Myelination Process Research Articles

The Use of Myelinating Cultures as a Screen of Glycomolecules for CNS Repair.

In vitro cell-based assays have been fundamental in modern drug discovery and have led to the identification of novel therapeutics. We have developed complex mixed central nervous system (CNS) cultures, which recapitulate the normal process of myelination over time and allow the study of several parameters associated with CNS damage, both during development and after injury or disease. In particular, they have been used as a reliable screen to identify drug candidates that may promote (re)myelination and/or neurite outgrowth. Previously, using these cultures, we demonstrated that a panel of low sulphated heparin mimetics, with structures similar to heparan sulphates (HSs), can reduce astrogliosis, and promote myelination and neurite outgrowth. HSs reside in either the extracellular matrix or on the surface of cells and are thought to modulate cell signaling by both sequestering ligands, and acting as co-factors in the formation of ligand-receptor complexes. In this study, we have used these cultures as a screen to address the repair potential of numerous other commercially available sulphated glycomolecules, namely heparosans, ulvans, and fucoidans. These compounds are all known to have certain characteristics that mimic cellular glycosaminoglycans, similar to heparin mimetics. We show that the N-sulphated heparosans promoted myelination. However, O-sulphated heparosans did not affect myelination but promoted neurite outgrowth, indicating the importance of structure in HS function. Moreover, neither highly sulphated ulvans nor fucoidans had any effect on remyelination but CX-01, a low sulphated porcine intestinal heparin, promoted remyelination in vitro. These data illustrate the use of myelinating cultures as a screen and demonstrate the potential of heparin mimetics as CNS therapeutics.

A simple electrical stimulation cell culture system on the myelination of dorsal root ganglia and Schwann cells.

A large number of animal experiments and clinical trials have confirmed that electrical stimulation can accelerate the growth of axons and recovery of motor function, all of which are inseparable from the formation of myelin. Therefore, establishment of a suitable electrical stimulation platform to study the effects of electrical stimulation on the myelin process of dorsal root ganglia and Schwann cells is of great significance for understanding the recovery of electrical stimulation. We designed a simple conductive glass cell culture system to overcome the shortcomings of direct contact of the electrode with the culture solution, and the number of culture chambers can be selected based on the purpose of the experiment in order to reduce experimental time and cost.

3,5-T2 and 3,3\u2032,5-T3 Regulate Cerebellar Thyroid Hormone Signalling and Myelin Molecular Dynamics in Tilapia

In contrast to mammalian adults, myelination in teleosts occurs throughout their lifespan and most of the progenitor cells are originated in the cerebellum. To understand the role that thyroid hormones (THs) play in juvenile cerebellar myelination in teleosts, we identified and localised the expression of genes involved in TH signalling (mct8, oatp1c1, dio2, dio3, thraa and l-thrb1) and analysed the effects of the two bioactive THs, T2 and T3, upon their regulation, as well as upon some structural components of the myelination process. Ex vivo approaches using organotypic cerebellar cultures followed by FISH and qPCR showed gene-specific localisation and regulation of TH signalling genes in the cerebellar nuclei. In vivo approaches using methimazole (MMI)-treated juvenile tilapias replaced with low doses of T3 and T2 showed by immunofluorescence that myelin fibres in the cerebellum are more abundant in the granular layer and that their visible size is reduced after MMI treatment but partially restored with TH replacement, suggesting that low doses of TH promote the re-myelination process in an altered condition. Together, our data support the idea that T2 and T3 promote myelination via different pathways and prompt T2 as a target for further analysis as a promising therapy for hypomyelination.

The Potential of Stem Cell Therapy to Repair White Matter Injury in Preterm Infants: Lessons Learned From Experimental Models.

Diffuse white matter injury (dWMI) is a major cause of morbidity in the extremely preterm born infant leading to life-long neurological impairments, including deficits in cognitive, motor, sensory, psychological, and behavioral functioning. At present, no treatment options are clinically available to combat dWMI and therefore exploration of novel strategies is urgently needed. In recent years, the pathophysiology underlying dWMI has slowly started to be unraveled, pointing towards the disturbed maturation of oligodendrocytes (OLs) as a key mechanism. Immature OL precursor cells in the developing brain are believed to be highly sensitive to perinatal inflammation and cerebral oxygen fluctuations, leading to impaired OL differentiation and eventually myelination failure. OL lineage development under normal and pathological circumstances and the process of (re)myelination have been studied extensively over the years, often in the context of other adult and pediatric white matter pathologies such as stroke and multiple sclerosis (MS). Various studies have proposed stem cell-based therapeutic strategies to boost white matter regeneration as a potential strategy against a wide range of neurological diseases. In this review we will discuss experimental studies focusing on mesenchymal stem cell (MSC) therapy to reduce white matter injury (WMI) in multiple adult and neonatal neurological diseases. What lessons have been learned from these previous studies and how can we translate this knowledge to application of MSCs for the injured white matter in the preterm infant? A perspective on the current state of stem cell therapy will be given and we will discuss different important considerations of MSCs including cellular sources, timing of treatment and administration routes. Furthermore, we reflect on optimization strategies that could potentially reinforce stem cell therapy, including preconditioning and genetic engineering of stem cells or using cell-free stem cell products, to optimize cell-based strategy for vulnerable preterm infants in the near future.

Myelin and Modeling: Bootstrapping Cortical Microcircuits.

Histological studies of myelin-stained sectioned cadaver brain and in vivo myelin-weighted magnetic resonance imaging (MRI) show that the cerebral cortex is organized into cortical areas with generally well-defined boundaries, which have consistent internal patterns of myelination. The process of myelination is largely driven by neural experience, in which the axonal passage of action potentials stimulates neighboring oligodendrocytes to perform their task. This bootstrapping process, such that the traffic of action potentials facilitates increased traffic, suggests the hypothesis that the specific pattern of myelination (myeloarchitecture) in each cortical area reveals the principal cortical microcircuits required for the function of that area. If this idea is correct, the observable sequential maturation of specific brain areas can provide evidence for models of the stages of cognitive development.

Cyclin-dependent Kinase 18 Promotes Oligodendrocyte Precursor Cell Differentiation through Activating the Extracellular Signal-Regulated Kinase Signaling Pathway.

The correct differentiation of oligodendrocyte precursor cells (OPCs) is essential for the myelination and remyelination processes in the central nervous system. Determining the regulatory mechanism is fundamental to the treatment of demyelinating diseases. By analyzing the RNA sequencing data of different neural cells, we found that cyclin-dependent kinase 18 (CDK18) is exclusively expressed in oligodendrocytes. In vivo studies showed that the expression level of CDK18 gradually increased along with myelin formation during development and in the remyelination phase in a lysophosphatidylcholine-induced demyelination model, and was distinctively highly expressed in oligodendrocytes. In vitro overexpression and interference experiments revealed that CDK18 directly promotes the differentiation of OPCs, without affecting their proliferation or apoptosis. Mechanistically, CDK18 activated the RAS/mitogen-activated protein kinase kinase 1/extracellular signal-regulated kinase pathway, thus promoting OPC differentiation. The results of the present study suggest that CDK18 is a promising cell-type specific target to treat demyelinating disease.

Direct Binding of the Flexible C-Terminal Segment of Periaxin to β4 Integrin Suggests a Molecular Basis for CMT4F.

The process of myelination in the nervous system requires a coordinated formation of both transient and stable supramolecular complexes. Myelin-specific proteins play key roles in these assemblies, which may link membranes to each other or connect the myelinating cell cytoskeleton to the extracellular matrix. The myelin protein periaxin is known to play an important role in linking the Schwann cell cytoskeleton to the basal lamina through membrane receptors, such as the dystroglycan complex. Mutations that truncate periaxin from the C terminus cause demyelinating peripheral neuropathy, Charcot-Marie-Tooth (CMT) disease type 4F, indicating a function for the periaxin C-terminal region in myelination. We identified the cytoplasmic domain of β4 integrin as a specific high-affinity binding partner for periaxin. The C-terminal region of periaxin remains unfolded and flexible when bound to the third fibronectin type III domain of β4 integrin. Our data suggest that periaxin is able to link the Schwann cell cytoplasm to the basal lamina through a two-pronged interaction via different membrane protein complexes, which bind close to the N and C terminus of this elongated, flexible molecule.

Molecular Genetic Diversity and DNA Diagnostics of Hereditary Spastic Paraplegia

Abstract—Hereditary spastic paraplegia (HSP) is a heterogeneous group of neurodegenerative disorders predominantly characterized by damage to the pyramidal tract. The major, single symptom of HSP is progressive weakness and spasticity of lower extremities, which ultimately leads to difficulties in walking. HSP has various types of inheritance and clinical characteristics. The development of molecular genetics and the emergence of high-throughput sequencing techniques, such as New Generation Sequencing (NGS) in particular, revealed a pronounced genetic heterogeneity of HSP. Currently, about 80 genetic loci for HSP are designated as spastic paraplegia genes (SPGs) (with numbering according to the mapping history), and the vast majority of causative genes within loci have been identified. The most frequent autosomal dominant forms of HSP are SPG4 and SPG3; SPG11, SPG15, SPG7 are common autosomal recessive HSPs. The mechanisms of HSP molecular pathogenesis are variable and encompassing defects of neuronal membrane transport and the disturbance of myelination processes, lipid metabolism, mitochondrial dysfunctions, etc. The differential diagnosis for HSPs is difficult due to the existence of many geno- and phenocopies within the other neuropathologies that can be partially distinguished by neuroimaging methods. The only method for the precise diagnostics of HSP and the number of genomic copies is DNA diagnostics—the search for mutations in individual genes and/or the analysis of many genes simultaneously by NGS methods (gene panel, whole-exome sequencing, and whole-genome sequencing). Such DNA diagnostics opens up the opportunity to perform prenatal or preimplantation analysis of the fetus for families carrying HSP mutations to predict the risk of HSP inheritance. Currently, HSP treatment is directed against individual symptoms (antispastic drugs, etc.), and the discovery of the pathogenic action of different genes requires the development of new therapeutic approaches.

Benchmark on Automatic 6-month-old Infant Brain Segmentation Algorithms: The iSeg-2017 Challenge.

Accurate segmentation of infant brain magnetic resonance (MR) images into white matter (WM), gray matter (GM), and cerebrospinal fluid (CSF) is an indispensable foundation for early studying of brain growth patterns and morphological changes in neurodevelopmental disorders. Nevertheless, in the isointense phase (approximately 6-9 months of age), due to inherent myelination and maturation process, WM and GM exhibit similar levels of intensity in both T1-weighted (T1w) and T2-weighted (T2w) MR images, making tissue segmentation very challenging. Despite many efforts were devoted to brain segmentation, only few studies have focused on the segmentation of 6-month infant brain images. With the idea of boosting methodological development in the community, iSeg-2017 challenge (http://iseg2017.web.unc.edu) provides a set of 6-month infant subjects with manual labels for training and testing the participating methods. Among the 21 automatic segmentation methods participating in iSeg-2017, we review the 8 top-ranked teams, in terms of Dice ratio, modified Hausdorff distance and average surface distance, and introduce their pipelines, implementations, as well as source codes. We further discuss limitations and possible future directions. We hope the dataset in iSeg-2017 and this review article could provide insights into methodological development for the community.

Independent and cooperative roles of the Mek/ERK1/2-MAPK and PI3K/Akt/mTOR pathways during developmental myelination and in adulthood.

Multiple extracellular and intracellular signals regulate the functions of oligodendrocytes as they progress through the complex process of developmental myelination and then maintain a functionally intact myelin sheath throughout adult life, preserving the integrity of the axons. Recent studies suggest that Mek/ERK1/2-MAPK and PI3K/Akt/mTOR intracellular signaling pathways play important, often overlapping roles in the regulation of myelination. However, it remains poorly understood whether they function independently, sequentially, or converge using a common mechanism to facilitate oligodendrocyte differentiation, myelin growth, and maintenance. To address these questions, we analyzed multiple genetically modified mice and asked whether the deficits due to the conditional loss-of-function of ERK1/2 or mTOR could be abrogated by simultaneous constitutive activation of PI3K/Akt or Mek, respectively. From these studies, we concluded that while PI3K/Akt, not Mek/ERK1/2, plays a key role in promoting oligodendrocyte differentiation and timely initiation of myelination through mTORC1 signaling, Mek/ERK1/2-MAPK functions largely independently of mTORC1 to preserve the integrity of the myelinated axons during adulthood. However, to promote the efficient growth of the myelin sheath, these two pathways cooperate with each other converging at the level of mTORC1, both in the context of normal developmental myelination or following forced reactivation of the myelination program during adulthood. Thus, Mek/ERK1/2-MAPK and the PI3K/Akt/mTOR signaling pathways work both independently and cooperatively to maintain a finely tuned, temporally regulated balance as oligodendrocytes progress through different phases of developmental myelination into adulthood. Therapeutic strategies aimed at targeting remyelination in demyelinating diseases are expected to benefit from these findings.

Transferrin Enhances Microglial Phagocytic Capacity.

Transferrin (Tf) is a glycoprotein playing a critical role in iron homeostasis and transport and distribution throughout the body and within tissues and cells. This molecule has been shown to accelerate the process of myelination and remyelination in the central nervous system (CNS) in vivo and induce oligodendroglial cell maturation in vitro. While the mechanisms involved in oligodendroglial precursor cell (OPC) differentiation have not been fully elucidated yet, our group has previously described the first molecular events taking place in OPC in response to extracellular Tf. Here, we show the effect of Tf on the different glial cell populations. We demonstrate that, after a CNS demyelinating injury, Tf can be incorporated by all glial cells-i.e., microglia, astrocytes, and OPC-and that, acting on microglial cells in vitro, Tf increases microglial proliferation rates and phagocytic capacity. It may be then speculated that the in vivo correlation of this process could generate a favorable microenvironment for OPC maturation and remyelination.

Epigenetic regulation of myelination in health and disease.

Myelin is lipid-rich structure that is necessary to avoid leakage of electric signals and to ensure saltatory impulse conduction along axons. Oligodendrocytes in central nervous system (CNS) and Schwann cells in peripheral nervous system (PNS) are responsible for myelin formation. Axonal demyelination after injury or diseases greatly impairs normal nervous system function. Therefore, understanding how the myelination process is programmed, coordinated, and maintained is crucial for developing therapeutic strategies for remyelination in the nervous system. Epigenetic mechanisms have been recognized as a fundamental contributor in this process. In recent years, histone modification, DNA modification, ATP-dependent chromatin remodeling, and non-coding RNA modulation are very active area of investigation. We will present a conceptual framework that integrates crucial epigenetic mechanisms with the regulation of oligodendrocyte and Schwann cell lineage progression during development and myelin degeneration in pathological conditions. It is anticipated that a refined understanding of the molecular basis of myelination will aid in the development of treatment strategies for debilitating disorders that involve demyelination, such as multiple sclerosis in the CNS and neuropathies in the PNS.

Therapeutic potential of conditioned medium derived from oligodendrocytes cultured in a self-assembling peptide nanoscaffold in experimental autoimmune encephalomyelitis

The use of neurotrophic factors is considered to be a novel therapeutic approach for restoring and/or maintaining neurological function in neurodegenerative disorders, such as multiple sclerosis (MS). Various studies have shown that conditioned medium produced by oligodendrocyte (OL-CM) contain a variety of neurotrophic factors. Here, we investigated the restorative effects of OL-CM, collected from oligodendrocytes cultured in a self-assembling peptide hydrogels scaffold (PuraMatrix), in experimental autoimmune encephalomyelitis (EAE) mouse model. Neural stem/progenitor cells, isolated from the embryonic mouse brain, were cultured and differentiated into oligodendrocyte. Cell viability and proliferation of oligodendrocytes were assessed by live/dead and MTT assays. Motor functions, myelination, cell infiltration, gliosis, and inflammatory process were assessed in EAE mice after intracranial injection of OL-CM at different concentrations. Application of OL-CM improved clinical score and neurological function in EAE mice and reduced the inflammatory cell infiltration and demyelination. Furthermore, administration of OL-CM reduced the expression of pro-inflammatory cytokines and suppressed the activation of NLRP3-inflammasome complex in EAE mice. These data suggest the potential therapeutic effect of OL-CM for MS treatment.

Effect of the CSF1R inhibitor PLX3397 on remyelination of corpus callosum in a cuprizone-induced demyelination mouse model.

Multiple sclerosis (MS) is a chronic inflammatory disease affecting the central nervous system (CNS). Despite introducing multiple immunomodulatory approaches for MS, there are still major concerns about possible ways for improving remyelination in this disease. Microglia exert essential roles in regulation of myelination processes, and interaction between colony-stimulating factor 1 (CSF1) with its receptor CSF1R is considered as a key regulator of microglial differentiation and survival. The aim of this study was to investigate possible roles for a CSF1R inhibitor PLX3397 in recovery of central myelination processes. Chronic demyelination was induced in mice by addition of 0.2% cuprizone to the chow for 12 weeks. Next, animals were undergoing a diet containing 290 mg/kg PLX3397 to induce microglial ablation. The PLX3397 treatment caused a significant decrease in the rate of expression for the CSF1/CSF1R axis, and a reduction in the protein expressions for the microglial marker Iba-1 and for the oligodendrocyte marker Olig-2. Findings from Luxol fast blue (LFB) staining and transmission electron microscopy (TEM) showed an increase in the rate of myelination for the mice receiving PLX3397. The rate of destruction in the nerve fibers and the extent of the gaps formed between layers of myelin sheaths was also reduced after the treatment with PLX3397. In addition, animals experienced an improvement in recovery of motor deficit after receiving PLX3397 (for all P < 0.05). It could be concluded that PLX3397 could retain myelination in the MS model possibly through regulation of the myelin environment.

Hierarchically aligned gradient collagen micropatterns for rapidly screening Schwann cells behavior.

To penetrate the effect of protein gradient micropattern on peripheral nerve regeneration, the hierarchically aligned gradient collagen micropattern was prepared by micromoulding method and the influence on Schwann cells growth behavior was studied. The morphology, wettability, stability and component variation of the micropatterns were firstly characterized. Then, Schwann cells were cultured and the related mechanism was penetrated. The results showed that the gradient collagen micropattern could be well fabricated. The surface wettability varied with the change of collagen concentration, and the prepared gradient micropattern showed a good stability after PBS immersion for 15 days. The results of Schwann cells culture and morphological index analysis displayed that the prepared gradient collagen micropatten could well regulate the orientation growth of Schwann cells, while a much better cell alignment growth was obtained on the gradient micropattern with higher collagen concentration and wider pattern size. PCR and WB showed that the micropattern structure could effectively up-regulate the key specific genes for axon regeneration and myelination process. Overall, the study provides a systematic and facile method for understanding the effect of various sized micropatterns on cell behavior, which may have a great significance for the development of artificial implants for tissue engineering and regenerative medicine.

Evaluation of apamin effects on myelination process in C57BL/6 mice model of multiple sclerosis.

Multiple sclerosis (MS) is a demyelinating disease that causes chronic inflammation in the central nervous system. The aim of this study was to investigate the effects of apamin administration on myelination process. MS was induced by feeding cuprizone pellets (0.2%) for 6 weeks (demyelination phase) followed by normal feeding for additional 2 weeks (remyelination phase). Briefly, C57BL/6 male mice were randomly divided into six groups. Group 1, received the regular food pellets. Group 2 contained two subgroups of 6 animals each (n = 2 × 6). First group received cuprizone for 6 weeks and the sacrificed while the second group after 6 weeks of cuprizone, received no treatment for additional 2 weeks. Group 3 (co-treatment group) was composed of two subgroups of 6 animals each (n = 2 × 6). Both subgroups received apamin (100 μg/kg) intraperitoneally twice a week for 6 weeks. First subgroup terminated at this time and the second subgroup was fed normal diet for two additional weeks. Group 4 (post-treatment, n = 6) received apamin (100 μg/kg) intraperitoneally twice a week for 2 weeks after cuprizone secession. Groups 5 and 6 (vehicle, n = 6 in each group) received phosphate buffered saline as the vehicle of apamin during demyelination and remyelination phase. At the end of each phase, mice were deeply anesthetized and perfused. Groups 5 and 6 (vehicle) received PBS as the vehicle during both phases. Mice were anesthetized, perfused with PBS through their heart, and their brains were removed. Brain sections stained with luxol fast blue and the images were analyzed. Apamin co-treatment significantly increased the myelin content as compared to the cuprizone group. Also, mild elevation in the myelinated areas was observed with apamin post-treatment in comparison with remyelination phase. Our results revealed that apamin prevents myelin destruction more significantly as compared to remyelination process. This observation explains the possible role of apamin in inhibiting the activation of the microglia cells than stimulation of the oligodendrocytic precursor cells.

Recent Advances in Live Imaging of Cells of the Oligodendrocyte Lineage.

Myelination is an important process that takes place also in the periphery during development and in theadulthood. Myelin serves as an electric isolator for axons, leading to a fast conduction of the action potential, and provides trophic support for the axon, both aspects highly important for the proper function of the nervous system. In the central nervous system, myelination starts shortly after birth and cells from the oligodendrocyte lineage tightly regulate this process during the whole life span. Initially, it was thought that under physiological conditions myelin generation only occurs in early postnatal development and that myelination stops at early adult ages. Historically, the process of myelination has mainly been studied in fixed tissue, and predominantly analyzed by electron microscopy, bringing valuable insights in the structure and distribution of myelin in the central nervous system. Nevertheless, the outdated notion of the static nature of myelin during adulthood was challenged in the past decades by the development of new techniques bringing in a new picture of a lively structure that is in constant remodeling under physiological and disease conditions. As fixed tissue can only provide information at a specific timepoint, the necessity of new techniques to study this process in vivo has become clear. In this chapter, we will review some of the latest techniques developed in order to study myelin and the oligodendrocyte lineage, as these cells are important for the formation and restructuration of the myelin. We will also introduce a protocol to prepare a cranial window to study NG2-glia (also known as oligodendrocyte progenitor cells) of the cerebral cortex in vivo, by 2-photon laser scanning microscopy. However, this technique can also be performed to study other cell populations or structures such as myelin, which will be discussed in this chapter as well. Despite being simple, this protocol has shown to be powerful to study the oligodendrocyte lineage and potentially is applicable to study myelin in vivo, which could turn into a key technique in the understanding of myelination and other functions that the oligodendrocyte lineage might have under physiological and disease conditions.

Study of Myelin Gene Expression in the Central Nervous System Using Real-Time PCR.

Myelin sheaths are crucial for the survival and maintenance of the axons and the rapid propagation of the action potential. The glial cells involved are Schwann cells in the peripheral nervous system (PNS) and oligodendrocytes in the central nervous system (CNS). One oligodendrocyte may myelinate over 40 axons. In the CNS, myelin is composed of several layers of cytoplasmic membrane from oligodendrocytes stabilized by structural myelin-specific proteins such as proteolipid protein (PLP) and myelin basic protein (MBP). Those genes are expressed during myelination and then silenced. They can be re-expressed after demyelinating episodes, where they contribute to remyelination. Demyelination occurs after injuries of the CNS such as traumatic brain injury or during acute episodes of neurodegeneration observed in demyelinating and neurodegenerative diseases. Remyelination process is achieved by oligodendrocytes newly generated following the recruitment and differentiation of oligodendrocyte precursor cells (OPCs). Failure of remyelination process leads to irreversible axonal loss, functional impairment, and finally decreased cognitive performances. Several techniques have been described to study myelination and remyelination in culture systems. In this chapter, we explain how we can study myelin genes' expression in oligodendrocytes by real-time polymerase chain reaction (RT-PCR) using specific primers for plp and mbp. This technique can be crucial and prompt to determine the effect of specific chemicals (such as pesticides) on the myelination process in oligodendrocytes.

Myelin Ultrastructure Terminology in Disease and Recovery Processes.

Ultrastructural evaluation of myelin coat helps to understand the possible background of pathological changes leading to deterioration or complete loss of nerve functions. A number of terms were previously introduced to describe the fine structural changes in myelin under various conditions. We believe that using a common terminology will be helpful to interpret the structure/function relationship in neurological disorders empowering the diagnosis and possible therapeutical approaches. In this paper, we present examples of ultrastructural changes in myelin during myelination, demyelination, re-myelination and dysmyelination processes and we reviewed the terminology previously used.We tried to include all studies reporting ultrastructural findings with no limitation to the experimental conditions, the species used and the disorders. The terminology used to describe the structural findings included compacted myelin, partially compacted myelin, noncompacted myelin, redundancy (hypermyelination, tomacula, myelinosome), splitting, complete circular splitting, myelin degradation, concentric lamellar bodies (myelin figures), loss of myelin lamellae, polyaxonal Schwann cells and necrotic cell debris.Ultrastructural data described in this paper aimed to provide a guide for future studies. We concluded that the evaluation of ultrastructural changes in any neurological disorder is greatly valuable for a better understanding of pathological and physiological changes occured. We also believe that supporting the ultrastructural findings with quantitative methods in the future will be of great value.

A method to deliver patterned electrical impulses to Schwann cells cultured on an artificial axon

Information from the brain travels back and forth along peripheral nerves in the form of electrical impulses generated by neurons and these impulses have repetitive patterns. Schwann cells in peripheral nerves receive molecular signals from axons to coordinate the process of myelination. There is evidence, however, that non-molecular signals play an important role in myelination in the form of patterned electrical impulses generated by neuronal activity. The role of patterned electrical impulses has been investigated in the literature using co-cultures of neurons and myelinating cells. The co-culturing method, however, prevents the uncoupling of the direct effect of patterned electrical impulses on myelinating cells from the indirect effect mediated by neurons. To uncouple these effects and focus on the direct response of Schwann cells, we developed an in vitro model where an electroconductive carbon fiber acts as an artificial axon. The fiber provides only the biophysical characteristics of an axon but does not contribute any molecular signaling. In our “suspended wire model”, the carbon fiber is suspended in a liquid media supported by a 3D printed scaffold. Patterned electrical impulses are generated by an Arduino 101 microcontroller. In this study, we describe the technology needed to set-up and eventually replicate this model. We also report on our initial in vitro tests where we were able to document the adherence and ensheath of human Schwann cells to the carbon fiber in the presence of patterned electrical impulses (hSCs were purchased from ScienCell Research Laboratories, Carlsbad, CA, USA; ScienCell fulfills the ethic requirements, including donor’s consent). This technology will likely make feasible to investigate the response of Schwann cells to patterned electrical impulses in the future.