Stereological method for objectively quantifying myelin sheaths in the rat hippocampus

In the present study,tissue blocks were randomly sampled from the entire hippocampus of 6-week-old Long-Evans rats.Isotropic,uniform and random sections,60 nm thick,were prepared by isector.Fifteen fields of view were randomly selected for each section and photographed using a transmission electron microscope.The mean internal and external diameters of the myelin sheaths were obtained by measuring the longest profile diameter perpendicular to its longest axis.The inner and outer perimeters of the myelin sheaths were estimated using the equidistant parallel test lines.The thickness of the myelin sheaths was estimated by direct orthogonal measurements in uniform,random locations.These stereological methods should permit an unbiased quantitative assessment of changes in the myelin sheaths of myelinated fibers in the hippocam-pus.

Human umbilical cord Wharton's jelly-derived oligodendrocyte precursor-like cells for axon and myelin sheath regeneration

Human umbilical mesenchymal stem cells from Wharton＇s jelly of the umbilical cord were induced to differentiate into oligodendrocyte precursor-like cells in vitro. Oligodendrocyte precursor cells were transplanted into contused rat spinal cords. Immunofluorescence double staining indicated that transplanted cells survived in injured spinal cord, and differentiated into mature and immature oligodendrocyte precursor cells. Biotinylated dextran amine tracing results showed that cell transplantation promoted a higher density of the corticospinal tract in the central and caudal parts of the injured spinal cord. Luxol fast blue and toluidine blue staining showed that the volume of residual myelin was significantly increased at 1 and 2 mm rostral and caudal to the lesion epicenter after cell transplantation. Furthermore, immunofluorescence staining verified that the newly regenerated myelin sheath was derived from the central nervous system. Basso, Beattie and Bresnahan testing showed an evident behavioral recovery. These results suggest that human umbilical mesenchymal stem cell-derived oligodendrocyte precursor cells promote the regeneration of spinal axons and myelin sheaths.

Salvianolic acid B protects the myelin sheath around injured spinal cord axons

Salvianolic acid B,an active pharmaceutical compound present in Salvia miltiorrhiza,exerts a neuroprotective effect in animal models of brain and spinal cord injury.Salvianolic acid B can promote recovery of neurological function;however,its protective effect on the myelin sheath after spinal cord injury remains poorly understood.Thus,in this study,in vitro tests showed that salvianolic acid B contributed to oligodendrocyte precursor cell differentiation,and the most effective dose was 20 μg/m L.For in vivo investigation,rats with spinal cord injury were intraperitoneally injected with 20 mg/kg salvianolic acid B for 8 weeks.The amount of myelin sheath and the number of regenerating axons increased,neurological function recovered,and caspase-3 expression was decreased in the spinal cord of salvianolic acid B-treated animals compared with untreated control rats.These results indicate that salvianolic acid B can protect axons and the myelin sheath,and can promote the recovery of neurological function.Its mechanism of action is likely to be associated with inhibiting apoptosis and promoting the differentiation and maturation of oligodendrocyte precursor cells.

Axonal damage in myelin oligodendrocyte glycoprotein peptide-induced experimental autoimmune encephalomyelitis in a C57BL/6 mouse model may be not secondary to inflammatory demyelination

The present study established a chronic experimental autoimmune encephalomyelitis model in C57BL/6 mice induced by myelin oligodendrocyte glycoprotein peptides and complete Freund＇s adjuvant. Onset latency was 12 days, with an incidence rate of 100%. Neuropathological characteristics included perivascular inflammatory cell infiltration, demyelination, neuronal degeneration, and axonal damage within cerebral and myelic white matter. Electron microscopy revealed swollen mitochondria, complete organ disappearance, and fused or broken myelin sheath structure, which were accompanied by myelin sheath reconstruction. Moreover, axonal damage was not consistent with demyelination distribution, and severity of axonal damage did not correlate with demyelination. Results suggested that axonal damage in an experimental autoimmune encephalomyelitis model is not secondary to inflammatory demyelination.

Insights into myelin dysfunction in schizophrenia and bipolar disorder

Schizophrenia and bipolar disorder are disabling psychiatric disorders with a worldwide prevalence of approximately 1%.Both disorders present chronic and deteriorating prognoses that impose a large burden,not only on patients but also on society and health systems.These mental illnesses share several clinical and neurobiological traits;of these traits,oligodendroglial dysfunction and alterations to white matter(WM)tracts could underlie the disconnection between brain regions related to their symptomatic domains.WM is mainly composed of heavily myelinated axons and glial cells.Myelin internodes are discrete axon-wrapping membrane sheaths formed by oligodendrocyte processes.Myelin ensheathment allows fast and efficient conduction of nerve impulses through the nodes of Ranvier,improving the overall function of neuronal circuits.Rapid and precisely synchronized nerve impulse conduction through fibers that connect distant brain structures is crucial for higher-level functions,such as cognition,memory,mood,and language.Several cellular and subcellular anomalies related to myelin and oligodendrocytes have been found in postmortem samples from patients with schizophrenia or bipolar disorder,and neuroimaging techniques have revealed consistent alterations at the macroscale connectomic level in both disorders.In this work,evidence regarding these multilevel alterations in oligodendrocytes and myelinated tracts is discussed,and the involvement of proteins in key functions of the oligodendroglial lineage,such as oligodendrogenesis and myelination,is highlighted.The molecular components of the axo-myelin unit could be important targets for novel therapeutic approaches to schizophrenia and bipolar disorder.

Action potential-simulated weak electric fields can directly initiate myelination

BACKGROUND： Myelination is a process whereby glial cells identify, adhere, wrap and enclose axons to form a spiral myelin sheath. OBJECTIVE： To investigate the effects of action potential-simulated weak electric fields on myelination in the central nervous system. DESIGN AND SETTING： This single-sample observation study was performed at the 324 Hospital of Chinese PLA. MATERIALS： Two 5 μm carbon fibers were provided by the Institute of Neuroscience, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences. One Sprague Dawley rat, aged 1 day, was used. METHODS： Cerebral cortex was harvested from the rat to prepare a suspension [（1 2）× 10^5/mL] containing neurons and glial cells. To simulate the axon, carbon fibers were placed at the bottom of the neuron-glial cell coculture dish, and were electrified with a single phase square wave current, 1×10^-2, 1×10^-3, 1×10^-4, and 1×10^-5 seconds, 1 Hz, 40 mV, and 10 μA, 30 minutes each, once a day for 10 consecutive days to simulate weak negative electric fields during action potential conduction. MAIN OUTCOME MEASURES： Glial cell growth and wrapping of carbon fibers were observed by phase contrast microscopy and immunohistochemistry. RESULTS： On culture day 7, cell groups were found to adhere to negative carbon fibers in the 1 × 10^-3 seconds square wave group. Cell membrane-like substances grew out of cell groups, wrapped the carbon fibers, and stretched to the ends of carbon fibers. Only some small and round cells close to negative carbon fibers were found on culture day 12. In the 1 × 10^-4 and 1 × 10^-3 seconds square wave groups, the negative carbon fibers were wrapped by oligodendrocytes or their progenitor cells. CONCLUSION： The local negative electric field which is generated by action potentials at 1×（10^-4-10^-3） seconds, 40 mV can directly initiate and participate in myelination in the central nervous system.

Research advances in experiments on correlation between schizophrenia and myelin sheath abnormality

Schizophrenia,as a common mental disease,seriously threatens the physical and mental health of human beings.It is characterized by many mental and behavioral disorders,such as uncoordinated thinking and emotional mode which separated from the actual living environment.Patients with schizophrenia are prone to relapse and deterioration due to their long course of disease,resulting in the loss of labor force.It has been proved that the occurrence and development of schizophrenia is closely related to the abnormal development of oligodendrocytes,which have the function of myelin formation and the dysfunction of myelin sheath itself.For a comprehensive understanding of myelin abnormal effects on the pathogenesis of schizophrenia,this paper is to review the literature,then summarize and discuss the animal experimental literature related to the abnormal myelin sheath in schizophrenia from the perspectives of behavioristics,neuroimaging,protein expression and stereology,in order to further clarify the influence of the abnormal myelin sheath on the pathogenesis of schizophrenia and provide ideas for the diagnosis of schizophrenia and the research and development of new drugs.

Role of myelin sheath energy metabolism in neurodegenerative diseases

More than any other organs,brain energy demand is entirely dependent on glucose catabolism through the oxidative phosphorylation（OXPHOS）.Glucose is the major cerebral energy substrate in the nervous system（NS）.Ketone bodies can be utilized as an additional substrate,but in any case,neurons critically depend on oxygen supply.

A hyaluronic acid granular hydrogel nerve guidance conduit promotes regeneration and functional recovery of injured sciatic nerves in rats

A hyaluronic acid granular hydrogel can promote neuronal and astrocyte colony formation and axonal extension in vitro,suggesting that the hydrogel can simulate an extracellular matrix structure to promote neural regeneration.However,in vivo experiments have not been conducted.In this study,we transplanted a hyaluronic acid granular hydrogel nerve guidance conduit to repair a 10-mm long sciatic nerve gap.The Basso,Beattie,and Bresnahan locomotor rating scale,sciatic nerve compound muscle action potential recording,Fluoro-Gold retrograde tracing,growth related protein 43/S100 immunofluorescence staining,transmission electron microscopy,gastrocnemius muscle dry/wet weight ratio,and Masson’s trichrome staining results showed that the nerve guidance conduit exhibited similar regeneration of sciatic nerve axons and myelin sheath,and recovery of the electrophysiological function and motor function as autologous nerve transplantation.The conduit results were superior to those of a bulk hydrogel or silicone tube transplant.These findings suggest that tissue-engineered nerve conduits containing hyaluronic acid granular hydrogels effectively promote the morphological and functional recovery of the injured sciatic nerve.The nerve conduits have the potential as a material for repairing peripheral nerve defects.

Serum response factor promotes axon regeneration following spinal cord transection injury

Studies have snown that serum response factor is beneficaial for axonar regeneration of peripheral herves.However,Its role after central nervous system injury remains unclear. In this study,we established a rat model of T9-T10 spinal cord transection injury.We found that the expression of serum response factor in injured spinal cord gray matter neurons gradually increased with time,reached its peak on the 7^(th) day,and then gradually decreased.To investigate the role of serum response factor,we used lentivirus vecto rs to ove rexpress and silence serum response factor in spinal cord tissue.We found that overexpression of serum response factor promoted motor function recovery in rats with spinal cord injury.Qualitative observation of biotinylated dextran amine anterograde tra cing showed that ove rexpression of serum response factor increased nerve fibers in the injured spinal co rd.Additionally,transmission electron microscopy showed that axon and myelin sheath morphology was restored.Silencing serum response factor had the opposite effects of ove rexpression.These findings suggest that serum response factor plays a role in the recovery of motor function after spinal cord injury.The underlying mechanism may be related to the regulation of axonal regeneration.

Reduced graphene oxide-embedded nerve conduits loaded with bone marrow mesenchymal stem cell-derived extracellular vesicles promote peripheral nerve regeneration

We previously combined reduced graphene oxide(rGO)with gelatin-methacryloyl(GelMA)and polycaprolactone(PCL)to create an rGO-GelMA-PCL nerve conduit and found that the conductivity and biocompatibility were improved.However,the rGO-GelMA-PCL nerve conduits differed greatly from autologous nerve transplants in their ability to promote the regeneration of injured peripheral nerves and axonal sprouting.Extracellular vesicles derived from bone marrow mesenchymal stem cells(BMSCs)can be loaded into rGO-GelMA-PCL nerve conduits for repair of rat sciatic nerve injury because they can promote angiogenesis at the injured site.In this study,12 weeks after surgery,sciatic nerve function was measured by electrophysiology and sciatic nerve function index,and myelin sheath and axon regeneration were observed by electron microscopy,immunohistochemistry,and immunofluorescence.The regeneration of microvessel was observed by immunofluorescence.Our results showed that rGO-GelMA-PCL nerve conduits loaded with BMSC-derived extracellular vesicles were superior to rGO-GelMA-PCL conduits alone in their ability to increase the number of newly formed vessels and axonal sprouts at the injury site as well as the recovery of neurological function.These findings indicate that rGO-GelMA-PCL nerve conduits loaded with BMSC-derived extracellular vesicles can promote peripheral nerve regeneration and neurological function recovery,and provide a new direction for the curation of peripheral nerve defect in the clinic.

Collagen/heparan sulfate porous scaffolds loaded with neural stem cells improve neurological function in a rat model of traumatic brain injury

One reason for the poor therapeutic effects of stem cell transplantation in traumatic brain injury is that exogenous neural stem cells cannot effectively migrate to the local injury site,resulting in poor adhesion and proliferation of neural stem cells at the injured area.To enhance the targeted delivery of exogenous stem cells to the injury site,cell therapy combined with neural tissue engineering technology is expected to become a new strategy for treating traumatic brain injury.Collagen/heparan sulfate porous scaffolds,prepared using a freeze-drying method,have stable physical and chemical properties.These scaffolds also have good cell biocompatibility because of their high porosity,which is suitable for the proliferation and migration of neural stem cells.In the present study,collagen/heparan sulfate porous scaffolds loaded with neural stem cells were used to treat a rat model of traumatic brain injury,which was established using the controlled cortical impact method.At 2 months after the implantation of collagen/heparan sulfate porous scaffolds loaded with neural stem cells,there was significantly improved regeneration of neurons,nerve fibers,synapses,and myelin sheaths in the injured brain tissue.Furthermore,brain edema and cell apoptosis were significantly reduced,and rat motor and cognitive functions were markedly recovered.These findings suggest that the novel collagen/heparan sulfate porous scaffold loaded with neural stem cells can improve neurological function in a rat model of traumatic brain injury.This study was approved by the Institutional Ethics Committee of Characteristic Medical Center of Chinese People’s Armed Police Force,China(approval No.2017-0007.2)on February 10,2019.

The mechanism of astragaloside Ⅳ promoting sciatic nerve regeneration

3-O-beta-D-xylopyranosyl-6-O-beta-D-glucopyranosyl-cycloastragenol （astragaloside IV）, the main active component of the traditional Chinese medicine astragalus membranaceus, has been shown to be neuroprotective. This study investigated whether astragaloside IV could promote the repair of injured sciatic nerve. Denervated sciatic nerve of mice was subjected to anastomosis. The mice were intraperitoneally injected with 10, 5, 2.5 mg/kg astragaloside IV per day for 8 consecutive days Western blot assay and real-time PCR results demonstrated that growth-associated protein-43 ex- pression was upregulated in mouse spinal cord segments L4-6 after intervention with 10, 5, 2.5 mg/kg astragaloside IV per day in a dose-dependent manner. Luxol fast blue staining and elec- trophysiological detection suggested that astragaloside IV elevated the number and diameter of myelinated nerve fibers, and simultaneously increased motor nerve conduction velocity and action potential amplitude in the sciatic nerve of mice. These results indicated that astragaloside IV con- tributed to sciatic nerve regeneration and functional recovery in mice. The mechanism underlying this effect may be associated with the upregulation of growth-associated protein-43 expression.

An inside-out vein graft filled with platelet-rich plasma for repair of a short sciatic nerve defect in rats

Platelet-rich plasma containing various growth factors can promote nerve regeneration. An inside-out vein graft can substitute nerve autograft to repair short nerve defects. It is hypothesized that an inside-out vein graft filled with platelet-rich plasma shows better effects in the repair of short sciatic nerve defects. In this study, an inside-out vein autograft filled with platelet-rich plasma was used to bridge a 10 mm-long sciatic nerve defect in rats. The sciatic nerve function of rats with an inside-out vein autograft filled with platelet-rich plasma was better improved than that of rats with a simple inside-out vein autograft. At 6 and 8 weeks, the sciatic nerve function of rats with an inside-out vein autograft filled with platelet-rich plasma was better than that of rats undergoing nerve autografting. Compared with the sciatic nerve repaired with a simple inside-out vein autograft, the number of myelinated axons was higher, axon diameter and myelin sheath were greater in the sciatic nerve repaired with an inside-out vein autograft filled with plateletrich plasma and they were similar to those in the sciatic nerve repaired with nerve autograft. These findings suggest that an inside-out vein graft filled with platelet-rich plasma can substitute nerve autograft to repair short sciatic nerve defects.

Curcumin upregulates S100 expression and improves regeneration of the sciatic nerve following its complete amputation in mice

The repair of peripheral nerve injury after complete amputation is difficult,and even with anastomosis,the rapid recovery of nerve function remains challenging.Curcumin,extracted from plants of the genus Curcuma,has been shown to have anti-oxidant and anti-inflammatory properties and to improve sciatic nerve crush injury in rats.Here,we determined whether curcumin had neuroprotective effects following complete peripheral nerve amputation injury.BALB/c mice underwent complete sciatic nerve amputation,followed by an immediate epineurium anastomosis.Mice were intragastrically administered curcumin at doses of 40（high）,20（moderate）,and 10 mg/kg/d（low） for 1 week.We found that myelin in the mice of the high- and moderate-dose curcumin groups appeared with regular shape,uniform thickness,clear boundary,and little hyperplasia surrounding the myelin.High and moderate doses of curcumin markedly improved both action potential amplitude of the sciatic nerves and the conduction velocity of the corresponding motor neurons,and upregulated m RNA and protein expression of S100,a marker for Schwann cell proliferation,in L4–6 spinal cord segments.These results suggest that curcumin is effective in promoting the repair of complete sciatic nerve amputation injury and that the underlying mechanism may be associated with upregulation of S100 expression.

Biomimetic chitosan scaffolds with long-term controlled release of nerve growth factor repairs 20-mm-long sciatic nerve defects in rats

Although autogenous nerve transplantation is the gold standard for treating peripheral nerve defects of considerable length,it still has some shortcomings,such as insufficient donors and secondary injury.Composite chitosan scaffolds loaded with controlled release of nerve growth factor can promote neuronal survival and axonal regeneration after short-segment sciatic nerve defects.However,the effects on extended nerve defects remain poorly understood.In this study,we used chitosan scaffolds loaded with nerve growth factor for 8 weeks to repair long-segment(20 mm)sciatic nerve defects in adult rats.The results showed that treatment markedly promoted the recovery of motor and sensory functions.The regenerated sciatic nerve not only reconnected with neurons but neural circuits with the central nervous system were also reconstructed.In addition,the regenerated sciatic nerve reconnected the motor endplate with the target muscle.Therefore,this novel biomimetic scaffold can promote the regeneration of extended sciatic nerve defects and reconstruct functional circuits.This provides a promising method for the clinical treatment of extended peripheral nerve injury.This study was approved by the Animal Ethics Committee of Capital Medical University,China(approval No.AEEI-2017-033)on March 21,2017.

L-carnitine alleviates sciatic nerve crush injury in rats:functional and electron microscopy assessments

Several studies have demonstrated that L-carnitine exhibits neuroprotective effects on injured sciatic nerve of rats with diabetes mellitus. It is hypothesized that L-carnitine exhibits neuro-protective effects on injured sciatic nerve of rats. Rat sciatic nerve was crush injured by a forceps and exhibited degenerative changes. After intragastric administration of 50 and 100 mg/kg L-carnitine for 30 days, axon area, myelin sheath area, axon diameter, myelin sheath diameter, and numerical density of the myelinated axons of injured sciatic nerve were similar to normal, and the function of injured sciatic nerve also improved signiifcantly. These ifndings suggest that L-carnitine exhibits neuroprotective effects on sciatic nerve crush injury in rats.

Human bone marrow mesenchymal stem cell transplantation attenuates axonal injury in stroke rats

Previous studies have shown that transplantation of human bone marrow mesenchymal stem cells promotes neural functional recovery after stroke, but the neurorestorative mechanisms remain largely unknown. We hypothesized that functional recovery of myelinated axons may be one of underlying mechanisms. In this study, an ischemia/reperfusion rat model was established using the middle cerebral artery occlusion method. Rats were used to test the hypothesis that intravenous transplantation of human bone marrow mesenchyrnal stem cells through the femoral vein could exert neuroprotective effects against cerebral ischemia via a mechanism associated with the ability to attenuate axonal injury. The results of behavioral tests, infarction volume analysis and immunohistochemistry showed that cerebral ischemia caused severe damage to the myelin sheath and axons. After rats were intravenously transplanted with human bone marrow mesenchymal stem cells, the levels of axon and myelin sheath-related proteins, including microtubule-associated protein 2, myelin basic protein, and growth-associated protein 43, were elevated, infarct volume was decreased and neural function was improved in cerebral ischemic rats. These findings suggest that intravenously transplanted human bone marrow mesenchymal stem cells promote neural function. Possible mechanisms underlying these beneficial effects include resistance to demyelination after cerebral ischemia, prevention of axonal degeneration, and promotion of axonal regeneration.

Ursolic acid induces neural regeneration after sciatic nerve injury

In this study, we aimed to explore the role of ursolic acid in the neural regeneration of the injured sciatic nerve. BALB/c mice were used to establish models of sciatic nerve injury through unilateral sciatic nerve complete transection and microscopic anastomosis at 0.5 cm below the ischial tuber-osity. The successful y generated model mice were treated with 10, 5, or 2.5 mg/kg ursolic acid via intraperitoneal injection. Enzyme-linked immunosorbent assay results showed that serum S100 protein expression level gradual y increased at 1-4 weeks after sciatic nerve injury, and significantly decreased at 8 weeks. As such, ursolic acid has the capacity to significantly increase S100 protein expression levels. Real-time quantitative PCR showed that S100 mRNA expression in the L 4-6 segments on the injury side was increased after ursolic acid treatment. In addition, the muscular mass index in the soleus muscle was also increased in mice treated with ursolic acid. Toluidine blue staining revealed that the quantity and average diameter of myelinated nerve fibers in the injured sciatic nerve were significantly increased after treatment with ursolic acid. 10 and 5 mg/kg of ursolic acid produced stronger effects than 2.5 mg/kg of ursolic acid. Our findings indicate that ursolic acid can dose-dependently increase S100 expression and promote neural regeneration in BALB/c mice fol owing sciatic nerve injury.

Expression pattern of neuregulin-1 type Ⅲ during the development of the peripheral nervous system

Neuregulin-1 type Ⅲ is a key regulator in Schwann cell proliferation, committing to a myelinat- ing fate and regulating myelin sheath thickness. However, the expression pattern of neuregulin- 1 type III in the peripheral nervous system during developmental periods （such as the premyelin- ating stage, myelinating stage and postmyelinating stage） has rarely been studied. In this study, dorsal root ganglia were isolated from rats between postnatal day 1 and postnatal day 56. The expression pattern of neuregulin-1 type III in dorsal root ganglia neurons at various develop- mental stages were compared by quantitative real-time polymerase chain reaction, western blot assay and immunofluorescent staining. The expression of neuregulin-I type Ⅲ mRNA reached its peak at postnatal day 3 and then stabilized at a relative high expression level from postnatal day 3 to postnatal day 56. The expression of neuregulin-1 type III protein increased gradually from postnatal day 1, reached a peak at postnatal day 28, and then decreased at postnatal day 56. Immunofluorescent staining results showed a similar tendency to western blot assay results. Experimental findings indicate that the expression of neuregulin-1 type III in rat dorsal root ganglion was increased during the premyelinating （from postnatal day 2 to postnatal day 5） and myelinating stage （from postnatal day 5 to postnatal day 10）, but remained at a high level in the postmyelinating stage （after postnatal day 10）.