Rat Nerve Tissues Research Articles

Fabrication of Anatomically Equivalent Pectin-Based Multifilament Nerve Conduits.

Reuniting denuded nerve ends after a long segmental peripheral nerve defect is challenging due to delayed axonal regeneration and incomplete, nonspecific reinnervation, as conventional hollow nerve guides fail to ensure proper fascicular complementation and obstruct axonal guidance across the defects. This study focuses on fabricating multifilament conduits using a plant-derived anionic polysaccharide, pectin, where the abundant availability of carboxylate (COO-) functional groups in pectin facilitates instantaneous sol-gel transition upon interaction with divalent cations. Despite their advantages, pectin hydrogels encounter structural instability under physiological conditions. Hence, pectin is conjugated with light-sensitive methacrylate residues (49.8% methacrylation) to overcome these issues, enabling the fabrication of dual cross-linked multifilament nerve conduits through an ionic interaction-driven, template-free 3D wet writing process, followed by photo-cross-linking at 525 nm. The anatomical equivalence including peri-, epi-, and endoneurium structures of the customized multifilament conduits was confirmed through scanning electron micrographs and micro-CT analysis of rat and goat sciatic nerve tissues. Furthermore, the fabricated multifilament nerve conduits demonstrated cytocompatibility and promoted the expression of neuron-specific intermediate filament protein (NF-200) in PC12 cells and neurite outgrowth of 16.90 ± 1.82 μm on day 14. Micro-CT imaging of an anastomosed native goat sciatic nerve with an 8-filament conduit demonstrated precise fascicular complementation in an ex vivo interpositional goat model. This approach not only eliminates the need for a suture-intensive ligation process but also highlights the customizability of multifilament conduits to meet patient- and injury-specific needs.

Effect of New Antidiabetics on Steatosis in Nerve Tissues and Nerve Conduction Velocity: Possible Role of Nerve Growth Factor (NGF)/Synaptophysin and Nrf2/HO-1 Pathways.

The current study aims to investigate the impact of the GLP1 analog (semaglutide) and SGLT2 inhibitor (dapagliflozin) on nerve functions, morphology, and the underlying mechanisms involving nerve growth factor (NGF)/synaptophysin and Nrf2/HO-1 pathways in obese rats. Forty male Sprague Dawley rats, aged six to eight weeks, were classified into five groups; normal group (high-fat diet {HFD} for 12 weeks, metformin group (HFD for 12 weeks + metformin in last four weeks), dapagliflozin group (HFD for 12 weeks +dapagliflozin in last four weeks, semaglutide group (HFD for 12 weeks + semaglutide in last four weeks). At the end of the experiment, the sciatic nerve was collected for nerve conduction study, oxidative stress marker (malondialdehyde, i.e., MDA), real-time polymerase chain reaction (PCR) study (for HO-1 and Nrf2), oil red O staining, electron microscopic examination and immunohistochemistry for NGF and synaptophysin. The HFD group showed a significant rise in blood glucose, serum lipids, homeostatic model assessment (HOMA) index, lipid deposition in nerve tissues, and lipid peroxidation (MDA) in nerve tissues with significant attenuation in nerve conduction velocity (NCV), the expression of Nrf2 and HO-1 genes and significant attenuation in area stained with NGF and synaptophysin. On the other hand, pretreatment with either dapagliflozin or semaglutide led to considerable enhancement in the deteriorated serum and nerve tissue parameters and reversed the pathological changes. New antidiabetic drugs like SGLT2 inhibitors (more powerful) and GLP1 analog might have neuroprotective beneficial effects besides controlling the glycemic state in obese rats. This effect may result from reduced oxidative stress and increased Nrf2 levels, HO-1, synaptophysin, and NGF in the nerve tissues of obese rats.

LncRNA GAS5 modulates Schwann cell function and enhances facial nerve injury repair via the miR-138-5p/CXCL12 axis.

Facial nerve is an integral part of peripheral nerve. Schwann cells are important microglia involved in the repair and regulation of facial nerve injury. LncRNA growth arrest‑specific transcript 5 (GAS5) is involved in the behavioral regulation of Schwann cell and the regeneration of peripheral nervous system. However, there is little research about the effect of GAS5 on the repair of facial nerve injury (FNI) by regulating Schwann cells. This study aimed to investigate the role of GAS5 in Schwann cell function and FNI repair, focusing on the miR-138-5p/CXCL12 axis. Hematoxylin and eosin staining, Luxol fast blue staining, transmission electron microscope, and immunofluorescence (IF) experiments were used to verify the effect of GAS5 on FNI rats. Reverse transcription real-time polymerase chain reaction was performed to detect GAS5, miR-138-5p, and C-X-C motif chemokine ligand 12 (CXCL12) mRNA expression. IF staining was used to detect the inflorescence of S100 calcium binding protein B (S100β), SRY-box transcription factor 10 (SOX10), and tubulin beta 3 class III (β-Tubulin III). Glial fibrillary acidic protein (GFAP), nerve growth factor receptor (NGFR), S100β, brain derived neurotrophic factor (BDNF), ciliary neurotrophic factor (CNTF), and CXCL12 proteins were detected using western blot. The5-bromo-2'-deoxyuridine staining, Transwell, and flow cytometry assays were conducted to detect Schwann cell function. Dual-luciferase, RNA immunoprecipitation, and RNA pulldown assaywere used to identify the interaction among GAS5, miR-138-5p, and CXCL12. Results found thatGAS5 was downregulated in facial nerve tissues of FNI rats. Overexpressed GAS5 decreased facial grading, inhibited demyelination, and promoted proliferation, migration, and suppressed apoptosis of Schwann cells. Mechanistically, GAS5 was a sponge of miR-138-5p and positively regulated CXCL12 expression. GAS5 inhibition repressed CXCL12 expression and decreased cell proliferationand migration, increased apoptosis rate of Schwann cells by sponging miR-138-5p. In conclusion,overexpression of GAS5 accelerates facial nerve repair in FNI rats by regulating miR-138-5p/CXCL12 axis.

LKB1 and CRMP1 cooperatively promote the repair of the sciatic nerve injury.

After peripheral nervous system injury, Schwann cells (SCs) can repair axons by providing a growth-promoting microenvironment. The aim of this study is to explore the effects and mechanisms of LKB1 and CRMP1 on the repair of sciatic nerve injury (SNI). The expressions of LKB1 and CRMP1 were changed in rats with SNI from 12h to 4 weeks by hematoxylin-eosin staining, RT-PCR assay, immunohistochemical staining, and western blotting. Immunofluorescence results show that LKB1 and CRMP1 are co-localized in the regenerated axons of the sciatic nerve tissue of SNI rats. Co-immunoprecipitation indicates that LKB1 interacts with CRMP1. LKB1 interference suppresses the phosphorylation level of CRMP1. Overexpression of LKB1 and CRMP1 promotes the invasion and migration of SCs, and nerve cell protuberance extends. The structure of the myelin sheath in the sciatic nerve of the model group was found to be loose and disordered. Rats in the model group had higher pain thresholds and heat sensitivity response times than those in the control group. Nerve conduction velocity, the latency of action potential, and the peak value of compound muscle action potential in the SNI group were significantly lower than those in the control group, and the muscle atrophy was severe. Overexpression of LKB1 may significantly improve the above conditions. However, the function of LKB1 to improve SNI is abolished by the interference of CRMP1. In summary, the interaction between LKB1 and CRMP promotes the migration and differentiation of SCs and the extension of neurons, thereby improving the repair of nerve injury.

Regenerative capacity of neural tissue scales with changes in tissue mechanics post injury

Spinal cord injuries have devastating consequences for humans, as mammalian neurons of the central nervous system (CNS) cannot regenerate. In the peripheral nervous system (PNS), however, neurons may regenerate to restore lost function following injury. While mammalian CNS tissue softens after injury, how PNS tissue mechanics changes in response to mechanical trauma is currently poorly understood. Here we characterised mechanical rat nerve tissue properties before and after in vivo crush and transection injuries using atomic force microscopy-based indentation measurements. Unlike CNS tissue, PNS tissue significantly stiffened after both types of tissue damage. This nerve tissue stiffening strongly correlated with an increase in collagen I levels. Schwann cells, which crucially support PNS regeneration, became more motile and proliferative on stiffer substrates in vitro, suggesting that changes in tissue stiffness may play a key role in facilitating or impeding nervous system regeneration.

Effect of Physical Training on the Morphology of Parasympathetic Atrial Ganglia after Unilateral Vagotomy in Rats.

Cardiac denervation is a serious problem in a number of patients, including patients after heart transplantation. The status of the parasympathetic ganglia after crossing the preganglionic fibers of the vagus nerve has not been enough studied. The aim of our study was to assess the effect of physical training on the morphological parameters of the parasympathetic atrial ganglia and autonomic regulation of heart rate after right- and left-sided vagotomy in rats. Morphometric characteristics of the right atrial ganglia were evaluated using an immunohistochemical method after a study that included a three-time assessment of heart rate variability. It was found that right-sided vagotomy leads to both an increase in the volume of ganglion and autonomic dysfunction. No significant change in the number of nerve cells was found in animals with false and left-sided vagotomy while maintaining preganglionic innervation after the physical training, whereas exercises led to a decrease in the volume of nerve tissue of rats with right-sided denervation. It was also found that in animals with preserved vagal innervation, the volume of atrial ganglion tissue correlates with overall heart rate variability and a normalized parasympathetic component. Therefore, a positive effect from regular physical activity on parasympathetic regulation can be expected only if preganglionic vagal influence is preserved.

Isolation, culture and inflammatory characteristics of astrocytes in aged rats nervous tissue

Objective: To establish an optimized method for the isolation and purification of astrocytes from the neural tissues of young and aged rats. Then, the morphological and functional differences of astrocytes between young and aged rats were compared to explore the functional changes of astrocytes after aging and its possible mechanism in the aging process. Methods: Young (2 months old) and aged (20 months old) SD rats were used. Astrocytes in brain and spinal cord tissue were purified by 50% - 35% percoll density gradient centrifugation. Each group of cells was set up with three duplicate wells. After 72 h of culture, Glial fibrillary acidic protein (GFAP) which was astrocyte specific marker were detected by immunofluorescence to evaluate the morphological characteristics. Cell senescence markers (p16 and p21) and β- Galactosidase were detected by qPCR and staining respectively. The expressions of pro-inflammatory cytokines (IL-1β, TNF-α) and anti-inflammatory cytokines were detected by qPCR. Results: Using 50%-35% percoll gradient separation, astrocytes were obtained with large number, good activity and purity of more than 95%, which could be used in subsequent experiments. Compared with the astrocytes in the nerve tissue of young rats, the astrocytes in the nervous tissue of the aged rats had fewer protrusions and tended to be activated in cell morphology; the positive rate of β -galactosidase staining was increased significantly and the expressions of p16 and p21 were increased (P<0.01). The expressions of pro-inflammatory cytokines (IL-1β, TNF-α) were increased (P<0.05), and the expression of anti-inflammatory cytokine (IL-10) was decreased (P<0.05) in astrocytes of the aged rats nervous tissue. Conclusion: The percoll gradient of 50% - 35% could be used as a method for separation, purification and primary culture of astrocytes. With the increase of age, astrocytes undergo cellular senescence, showing a pro-inflammatory phenotype, promoting inflammaging of the nervous system, which may be one of the mechanisms of nervous system aging and neurodegenerative diseases.

Identification of prophylactic drugs for oxaliplatin-induced peripheral neuropathy using big data

BackgroundDrug repositioning is a cost-effective method to identify novel disease indications for approved drugs; it requires a shorter developmental period than conventional drug discovery methods. We aimed to identify prophylactic drugs for oxaliplatin-induced peripheral neuropathy by drug repositioning using data from large-scale medical information and life science information databases. MethodsHerein, we analyzed the reported data between 2007 and 2017 retrieved from the FDA’s database of spontaneous adverse event reports (FAERS) and the LINCS database provided by the National Institute of Health. The efficacy of the drug candidates for oxaliplatin-induced peripheral neuropathy obtained from the database analysis was examined using a rat model of peripheral neuropathy. Additionally, we compared the incidence of peripheral neuropathy in patients who received oxaliplatin at the Tokushima University Hospital, Japan. The effects of statins on the animal model were examined in six-week-old male Sprague–Dawley rats and seven or eight-week-old male BALB/C mice. Retrospective medical chart review included clinical data from Tokushima University Hospital from April 2009 to March 2018. ResultsSimvastatin, indicated for dyslipidemia, significantly reduced the severity of peripheral neuropathy and oxaliplatin-induced hyperalgesia. In the nerve tissue of model rats, the mRNA expression of Gstm1 increased with statin administration. A retrospective medical chart review using clinical data revealed that the incidence of peripheral neuropathy decreased with statin use. Conclusion and relevanceThus, drug repositioning using data from large-scale basic and clinical databases enables the discovery of new indications for approved drugs with a high probability of success.

Effect of Tacrolimus on Peripheral Nerve Regeneration in Allograft Transplantation: A Light and Electron Microscopic Study.

Peripheral nerve injuries are common in Europe; however, the treatment techniques may lead to disabilities. This study aimed to evaluate the effect of tacrolimus use on the capacity of the epineural sheath graft to improve its regeneration quality in rat sciatic nerves as a treatment option for nerve injuries. In the experimental process, 30 male Sprague Dawley were used as recipients and 10 Wistar rats were used as donors. Under anesthesia, all rats were operated on to resect the sciatic nerve. The nerve tissue of Wistar rats was used as allograft. In the autograft group, the resected nerve was reversed and sutured, resulting in an epineural sheath graft. For the allograft groups, rats were randomly divided into 2 groups as the tacrolimus-treated group and the nontreated group after allograft transplant. Tacrolimus was administered intramuscularly at 0.1 mg/kg daily for 12 weeks. After the treatment period, rats were killed and evaluated histomorphologically with light and electron microscopy. Histological examination showed no remarkable differences between different regions of the sciatic nerves (distal, middle, and proximal). The axonal density was decreased in the allograft groups compared with the autograft group (P < .001). Results showed that the number of mast cells was increased in the allograft group without tacrolimus treatment (P < .05). Similarly, there was a mild increase in mast cell count in the tacrolimus-treated allograft group. Our results showed that tacrolimus use in rats with implanted epineural nerve sheath supported recovery in terms of morphological and physiological regeneration of the nerve.

Silymarin alleviates docetaxel-induced central and peripheral neurotoxicity by reducing oxidative stress, inflammation and apoptosis in rats.

Docetaxel (DTX) is a chemotherapeutic agent used in the treatment of various malignancies but is often associated with central and peripheral neurotoxicity. The aim of this study was to investigate the neuroprotective effect of silymarin (SLM) against DTX-induced central and peripheral neurotoxicities in rats. Rats received 25 and 50mg/kg body weight SLM orally for seven consecutive days after receiving a single injection of 30mg/kg body weight DTX on the first day. SLM significantly decreased brain lipid peroxidation level and ameliorated brain glutathione (GSH), superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPx) activities in DTX-administered rats. SLM attenuated levels of nuclear factor kappa B (NF-κB), tumor necrosis factor-α (TNF-α), glial fibrillary acidic protein (GFAP) and activity of p38α mitogen-activated protein kinase (p38α MAPK) whereas caused an increase in levels of neural cell adhesion molecule (NCAM) in the brain and sciatic nerve of DTX-induced rats. In addition, SLM improved the histological structure of the brain and sciatic nerve tissues and decreased the expression of c-Jun N-terminal kinase (JNK) in the sciatic nerve whereas increased cyclic AMP response element binding protein (CREB) expression in the brain induced by DTX. Additionally, SLM markedly up-regulated the expression of nuclear factor erythroid 2-related factor 2 (Nrf2), heme oxygenase-1 (HO-1) and B-cell lymphoma-2 (Bcl-2) and downregulated the expression of Bcl-2 associatedXprotein (Bax) in the brain and sciatic nerve tissues of DTX-induced rats. Our results show that SLM can protect DTX-induced brain and sciatic nerve injuries by enhancing the antioxidant defense system and suppressing apoptosis and inflammation.

Description and validation of a simple histological nerve tissue scoring system for nerve allografts.

In nerve allograft development, currently used subjective histological scoring systems to evaluate nerve structure in experimental studies are not uniform and have not been validated. The aim of this study was to describe and validate a simple, fast and inexpensive method to compare structural properties of nerve allografts on a histological level. A total of 113 histological sections of rat (sciatic nerves) and human peripheral nerve segments (thoracodorsal and long thoracic nerve for motor and sural for sensory nerve) treated with various decellularization protocols were analyzed. Slides were stained with Hematoxylin & Eosin (H&E), Toluidine Blue and Laminin. A subjective scoring system using a 5-point scale was used to score each nerve section by four independent researchers. For validation of this score both inter- and intra-rater reliability were calculated using the Intraclass Correlation Coefficient (ICC). Overall, an excellent correlation between the different observers was found for the Toluidine Blue (ICC = 0.919, 95% CI: 0.891-0.941), H&E (ICC = 0.9, 95% CI: 0.865-0.927) and Laminin staining (ICC = 0.897, 95% CI: 0.860-0.926). Intra-rater reliability was good for all three staining techniques (ICC >0.8). This study demonstrates the validity of this simple scoring system to evaluate nerve structure after different processing protocols. All three evaluated staining techniques can reliably be used for both rat and human nerve tissue. We believe this method is suitable to compare different processing techniques to create processed nerve allografts, especially when the average scores of multiple raters are used.

Effects of immune cell-targeted treatments result from the suppression of neuronal oxidative stress and inflammation in experimental diabetic rats.

In this study, we hypothesized that reduction of immune cell activation as well as their oxidant or inflammatory mediators with minocycline (MCN), liposome-encapsulated clodronate (LEC), or anti-Ly6G treatments can be neuroprotective approaches in diabetic neuropathy. MCN (40mg/kg) for reduction of microglial activation, LEC (25mg/kg) for of macrophage inhibition, or anti-Ly6G (150μg/kg) for neutrophil suppression injected to streptozotocin (STZ)-induced diabetic rats twice, 3days, and 1week (half dose) after STZ. Animal mass and blood glucose levels were measured; thermal and mechanical sensitivities were tested for in pain sensations. The levels of chemokine C-X-C motif ligand 1 (CXCL1), CXCL8, and C-C motif ligand 2 (CCL2), CCL3, and total oxidant status (TOS) and total antioxidant status (TAS) were measured in the spinal cord and sciatic nerve tissues of rats. LEC significantly reduced the glucose level of diabetic rats compared with drug control. However, MCN or anti-LY6G did not change the glucose level. While diabetic rats showed a marked decrease in both thermal and mechanical sensations, all treatments alleviated these abnormal sensations. The levels of chemokines and oxidative stress parameters increased in diabetic rats. All drug treatments significantly decreased the CCL2, CXCL1, and CXCL8 levels of spinal cord tissues and ameliorated the neuronal oxidative stress compared with control treatments. Present findings suggest that the neuroprotective actions of MCN, LEC, or anti-Ly6G treatments may be due to the modulation of neuronal oxidative stress and/or inflammatory mediators of immune cells in diabetic rats with neuropathy.

2,5-hexanedione-induced deregulation of axon-related microRNA expression in rat nerve tissues

Exposure to organic solvent in industry, including n-hexane is correlated with central-peripheral axonopathy, which is mediated by its active metabolite, 2,5-hexanedione (HD). However, the underlying mechanism is still largely unknown. Recently identified microRNAs (miRNAs) may play important roles in toxicant exposure and in the process of toxicant-induced neuropathys. To examine the role of miRNAs in HD-induced toxicity, neuropathic animal model was successfully built. miRNA microarray analysis revealed 105 differentially expressed miRNAs after HD exposure. Bioinformatics analysis showed that “Axon” and “Neurotrophin Signaling Pathway” was the top significant GO term and pathway, respectively. 7 miRNAs both related to “Axon” and “Neurotrophin Signaling Pathway” were screened out and further confirmed by Real-Time PCR. Correspondingly, the deregulation expression levels of proteins of four target genes (GSK3β, Map3k1, BDNF and MAP1B) were further confirmed via western blot, verifying the results of gene target analysis. Taken together, our results showed that the axon-related miRNAs to be associated with MAP1B or neurotrophin signal pathways changed in nerve tissues following HD exposure. These miRNAs may play important roles in HD-induced neurotoxicity.

A green route for the fabrication of thermo-sensitive chitosan nerve conduits and their property evaluation

To explore a green route for the fabrication of thermo-sensitive chitosan nerve conduits, improve the mechanical properties and decrease the degradation rate of the chitosan nerve conduits. Taking advantage of the ionic specific effect of the thermo-sensitive chitosan, the strengthened chitosan nerve conduits were obtained by immersing the gel-casted conduits in salt solution for ion-induced phase transition, and rinsing, lyophilization, and 60Co sterilization afterwards. The nerve conduits after immersing in NaCl solutions for 0, 4, 12, 24, 36, 48, and 72 hours were obtained and characterized the general observation, diameters and mechanical properties. According to the above results, the optimal sample was chosen and characterized the microstructure, degradation properties, and cytocompatibility. The left sciatic nerve defect 15 mm in length was made in 20 male Sprague Dawley rats. The autologous nerves (control group, n=10) and the nerve conduits (experimental group, n=10) were used to repair the defects. At 8 weeks after operation, the compound muscle action potential (CMAP) was measured. The regenerated nerves were investigated by gross observation and toluidine blue staining. The gastrocnemius muscle was observed by HE staining. With the increased ionic phase transition time, the color of the conduit was gradually deepened and the diameter was gradually decreased, which showed no difference during 12 hours. The tensile strength of the nerve conduit was increased gradually. The ultimate tensile strength showed significant difference between the 48 hours and 12, 24, and 36 hours groups ( P<0.05), and no significant difference between the 48 hours and 72 hours groups ( P>0.05). As a result, the nerve conduit after ion-induced phase transition for 48 hours was chosen for further study. The scanning electron microscope (SEM) images showed that the nerve conduit had a uniform porous structure. The degradation rate of the the nerve conduit after ion-induced phase transition for 48 hours was significantly decreased as compared with that of the conduit without ion-induced phase transition. The nerve conduit could support the attachment and proliferation of rat Schwann cells on the inner surface. The animal experiments showed that at 8 weeks after operation, the CMAPs of the experimental and control groups were (3.5±0.9) and (4.3±1.1) m/V, respectively, which showed no significant difference between the two groups ( P<0.05), and were significantly lower than that of the contralateral site ［(45.6±5.6 m/V), P>0.05］. The nerve conduit of the experimental group could repair the nerve defect. There was no significant difference between the experimental and control groups in terms of the histomorphology of the regenerated nerve fibers and the gastrocnemius muscle. The green route for the fabrication of thermo-sensitive chitosan nerve conduits is free of any toxic reagents, and has simple steps, which is beneficial to the industrial transformation of the chitosan nerve conduit products. The prepared chitosan nerve conduit can be applied to rat peripheral nerve defect repair and nerve tissue engineering.

Carbon disulfide activates p62-Nrf2-keap1 pathway in rat nerve tissues.

Oxidative stress is associated with the pathogenesis of carbon disulfide (CS2) induced polyneuropathy. The nuclear factor erythroid 2-related factor 2 (Nrf2) plays a critical role in protecting cells against oxidative stress. However, whether there exists a Nrf2-mediated antioxidative machinery in CS2-induced neuropathy has not been elucidated. In the present study, male wistar rats were randomly divided into three experimental groups and one control group. The rats in experimental groups were treated with CS2 by gavage at dosages of 200, 400 and 600mg/kg/day respectively, six times per week for 6 weeks. Nrf2-keap1 antioxidative pathway and p62-related kinase signaling in rat nerve tissues was examined by western blotting and real-time PCR. The results demonstrated that CS2 treatment resulted in Nrf2 translocation from the cytosol to the nucleus in rat spinal cords. In the meantime, the expression of antioxidative enzymes such as NAD(P)H quinone oxidoreductase-1, heme oxygenase-1, and glutamate-cysteine ligase was significantly increased. Furthermore, CS2 treatment increased the level of p62 and its phosphorylation status, while decreased the level of keap1. In addition, CS2 also lead to the activation of CAMKK2 and ULK1 kinase signaling in rat spinal cords and sciatic nerves. Taken together, our results indicated that CS2 intoxication was associated with the activation of Nrf2-ARE antioxidative machinery, which might play a protective role against CS2-induced neuronal damage.

Autophagy is involved in the pathogenesis of experimental autoimmune neuritis in rats.

Recent studies have shown that autophagy is involved in peripheral nervous system disease. However, the role of autophagy in the pathogenesis of experimental autoimmune neuritis (EAN) remains unclear. Therefore, EAN was induced by a subcutaneous injection into both hind footpads of synthetic neuritogenic P2(57-81) peptide in male Lewis rats. The clinical evaluation was completed using a 10-point scale method. The histological alteration of sciatic nerves was analyzed by hematoxylin and eosin and luxol fast blue staining. The ultrastructure of sciatic nerves was analyzed by transmission electron microscopy. Expressions of beclin-1 and microtubule-associated protein light chain-3 (LC3) and p62/SQSTM1 were determined by western blot. 3-Methyladenine, the inhibitor of autophagy, was used in this research. Results showed that the clinical scores were significantly increased from day 6 to day 16 after immunization compared with the control group. Compared with the control group, the number of inflammatory cells and the histological score of sciatic nerves were significantly increased, expressions of beclin-1 and LC3-II and the ratio of LC3-II/LC3-I in the sciatic nerve were significantly increased, and the expression of p62 was significantly decreased in the EAN model group. Considerable double-membrane autophagosomes in axons and myelin sheaths of sciatic nerves were observed and the number of autophagosomes in axons and myelin sheaths of sciatic nerves in the EAN model group was obviously increased compared with the control group. 3-Methyladenine ameliorated the neurologic severity of EAN. Our results suggest that autophagy activity in nerve tissue of EAN rats is increased, which may be associated with the pathogenesis of EAN.

Effect of hyperbaric oxygen on expression of fractalkine in nerve tissues of rats with neuropathic pain

Objective To investigate the effect of hyperbaric oxygen on the expression of fractalkine (FKN) in the nerve tissues of rats with neuropathic pain (NP). Methods Thirty-two male Sprague-Dawley rats, weighing 250-280 g, aged 10-12 weeks, were divided into 4 groups (n= 8 each) using the random number table: control group (group C), sham operation group (group S), group NP, and hyperbaric oxygen group (group H). NP was induced by chronic constriction injury (CCI) in anesthetized rats. The left sciatic nerve was exposed, and 4 loose ligatures were placed on the sciatic nerve at 1 mm intervals with 4-0 silk thread. Group H received hyperbaric oxygen therapy once a day for 5 consecutive days starting from day 1 after CCI. The rats were placed into the hyperbaric oxygen chamber, which was pressurised to 2 atmosphere absolute at a rate of 10 kPa/min, and maintained at this level for 60 min. The pressure was then decreased to the normal pressure at a rate of 10 kPa/min. The mechanical paw withdrawal threshold (MWT) and thermal paw withdrawal latency (TWL) were measured at 1 day before CCI, and 3, 5, 7 and 14 days after CCI. After measurement of pain threshold at 3 and 7 days after CCI, 4 rats were selected and sacrificed. The sciatic nerve and lumbar segment of the spinal cord were removed for determination of the expression of FKN by Western blot. Results Compared with group C, the MWT was significantly decreased, and TWL was shortened at each time point after CCI, the expression of FKN in the sciatic nerve at 3 days after CCI, and in the sciatic nerve and spinal cord at 3 and 7 days after CCI was up-regulated in group NP (P 0.05). Compared with group NP, the MWT was significantly increased, and TWL was prolonged at each time point after CCI, and the expression of FKN in the sciatic nerve at 3 days after CCI, and in the sciatic nerve and spinal cord at 3 and 7 days after CCI was down-regulated in group H (P<0.05). Conclusion The mechanism by which hyperbaric oxygen mitigates NP is related to inhibition of over-expression of FKN in the nerve tissues of rats. Key words: Hyperbaric oxygenation; Neuralgia; Chemotactic factors

Effect of reboxetine on activity of carboxypeptidase E in the nerve tissue of rats

Depression is one of the most common mental disorders, but its etiology is not completely understood. It is assumed that peptidergic system components are involved in the formation of this pathology. Neuropeptides play an important role in the regulation of mental and emotional states. Сarboxypeptidase E is a key enzyme of peptide processing; it regulates neuropeptide levels in the various structures of the nervous system. We have studied effects of a single dose of reboxetine on the activity of carboxypeptidase E in various brain regions and the adrenal glands of rats. The reboxetine injection decreased carboxypeptidase E activity in the pituitary gland (12 h after injection), in the pituitary gland, the quadrigeminal bodies, the medulla oblongata, the hypothalamus, the hippocampus and the amygdala (24 h after injection), in the pituitary gland and striatum (72 h after injection). The enzyme activity in adrenal glands remained basically unchanged. Apparently, the decrease of carboxypeptidase E activity may influence the level of regulatory peptides involved in the pathogenesis of depression.

2,5-Hexanedione altered the degradation of low-molecular-weight neurofilament in rat nerve tissues

Occupational exposure to n-hexane produces a central–peripheral distal axonopathy, which is characterized by giant axonal swellings filled with neurofilaments (NFs). To investigate the change of NFs degradation and their possible role in n-hexane neuropathy, adult male Wistar rats were administered intraperitoneally at a dosage of 400mg/kg/day 2,5-hexanedione (2,5-HD) for 4weeks. The time course of low-molecular-weight neurofilament (NF-L) degradation and autophagy-related protein in rat sciatic nerves and spinal cords was determined by Western blotting. The results demonstrated that the administration of 2,5-HD inhibited NF-L degradation to an undetectable level in sciatic nerves. Furthermore, a significant reduction of NF-L degradation in spinal cords was observed in the early stage of 2,5-HD exposure. In the meantime, 2,5-HD significantly decreased the level of Beclin-1, a key autophagy-regulated protein in sciatic nerves of rats while increased the level of P62, a selective substrate of autophagy degrading pathway, which indicated a dysfunctional autophagy in rat nerve tissues. Collectively, our findings suggested that the inhibition of autophagy by 2,5-HD might be responsible for the reduction of NF-L degradation in rat sciatic nerves, and involved in the pathogenesis of 2,5-HD-induced axonopathy.

Hydroxycloroquine-induced oxidative stress on sciatic nerve and muscle tissue of rats

The aim of this study was to investigate the role of hydroxychloroquine (HCQ)-induced oxidative stress on sciatic nerve and muscle tissues of rats. The oxidant/antioxidant parameters in the sciatic nerve and muscle tissues were analyzed, and stereological analysis of the sciatic nerve was performed. Levels of malondialdehyde and nitric oxide in the tissues were significantly higher in the HCQ group than in the control group (p < 0.05). In addition, activities of superoxide dismutase and glutathione peroxidase were found to be significantly higher in the HCQ group than the control group (p < 0.05). There were significant decreases in nerve fiber diameter and myelin sheet thickness in the HCQ group compared with the control group (p < 0.05). These results revealed that HCQ might increase oxidative stress on sciatic nerve and muscle tissues of rats, which may correlate with axonal atrophy in sciatic nerves.