n=6), SCI group ( n=18), and SCI+M2-MG group ( n=18). In sham group, only the laminae of T 10 level were removed; SCI group and SCI+M2-MG group underwent SCI modeling, and SCI+M2-MG group was simultaneously injected with M2-MG. The survival of mice in each group was observed after operation. At immediate (0), 3, 7, 14, 21, and 28 days after operation, the motor function of mice was evaluated by Basso Mouse Scale (BMS) score, and the gait was evaluated by footprint experiment at 28 days. The spinal cord tissue was taken after operation for immunofluorescence staining, in which glial fibrillary acidic protein (GFAP) staining at 7, 14, and 28 days was used to observe the injured area of the spinal cord, neuronal nuclei antigen staining at 28 days was used to observe the survival of neurons, and GFAP/C3 double staining at 7 and 14 days was used to observe the changes in the number of A1 astrocytes. Results: The purity of MG in vitro reached 90%, and the most of the cells were polarized into M2 phenotype identified by Arg-1 immunofluorescence staining. M2-MG promoted the axon growth when co-cultured with DRGs in vitro ( P<0.05). All groups of mice survived until the experiment was completed. The hind limb motor function of SCI group and SCI+M2-MG group gradually recovered over time. Among them, the SCI+M2-MG group had significantly higher BMS scores than the SCI group at 21 and 28 days ( P<0.05), and the dragging gait significantly improved at 28 days, but it did not reach the level of the sham group. Immunofluorescence staining showed that compared with the SCI group, the SCI+M2-MG group had a smaller injury area at 7, 14, and 28 days, an increase in neuronal survival at 28 days, and a decrease in the number of A1 astrocytes at 7 and 14 days, with significant differences ( P<0.05). Conclusion: M2-MG transplantation improves the motor function of the hind limbs of SCI mice by promoting neuron survival and axon regeneration. This neuroprotective effect is related to the inhibition of A1 astrocytes polarization.

The involvement of Neuregulin-1 in the process of facial nerve injury repair through the utilization of dental pulp stem cells.

BACKGROUND: Facial nerve injury often results in poor prognosis due to the challenging process of nerve regeneration. Neuregulin-1, a human calmodulin, is under investigation in this study for its impact on the reparative capabilities of Dental Pulp Stem Cells (DPSCs) in facial nerve injury. METHODS: Lentivirus was used to transfect and construct Neuregulin-1 overexpressed DPSCs. Various techniques assessed the effects of Neuregulin-1: osteogenic induction, lipid induction, Reverse Transcription Polymerase Chain Reaction, Western Blot, Cell Counting Kit-8 assay, wound healing, immunofluorescence, Phalloidin staining, nerve stem action potential, Hematoxylin-eosin staining, transmission electron microscopy, and immunohistochemistry. RESULTS: Neuregulin-1 effectively enhanced the proliferation, migration, and cytoskeletal rearrangement of DPSCs, while simultaneously suppressing the expression of Ras homolog gene family member A (RhoA) and Microfilament actin (F-actin). These changes facilitated the neural differentiation of DPSCs. Additionally, in vivo experiments showed that Neuregulin-1 expedited the restoration of action potential in the facial nerve trunk, increased the thickness of the myelin sheath, and stimulated axon regeneration. CONCLUSION: Neuregulin-1 has the capability to facilitate the repair of facial nerve injuries by promoting the regenerative capacity of DPSCs. Thus, Neuregulin-1 is a significant potential gene in the reparative processes of nerve damage.