Myelin Thickness Research Articles

Promising application of a novel biomaterial, light chain of silk fibroin combined with NT3, in repairment of rat sciatic nerve defect injury

Autologous nerve transplantation is the gold standard for treating peripheral nerve defects, but it is associated with defects such as insufficient donor and secondary injury. Artificial nerve guidance conduits (NGCs) are now considered promising alternatives for bridging long nerve gaps, although exploring new biomaterials to construct NGCs remains challenging. Silk fibroin (SF) has good biocompatibility and can self-assemble in aqueous solutions. However, the lack of proximal neurotrophic factors after nerve injury is a major concern, leading to incomplete nerve regeneration. In this study, NT-3, a neurotrophin that promotes neuronal survival and differentiation, was bound to the light chain of silk fibroin (FIBL) in two ways: one was directly bound to FIBL (FIBL-NT3) and the other was a polypeptides-linker (FIBL-Linker-NT3). The design aimed to take advantage of silk fiber's character of self-assembly of heavy-light chains and test whether a flexible linker with NT3 molecule is easy to be a NT3 dimer, the active form. In vitro studies indicated that FIBL-Linker-NT3 combined with SF membranes promoted axon growth in adult rat dorsal root ganglion (DRG) neurons. Then we tested if FIBL-Linker-NT3 could self-assemble with the SF heavy chain (SFH). DTT (Dithiothreitol) was used to break the disulfide bonds between the SF light and heavy chains, and the light-chain protein was removed via dialysis. SFH was assembled using FIBL-Linker-NT3, as evidenced by the western blotting results that showed a high molecular band corresponding to SFH-FIBL-Linker-NT3. Chitosan scaffolds have been identified to provide a suitable microenvironment, so a chitosan/SF-FIBL-Linker-NT3 conduit was also constructed. Nerve transplantation of this conduit was evaluated in vivo in a rat sciatic nerve defect model. Immunohistochemical assays showed that the chitosan/SF-FIBL-Linker-NT3 group was superior to the chitosan/PBS, SF, PBS + FIBL-Linker-NT3 groups in nerve regeneration. In addition, the chitosan/SF-FIBL-Linker-NT3 conduit-transplanted group exhibited better recovery in terms of neurite length, sciatic functional index value, sensitivity to heat, time on the rotarod, wet weight ratio, cross-sectional area, compound muscle action potential, number of myelin layers, and myelin thickness in the nerve. Taking together, our study identified that FIBL-Linker-NT3 could promote axonal growth and regeneration in vivo and in vitro and is a promising candidate biomaterial for artificial NGCs.

VEGFA-modified DPSCs combined with LC-YE-PLGA NGCs promote facial nerve injury repair in rats

ObjectiveThe aim of this research was to investigate the effect of vascular endothelial growth factor A (VEGFA)-overexpressing rat dental pulp stem cells (rDPSCs) combined with laminin-coated and yarn-encapsulated poly(l-lactide-co-glycolide) (PLGA) nerve guidance conduit (LC-YE-PLGA NGC) in repairing 10 mm facial nerve injury in rats. Study DesignrDPSCs isolated from rat mandibular central incisor were cultured and identified in vitro and further transfected with the lentiviral vectors (Lv-VEGFA). To investigate the role and mechanisms of VEGFA in neurogenic differentiation in vitro, semaxanib (SU5416), Cell Counting Kit-8 (CCK-8), real-time quantitative polymerase chain reaction (qPCR) and Western blotting were performed. Ten-millimeter facial nerve defect models in rats were established and bridged by LC-YE-PLGA NGCs. The repair effects were detected by transmission electron microscopy (TEM), compound muscle action potential (CMAP), immunohistochemistry and immunofluorescence. ResultsExtracted cells exhibited spindle-shaped morphology, presented typical markers (CD44+CD90+CD34−CD45−), and presented multidirectional differentiation potential. The DPSCs with VEGFA overexpression were constructed successfully. VEGFA enhanced the proliferation and neural differentiation ability of rDPSCs, and the expression of neuron-specific enolase (NSE) and βIII-tubulin was increased. However, these trends were reversed with the addition of SU5416. This suggests that VEGFA mediates the above effects mainly through vascular endothelial growth factor receptor 2 (VEGFR2) binding. The LC-YE-NGC basically meet the requirements of facial nerve repair. For the in vivo experiment, the CMAP latency period was shorter in DPSCS-VEGFA-NGC group in comparison with other experimental groups, while the amplitude was increased. Such functional recovery correlated well with an increase in histological improvement. Further study suggested that VEGFA-modified DPSCs could increase the myelin number, thickness and axon diameter of facial nerve. NSE, βIII-tubulin and S100 fluorescence intensity and immunohistochemical staining intensity were significantly enhanced. ConclusionVEGFA-modified rDPSCs combined with LC-YE-PLGA NGCs have certain advantages in the growth and functional recovery of facial nerves in rats.

Node of Ranvier remodeling in chronic psychosocial stress and anxiety

Differential expression of myelin-related genes and changes in myelin thickness have been demonstrated in mice after chronic psychosocial stress, a risk factor for anxiety disorders. To determine whether and how stress affects structural remodeling of nodes of Ranvier, another form of myelin plasticity, we developed a 3D reconstruction analysis of node morphology in C57BL/6NCrl and DBA/2NCrl mice. We identified strain-dependent effects of chronic social defeat stress on node morphology in the medial prefrontal cortex (mPFC) gray matter, including shortening of paranodes in C57BL/6NCrl stress-resilient and shortening of node gaps in DBA/2NCrl stress-susceptible mice compared to controls. Neuronal activity has been associated with changes in myelin thickness. To investigate whether neuronal activation is a mechanism influencing also node of Ranvier morphology, we used DREADDs to repeatedly activate the ventral hippocampus-to-mPFC pathway. We found reduced anxiety-like behavior and shortened paranodes specifically in stimulated, but not in the nearby non-stimulated axons. Altogether, our data demonstrate (1) nodal remodeling of the mPFC gray matter axons after chronic stress and (2) axon-specific regulation of paranodes in response to repeated neuronal activity in an anxiety-associated pathway. Nodal remodeling may thus contribute to aberrant circuit function associated with anxiety disorders.

Treatment with the Glycosphingolipid Modulator THI Rescues Myelin Integrity in the Striatum of R6/2 HD Mice

Huntington's disease is one of the most common dominantly inherited neurodegenerative disorders caused by an expansion of a polyglutamine (polyQ) stretch in the N-terminal region of huntingtin (Htt). Among all the molecular mechanisms, affected by the mutation, emerging evidence proposes glycosphingolipid dysfunction as one of the major determinants. High levels of sphingolipids have been found to localize in the myelin sheaths of oligodendrocytes, where they play an important role in myelination stability and functions. In this study, we investigated any potential existing link between sphingolipid modulation and myelin structure by performing both ultrastructural and biochemical analyses. Our findings demonstrated that the treatment with the glycosphingolipid modulator THI preserved myelin thickness and the overall structure and reduced both area and diameter of pathologically giant axons in the striatum of HD mice. These ultrastructural findings were associated with restoration of different myelin marker protein, such as myelin-associated glycoprotein (MAG), myelin basic protein (MBP) and 2', 3' Cyclic Nucleotide 3'-Phosphodiesterase (CNP). Interestingly, the compound modulated the expression of glycosphingolipid biosynthetic enzymes and increased levels of GM1, whose elevation has been extensively reported to be associated with reduced toxicity of mutant Htt in different HD pre-clinical models. Our study further supports the evidence that the metabolism of glycosphingolipids may represent an effective therapeutic target for the disease.

Morphometric Analysis of Retinal Ganglion Cell Axons in Normal and Glaucomatous Brown Norway Rats Optic Nerves.

A reference atlas of optic nerve (ON) retinal ganglion cell (RGC) axons could facilitate studies of neuro-ophthalmic diseases by detecting subtle RGC axonal changes. Here we construct an RGC axonal atlas for normotensive eyes in Brown Norway rats, widely used in glaucoma research, and also develop/evaluate several novel metrics of axonal damage in hypertensive eyes. Light micrographs of entire ON cross-sections from hypertensive and normotensive eyes were processed through a deep learning-based algorithm, AxoNet2.0, to determine axonal morphological properties and were semiquantitatively scored using the Morrison grading scale (MGS) to provide a damage score independent of AxoNet2.0 outcomes. To construct atlases, ONs were conformally mapped onto an ON "template," and axonal morphometric data was computed for each region. We also developed damage metrics based on myelin morphometry. In normotensive eyes, average axon density was ∼0.3 axons/µm2 (i.e., ∼80,000 axons in an ON). We measured axoplasm diameter, eccentricity, cross-sectional area, and myelin g-ratio and thickness. Most morphological parameters exhibited a wide range of coefficients of variation (CoV); however, myelin thickness CoV was only ∼2% in normotensive eyes. In hypertensive eyes, increased myelin thickness correlated strongly with MGS (P < 0.0001). We present the first comprehensive normative RGC axon morphometric atlas for Brown Norway rat eyes. We suggest objective, repeatable damage metrics based on RGC axon myelin thickness for hypertensive eyes. These tools can evaluate regional changes in RGCs and overall levels of damage in glaucoma studies using Brown Norway rats.

Pharmacological treatment promoting remyelination enhances motor function after internal capsule demyelination in mice.

Multiple sclerosis (MS) is an inflammatory demyelinating disease of the central nervous system characterized by remyelination failure, axonal degeneration, and progressive worsening of motor functions. Animal models of demyelination are frequently used to develop and evaluate therapies for MS. We recently reported that focal internal capsule (IC) demyelination in mice with lysophosphatidylcholine injection induced acute motor deficits followed by recovery through remyelination. However, it remains unknown whether the IC demyelination mouse model can be used to evaluate changes in motor functions caused by pharmacological treatments that promote remyelination using behavioral testing and histological analysis. In this study, we examined the effect of clemastine, an anti-muscarinic drug that promotes remyelination, in the mouse IC demyelination model. Clemastine administration improved motor function and changed forepaw preference in the IC demyelinated mice. Moreover, clemastine-treated mice showed increased mature oligodendrocyte density, reduced axonal injury, an increased number of myelinated axons and thicker myelin in the IC lesions compared with control (PBS-treated) mice. These results suggest that the lysophosphatidylcholine-induced IC demyelination model is useful for evaluating changes in motor functions following pharmacological treatments that promote remyelination.

Vitamin D3 Promotes Structural and Functional Recovery After Vincristine-Induced Peripheral Neuropathy in Rats: An Experimental Study.

Background Vincristine-induced peripheral neuropathy (VIPN) is a distal axonopathy characterized by the loss of distal myelinated axons. This study aimed to assess the potential neuroregenerative roles of vitamin D3 using functional and electron microscopic analyses in a rat model of VIPN. Methodology A total of 40female Wistar rats were randomly divided into four main groups: Group 1 (control, n = 10), Group 2 (vincristine, n =10), Group 3 (vincristine + vitamin D3, n = 10), and Group 4 (vincristine + vehicle, n = 10). Vincristine was administered intraperitoneally at a dose of 0.15 mg/kg, for two weeks, to induce peripheral neuropathy. Following successful induction, vitamin D3 (500 IU/kg/day) and vehicle treatments were applied weekly over four weeks. Structural (electron microscopic analysis) and functional analysis (von Frey test, pinch test, and electrophysiological analysis) were performed to assess functional recovery after peripheral nerve impairment. Results Withdrawal responses to mechanical allodynia and pinch tests were significantly higher in the vitamin D3-treated group (P< 0.05). The electrophysiological analysis also supported these results. Electron microscopic evaluation revealed that the remyelinated nerve fibers in the vitamin D3-treated group (Group 3) had thick myelin sheaths and normal axonal morphology. Conclusions Our study demonstrated that vitamin D3 could promote functional and structural recovery in a rat model of VIPN. Further studies should be conducted to elucidate the underlying mechanisms by which vitamin D3 exerts its regenerative effects in VIPN, using alternative administration protocols.

Secondary Degeneration Impairs Myelin Ultrastructural Development in Adulthood following Adolescent Neurotrauma in the Rat Optic Nerve

Adolescence is a critical period of postnatal development characterized by social, emotional, and cognitive changes. These changes are increasingly understood to depend on white matter development. White matter is highly vulnerable to the effects of injury, including secondary degeneration in regions adjacent to the primary injury site which alters the myelin ultrastructure. However, the impact of such alterations on adolescent white matter maturation is yet to be investigated. To address this, female piebald-virol-glaxo rats underwent partial transection of the optic nerve during early adolescence (postnatal day (PND) 56) with tissue collection two weeks (PND 70) or three months later (PND 140). Axons and myelin in the transmission electron micrographs of tissue adjacent to the injury were classified and measured based on the appearance of the myelin laminae. Injury in adolescence impaired the myelin structure in adulthood, resulting in a lower percentage of axons with compact myelin and a higher percentage of axons with severe myelin decompaction. Myelin thickness did not increase as expected into adulthood after injury and the relationship between the axon diameter and myelin thickness in adulthood was altered. Notably, dysmyelination was not observed 2 weeks postinjury. In conclusion, injury in adolescence altered the developmental trajectory, resulting in impaired myelin maturation when assessed at the ultrastructural level in adulthood.

Differential impacts of Cntnap2 heterozygosity and Cntnap2 null homozygosity on axon and myelinated fiber development in mouse.

Over the last decade, a large variety of alterations of the Contactin Associated Protein 2 (CNTNAP2) gene, encoding Caspr2, have been identified in several neuronal disorders, including neurodevelopmental disorders and peripheral neuropathies. Some of these alterations are homozygous but most are heterozygous, and one of the current challenges is to estimate to what extent they could affect the functions of Caspr2 and contribute to the development of these pathologies. Notably, it is not known whether the disruption of a single CNTNAP2 allele could be sufficient to perturb the functions of Caspr2. To get insights into this issue, we questioned whether Cntnap2 heterozygosity and Cntnap2 null homozygosity in mice could both impact, either similarly or differentially, some specific functions of Caspr2 during development and in adulthood. We focused on yet poorly explored functions of Caspr2 in axon development and myelination, and performed a morphological study from embryonic day E17.5 to adulthood of two major brain interhemispheric myelinated tracts, the anterior commissure (AC) and the corpus callosum (CC), comparing wild-type (WT), Cntnap2 -/- and Cntnap2 +/- mice. We also looked for myelinated fiber abnormalities in the sciatic nerves of mutant mice. Our work revealed that Caspr2 controls the morphology of the CC and AC throughout development, axon diameter at early developmental stages, cortical neuron intrinsic excitability at the onset of myelination, and axon diameter and myelin thickness at later developmental stages. Changes in axon diameter, myelin thickness and node of Ranvier morphology were also detected in the sciatic nerves of the mutant mice. Importantly, most of the parameters analyzed were affected in Cntnap2 +/- mice, either specifically, more severely, or oppositely as compared to Cntnap2 -/- mice. In addition, Cntnap2 +/- mice, but not Cntnap2 -/- mice, showed motor/coordination deficits in the grid-walking test. Thus, our observations show that both Cntnap2 heterozygosity and Cntnap2 null homozygosity impact axon and central and peripheral myelinated fiber development, but in a differential manner. This is a first step indicating that CNTNAP2 alterations could lead to a multiplicity of phenotypes in humans, and raising the need to evaluate the impact of Cntnap2 heterozygosity on the other neurodevelopmental functions of Caspr2.

Novel insights into axon diameter and myelin content in late childhood and adolescence.

White matter microstructural development in late childhood and adolescence is driven predominantly by increasing axon density and myelin thickness. Ex vivo studies suggest that the increase in axon diameter drives developmental increases in axon density observed with pubertal onset. In this cross-sectional study, 50 typically developing participants aged 8-18years were scanned using an ultra-strong gradient magnetic resonance imaging scanner. Microstructural properties, including apparent axon diameter $({d}_a)$, myelin content, and g-ratio, were estimated in regions of the corpus callosum. We observed age-related differences in ${d}_a$, myelin content, and g-ratio. In early puberty, males had larger ${d}_a$ in the splenium and lower myelin content in the genu and body of the corpus callosum, compared with females. Overall, this work provides novel insights into developmental, pubertal, and cognitive correlates of individual differences in apparent axon diameter and myelin content in the developing human brain.

The TGR5 Agonist INT-777 Promotes Peripheral Nerve Regeneration by Activating cAMP-dependent Protein Kinase A in Schwann Cells.

Schwann cell (SC) myelination is a pivotal event in the normal physiological functioning of the peripheral nervous system (PNS), where myelination is finely controlled by a series of factors within SCs to ensure timely onset and correct myelin thickness for saltatory conduction. Among these, cyclic AMP (cAMP) is a promising factor for driving myelin gene expression in SCs. It has been shown that TGR5 activation is often associated with increased production of cAMP. Therefore, we speculated that the G-protein-coupled receptor (TGR5) might be involved in the PNS myelination. To test this hypothesis, sciatic nerve crush-injured mice were treated with INT-777, a specific agonist of TGR5, which significantly improved remyelination and functional recovery. Furthermore, rats that underwent sciatic nerve transection were treated with INT-777, which also promoted nerve regeneration and functional recovery. In primary SCs, the stimulatory effect of INT-777 on myelin gene expression was largely counteracted by H89, a potent inhibitor of cAMP-dependent protein kinase A (PKA). Additionally, INT-777 stimulated cell migration was blunted in the presence of H89. Overall, these data indicate that INT-777 is capable of promoting peripheral nerve regeneration and functional recovery after injury, and these benefits are likely due to the activation of the TGR5/cAMP/PKA axis. As such, INT-777, together with other TGR5 agonists, may hold great therapeutic potential for treating peripheral nerve injury.

Early microglial response, myelin deterioration and lethality in mice deficient for very long chain ceramide synthesis in oligodendrocytes.

The sphingolipids galactosylceramide (GalCer), sulfatide (ST) and sphingomyelin (SM) are essential for myelin stability and function. GalCer and ST are synthesized mostly from C22-C24 ceramides, generated by Ceramide Synthase 2 (CerS2). To clarify the requirement for C22-C24 sphingolipid synthesis in myelin biosynthesis and stability, we generated mice lacking CerS2 specifically in myelinating cells (CerS2ΔO/ΔO ). At 6 weeks of age, normal-appearing myelin had formed in CerS2ΔO/ΔO mice, however there was a reduction in myelin thickness and the percentage of myelinated axons. Pronounced loss of C22-C24 sphingolipids in myelin of CerS2ΔO/ΔO mice was compensated by greatly increased levels of C18 sphingolipids. A distinct microglial population expressing high levels of activation and phagocytic markers such as CD64, CD11c, MHC class II, and CD68 was apparent at 6 weeks of age in CerS2ΔO/ΔO mice, and had increased by 10 weeks. Increased staining for denatured myelin basic protein was also apparent in 6-week-old CerS2ΔO/ΔO mice. By 16 weeks, CerS2ΔO/ΔO mice showed pronounced myelin atrophy, motor deficits, and axon beading, a hallmark of axon stress. 90% of CerS2ΔO/ΔO mice died between 16 and 26 weeks of age. This study highlights the importance of sphingolipid acyl chain length for the structural integrity of myelin, demonstrating how a modest reduction in lipid chain length causes exposure of a denatured myelin protein epitope and expansion of phagocytic microglia, followed by axon pathology, myelin degeneration, and motor deficits. Understanding the molecular trigger for microglial activation should aid the development of therapeutics for demyelinating and neurodegenerative diseases.

Behavioral and axon histomorphometric analyses after intraneural transplantation of human skin- and nerve-derived Schwann cells in nude rats.

We have recently isolated and expanded skin-derived Schwann cells (Sk-SCs) from human skin and showed that they are largely similar to nerve-derived Schwann cells (N-SCs). Here, we extend our investigation into functional assessments of the nude rats that received human Sk-SCs and N-SCs after intraneural delivery into crushed and decellularized tibial nerve in adult nude rats. Sk-SCs, N-SCs, dermal fibroblasts, or control culture medium was injected into the crushed and decellularized tibial nerve using in situ repeated freeze-thaw cycles. Animals were then subjected to a ladder rung walking test, nociceptive von Frey testing, and walking gait analysis weekly. Animals were euthanized 6 weeks after surgery, gastrocnemius and soleus muscles were weighed, distal nerves were harvested, and whole semithin cross-sections were analyzed using segmentation software. N-SC-injected and dermal fibroblast-injected animals improved significantly at 4 to 6 weeks postinjury in nociceptive assessment compared with medium-injected controls. Sk-SCs recovered more rapidly in tibial functional index at 2 weeks postinjury compared with medium-injected controls. No significant difference was observed for the ladder rung walking test or muscle weight ratio. Histologically, the number of myelinated axons was significantly higher in all cell injection groups compared with medium-injected controls. No significant difference was observed in g ratio, axon diameter, or myelin thickness. Cell injection significantly improved axon regeneration across an in situ decellularized nerve segment. However, a more human cell-permissive animal model is required to delineate functional differences between cell types for preclinical transplantation studies.

Sensitivity of the electrical response of a node of Ranvier model to alterations of the myelin sheath geometry.

Nodes of Ranvier play critical roles in the generation and transmission of action potentials. Alterations in node properties during pathology and/or development are known to affect the speed and quality of electrical transmission. From a modelling standpoint, nodes of Ranvier are often described by systems of ordinary differential equations neglecting or greatly simplifying their geometric structure. These approaches fail to accurately describe how fine scale alteration in the node geometry or in myelin thickness in the paranode region will impact action potential generation and transmission. Here, we rely on a finite element approximation to describe the three dimensional geometry of a node of Ranvier. With this, we are able to investigate how sensitive is the electrical response to alterations in the myelin sheath and paranode geometry. We could in particular investigate irregular loss of myelin, which might be more physiologically relevant than the uniform loss often described through simpler modelling approaches.

The long noncoding RNA FEDORA is a cell type- and sex-specific regulator of depression.

Women suffer from depression at twice the rate of men, but the underlying molecular mechanisms are poorly understood. Here, we identify marked baseline sex differences in the expression of long noncoding RNAs (lncRNAs), a class of regulatory transcripts, in human postmortem brain tissue that are profoundly lost in depression. One such human lncRNA, RP11-298D21.1 (which we termed FEDORA), is enriched in oligodendrocytes and neurons and up-regulated in the prefrontal cortex (PFC) of depressed females only. We found that virally expressing FEDORA selectively either in neurons or in oligodendrocytes of PFC promoted depression-like behavioral abnormalities in female mice only, changes associated with cell type-specific regulation of synaptic properties, myelin thickness, and gene expression. We also found that blood FEDORA levels have diagnostic implications for depressed women and are associated with clinical response to ketamine. These findings demonstrate the important role played by lncRNAs, and FEDORA in particular, in shaping the sex-specific landscape of the brain and contributing to sex differences in depression.

Histology-informed automatic parcellation of white matter tracts in the rat spinal cord.

The white matter is organized into "tracts" or "bundles," which connect different parts of the central nervous system. Knowing where these tracts are located in each individual is important for understanding the cause of potential sensorial, motor or cognitive deficits and for developing appropriate treatments. Traditionally, tracts are found using tracer injection, which is a difficult, slow and poorly scalable technique. However, axon populations from a given tract exhibit specific characteristics in terms of morphometrics and myelination. Hence, the delineation of tracts could, in principle, be done based on their morphometry. The objective of this study was to generate automatic parcellation of the rat spinal white matter tracts using the manifold information from scanning electron microscopy images of the entire spinal cord. The axon morphometrics (axon density, axon diameter, myelin thickness and g-ratio) were computed pixelwise following automatic axon segmentation using AxonSeg. The parcellation was based on an agglomerative clustering algorithm to group the tracts. Results show that axon morphometrics provide sufficient information to automatically identify some white matter tracts in the spinal cord, however, not all tracts were correctly identified. Future developments of microstructure quantitative MRI even bring hope for a personalized clustering of white matter tracts in each individual patient. The generated atlas and the associated code can be found at https://github.com/neuropoly/tract-clustering.

Assessment of Human Epineural Conduit of Different Size Diameters on Efficacy of Nerve Regeneration and Functional Outcomes.

Different types of nerve conduits are used to bridge peripheral nerve gaps when a tension-free repair is unattainable. To best support nerve regeneration, naturally occurring conduits have been tested. Since allografts offer an unlimited source of epineurium, we have developed human epineural conduit (hEC) as a novel technology to bridge nerve gaps. Considering acellular properties, and lack of immunogenic response, epineurium-derived conduits represent an attractive material, when compared with nerve allografts that require systemic immunosuppression. In this study, we introduce the hEC as a novel naturally occurring material applied for repair of nerve gaps after trauma. We tested the application of hEC created from human sciatic nerve in the restoration of 20 mm sciatic nerve defects in the nude rat model. Four experimental groups were studied: group 1: no repair control (n = 6), group 2: autograft control (n = 6), group 3: matched diameter hEC (n = 6), and group 4: large diameter hEC (n = 6). Functional tests of toe-spread and pin prick were performed at 1, 3, 6, 9, 12 weeks after repair. At 12 weeks, nerve samples were collected for immunostaining of Laminin B, S-100, glial fibrillary acidic protein (GFAP), nerve growth factor (NGF), vascular endothelial growth factor (VEGF), von Willebrand factor, and histomorphometric analysis of myelin thickness, axonal density, fiber diameter, and percentage of the myelinated nerve fibers. Muscle samples were gathered for gastrocnemius muscle index (GMI) and muscle fiber area ratio measurements. Best functional recovery, as well as GMI, was revealed for the autograft group, and was comparable to the matched hEC group. Significant differences were revealed between matched and large hEC groups in expression of S100 (p = 0.0423), NGF (p = 0.269), VEGF (p = 0.0003) as well as in percentage of myelinated fibers (p < 0.001) and axonal density (p = 0.0003). We established the feasibility of hEC creation. The innovative method introduces an alternative technique to autograft repair of nerve defects.

Autophagy Dysfunction in a Diabetic Peripheral Neuropathy Model.

The relationship between autophagy and diabetic peripheral neuropathy (DPN) has been highlighted in few reports. Using an animal model, the authors investigated the relationship between autophagy and DPN, focused particularly on changes in autophagy in Schwann cells. The ultrastructural features of DPN mice were evaluated in vivo using transmission electron microscopy. Dysfunction of autophagy in DPN was evaluated using immunofluorescence microscopy and Western blot analysis of proteins related to autophagy, including Beclin1, LC3, and p62. Reactive oxygen species levels were measured in vitro in glucose-treated Schwann cells. Dysfunction of autophagy in glucose-treated Schwann cells was examined by immunofluorescence microscopy and Western blot analysis. Reduced myelin thickness and axonal shrinkage were observed in the sciatic nerves of DPN mice. Reactive oxygen species levels were increased in Schwann cells treated with high glucose ( P < 0.05). The expression of Beclin1 was increased in DPN mice and Schwann cells treated with high glucose ( P < 0.05), whereas the expression of LC3-II/LC3-I ratio and p62 were decreased in DPN mice and Schwann cells treated with high glucose ( P < 0.05). These results suggest that increased levels of reactive oxygen species induced by high glucose may contribute to autophagy dysfunction in Schwann cells. Autophagy dysfunction especially in Schwann cells may be an underlying cause of DPN. This study presents the pathological mechanism of diabetic peripheral neuropathy.

Heterogeneity and regulation of oligodendrocyte morphology.

Oligodendrocytes form multiple myelin sheaths in the central nervous system (CNS), which increase nerve conduction velocity and are necessary for basic and higher brain functions such as sensory function, motor control, and learning. Structures of the myelin sheath such as myelin internodal length and myelin thickness regulate nerve conduction. Various parts of the central nervous system exhibit different myelin structures and oligodendrocyte morphologies. Recent studies supported that oligodendrocytes are a heterogenous population of cells and myelin sheaths formed by some oligodendrocytes can be biased to particular groups of axons, and myelin structures are dynamically modulated in certain classes of neurons by specific experiences. Structures of oligodendrocyte/myelin are also affected in pathological conditions such as demyelinating and neuropsychiatric disorders. This review summarizes our understanding of heterogeneity and regulation of oligodendrocyte morphology concerning central nervous system regions, neuronal classes, experiences, diseases, and how oligodendrocytes are optimized to execute central nervous system functions.

Enlargement of the Nerve Fibers of Silenced Lumbosacral Motoneurons in Cats.

Whether neuromuscular activity influences the size of motor nerves is controversial. All neuromuscular activity in cat hindlimbs was eliminated by spinal cord isolation (SCI), namely, spinal cord transection above and below the medial gastrocnemius (MG) and soleus (SOL) motoneuron pools and L5-S3 dorsal root transection. MG, SOL and sural (SUR) nerves were removed for size measurements, eight months after SCI surgery and from age-matched control cats. Nerve fiber number, the linear relationship between axon size and myelin thickness, and the bimodal distributions of nerve fiber area and diameter were maintained in all three nerves after SCI. The distributions of myelinated sensory fibers were unchanged in SUR nerves in contrast to the myelinated motor fibers in the MG and SOL nerves that were significantly larger. These findings provide evidence that all lumbar motoneurons survive SCI and that their nerve fibers enlarge. Thus, motor nerve fiber size in addition to the properties of the motoneurons and their muscle fibers is dynamic, responding to neuromuscular activity.