Cells Of Peripheral Nervous System Research Articles

Migraine and major depression: localizing shared genetic susceptibility in different cell types of the nervous systems.

There is a bidirectional relationship between migraine and major depression disorder (MDD). They likely share important risk genes associated with different cell types in the central nervous system (CNS) and peripheral nervous system (PNS). Profiling the expression of these genes in specific cell types is critical in understanding the pathophysiology of the relationship between migraine and MDD. Associated genes shared by migraine and MDD were identified by consolidating multiple curations of human disease-gene associations. Subsequently, the expression of overlapping genes was profiled and compared across the different cell types in CNS, PNS and neurovascular cells using eight single cell RNA sequencing datasets, including two human CNS datasets, two mouse CNS datasets, one human PNS dataset and three mouse PNS datasets. 45 shared genes between migraine and MDD were identified. Consistently found in all eight datasets, dopaminergic and serotonergic neurotransmitters were broadly expressed in CNS and PNS cell types. Glutamatergic and endocannabinoid genes were specifically expressed in CNS neurons and astrocytes. Synthesis and/or Release and Binding of Neuropeptides were specifically expressed in PNS peptidergic nociceptor (PEP). Genes related to inflammatory factors and immune responses were specifically expressed in CNS microglia. Among which, IL1B and COMT were highly expressed in CNS microglia cells. Single cell RNA sequencing of the CNS and PNS helps to identify the shared genes between migraine and MDD that are enriched in specific cell types. The findings provide new insight in understanding the underlying mechanism of action for the bidirectional co-morbidity between migraine and MDD.

Malignant Peripheral Nerve Sheath Tumors Activate Distinct Immunosuppressive Pathways Following Radiotherapy and are Associated with Immune Depletion In Vivo

Patients with neurofibromatosis type I, caused by NF1 loss, develop benign plexiform neurofibromas (pNF) in their peripheral nervous system (PNS). Malignant transformation of pNFs into malignant peripheral nerve sheath tumors (MPNSTs) occurs following CDKN2A/B and SUZ12 loss, a process associated with radiotherapy (RT). However, the molecular mechanisms underlying RT responses by different PNS cell types remain unclear. We hypothesized normal peripheral nerve cells, pNFs, and MPNSTs harbor distinct RT responses. Patient derived NF1 WT immortalized peripheral nerve cells (iPNs), NF1 mutant pNF cells, and NF1/CDKN2AB/SUZ12 mutant MPNST cells were used to study RT responses in vitro. CRISPRi was used to assess the functional effects of candidate gene repression. In vitro viability was measured by cell counts. Transcriptomic signatures were measured by bulk RNA-sequencing and integrated with single-cell RNA sequencing (scRNA-seq) data from patient-derived pNF and MPNST resection specimens. Radiation dose response curves revealed pNF cells (IC50 0.61 Gy) were more radiosensitive than MPNST cells (4.15 Gy). WT iPNs, NF1 deficient iPNs, and pNFs cells displayed no difference in cell viability (p = 0.67; t-test) following initiation of 2 Gy x 5 fractions, while MPNST cells were significantly more viable (p = 0.02; t-test). Principal component analysis of bulk RNA-sequencing data at 5 or 14 days following 2 Gy x 5 fractions revealed cell line of origin accounted for the greatest inter-sample variation (64.9% variance), with additional components separating samples based on radiation presence and timing. Using the most variable genes in PCA space to identify markers of RT response, iPNs and pNFs upregulated pro-apoptotic pathways (BAD, DAPK3) at 5 days post-radiation while MPNST cells alone upregulated pro-survival growth factor signaling). At 14 days post radiation, MPNST cells uniquely upregulated TGFβ signaling and interferon response circuits. Incorporation of scRNA-seq data revealed enrichment of growth factor signaling and TGFβ signatures in MPNSTs compared to pNF. Moreover, MPNST harbored significantly fewer immune cells than pNFs (p = 0.008, t-test), suggesting cell-autonomous signaling and crosstalk with the microenvironment are both critical to MPNST radioresistance. Our data indicate additional genetic hits beyond NF1 loss may be required for RT-associated malignant transformation of pNFs and radioresistance in MPNSTs. Analysis of transcriptomic responses to RT suggests that upregulated growth factor signaling and TGFβ-associated immunosuppression are distinct features of MPNST. Future work will focus on CRISPRi screens to unbiasedly nominate functional modifiers of RT response in NF1/CDKN2AB deficient tumors, which may be broadly useful in cancer.

Differentiation of peripheral sensory neurons from iPSCs derived from stem cells from human exfoliated deciduous teeth (SHED).

Peripheral nervous system (PNS) sensory alterations are present in several pathologies and syndromes. The use of induced pluripotent stem cell (iPSC) technology is an important strategy to produce sensory neurons in patients who are accomplished in terms of sensory symptoms. The iPSC technology relies on manipulating signaling pathways to resemble what occurs in vivo, and the iPSCs are known to carry a transcriptional memory after reprogramming, which can affect the produced cell. To this date, protocols described for sensory neuron production start using iPSCs derived from skin fibroblasts, which have the same ontogenetic origin as the central nervous system (CNS). Since it is already known that the cells somehow resemble their origin even after cell reprogramming, PNS cells should be produced from cells derived from the neural crest. This work aimed to establish a protocol to differentiate sensory neurons derived from stem cells from human exfoliated deciduous teeth (SHED) with the same embryonic origin as the PNS. SHED-derived iPSCs were produced and submitted to peripheral sensory neuron (PSN) differentiation. Our protocol used the dual-SMAD inhibition method, followed by neuronal differentiation, using artificial neurotrophic factors and molecules produced by human keratinocytes. We successfully established the first protocol for differentiating neural crest and PNS cells from SHED-derived iPSCs, enabling future studies of PNS pathologies.

Peripheral sensory neurons and non-neuronal cells express functional Piezo1 channels.

Here, we present evidence showing Piezo1 protein expression in the primary sensory neurons (PSNs) and non-neuronal cells of rat peripheral nervous system. Using a knockdown/knockout validated antibody, we detected Piezo1 immunoreactivity (IR) in ∼60% of PSNs of rat dorsal root ganglia (DRG) with higher IR density in the small- and medium-sized neurons. Piezo1-IR was clearly identified in DRG perineuronal glia, including satellite glial cells (SGCs) and Schwann cells; in sciatic nerve Schwann cells surrounding the axons and cutaneous afferent endings; and in skin epidermal Merkel cells and melanocytes. Neuronal and non-neuronal Piezo1 channels were functional since various cells (dissociated PSNs and SGCs from DRGs, isolated Schwann cells, and primary human melanocytes) exhibited a robust response to Piezo1 agonist Yoda1 by an increase of intracellular Ca2+ concentration ([Ca2+]i). These responses were abolished by non-specific Piezo1 antagonist GsMTx4. Immunoblots showed elevated Piezo1 protein in DRG proximal to peripheral nerve injury-induced painful neuropathy, while PSNs and SGCs from rats with neuropathic pain showed greater Yoda1-evoked elevation of [Ca2+]i and an increased frequency of cells responding to Yoda1, compared to controls. Sciatic nerve application of GsMTx4 alleviated mechanical hypersensitivity induced by Yoda1. Overall, our data show that Piezo1 is widely expressed by the neuronal and non-neuronal cells in the peripheral sensory pathways and that painful nerve injury appeared associated with activation of Piezo1 in PSNs and peripheral glial cells.

Inflammation in pathogenesis of chronic pain: Foe and friend.

Chronic pain is a refractory health disease worldwide causing an enormous economic burden on individuals and society. Accumulating evidence suggests that inflammation in the peripheral nervous system (PNS) and central nervous system (CNS) is the major factor in the pathogenesis of chronic pain. The inflammation in the early- and late phase may have distinctive effects on the initiation and resolution of pain, which can be viewed as friend or foe. On the one hand, painful injuries lead to the activation of glial cells and immune cells in the PNS, releasing pro-inflammatory mediators, which contribute to the sensitization of nociceptors, leading to chronic pain; neuroinflammation in the CNS drives central sensitization and promotes the development of chronic pain. On the other hand, macrophages and glial cells of PNS and CNS promote pain resolution via anti-inflammatory mediators and specialized pro-resolving mediators (SPMs). In this review, we provide an overview of the current understanding of inflammation in the deterioration and resolution of pain. Further, we summarize a number of novel strategies that can be used to prevent and treat chronic pain by controlling inflammation. This comprehensive view of the relationship between inflammation and chronic pain and its specific mechanism will provide novel targets for the treatment of chronic pain.

Shared Molecular Signatures Across Zika Virus Infection and Multiple Sclerosis Highlight AP-1 Transcription Factor as a Potential Player in Post-ZIKV MS-Like Phenotypes.

Zika virus (ZIKV) is an arbovirus of the Flaviviridae genus that has rapidly disseminated from across the Pacific to the Americas. Robust evidence has indicated a crucial role of ZIKV in congenital virus syndrome, including neonatal microcephaly. Moreover, emerging evidence suggests an association between ZIKV infection and the development of an extensive spectrum of central nervous system inflammatory demyelinating diseases (CNS IDD), such as multiple sclerosis-like clinical phenotypes. However, the underlying mechanisms of host-pathogen neuro-immune interactions remain to be elucidated. This study aimed to identify common transcriptional signatures between multiple sclerosis (MS) and ZIKV infection to generate molecular interaction networks, thereby leading to the identification of deregulated processes and pathways, which could give an insight of these underlying molecular mechanisms. Our investigation included publicly available transcriptomic data from MS patients in either relapse or remission (RR-MS) and datasets of subjects acutely infected by ZIKV for both immune peripheral cells and central nervous system cells. The protein-protein interaction (PPI) analysis showed upregulated AP-1 transcription factors (JUN and FOS) among the top hub and bottleneck genes in RR-MS and ZIKV data. Gene enrichment analysis retrieved a remarkable presence of ontologies and pathways linked to oxidative stress responses, immune cell function, inflammation, interleukin signaling, cell division, and transcriptional regulation commonly enriched in both scenarios. Considering the recent findings concerning AP-1 function in immunological tolerance breakdown, regulation of inflammation, and its function as an oxidative stress sensor, we postulate that the ZIKV trigger may contribute as a boost for the activation of such AP-1-regulated mechanisms that could favor the development of MS-like phenotypes following ZIKV infection in a genetically susceptible individual.

In Vitro Myelination of Peripheral Axons in a Coculture of Rat Dorsal Root Ganglion Explants and Schwann Cells.

The process of myelination is essential to enable rapid and sufficient signal transduction in the nervous system. In the peripheral nervous system, neurons and Schwann cells engage in a complex interaction to control the myelination of axons. Disturbances of this interaction and breakdown of the myelin sheath are hallmarks of inflammatory neuropathies and occur secondarily in neurodegenerative disorders. Here, we present a coculture model of dorsal root ganglion explants and Schwann cells, which develops a robust myelination of peripheral axons to investigate the process of myelination in the peripheral nervous system, study axon-Schwann cell interactions, and evaluate the potential effects of therapeutic agents on each cell type separately. Methodologically, dorsal root ganglions of embryonic rats (E13.5) were harvested, dissociated from their surrounding tissue, and cultured as whole explants for 3 days. Schwann cells were isolated from 3-week-old adult rats, and sciatic nerves were enzymatically digested. The resulting Schwann cells were purified by magnetic-activated cell sorting and cultured under neuregulin and forskolin-enriched conditions. After 3 days of dorsal root ganglion explant culture, 30,000 Schwann cells were added to one dorsal root ganglion explant in a medium containing ascorbic acid. The first signs of myelination were detected on day 10 of coculture, through scattered signals for myelin basic protein in immunocytochemical staining. From day 14 onward, myelin sheaths were formed and propagated along the axons. Myelination can be quantified by myelin basic protein staining as a ratio of the myelination area and axon area, to account for the differences in axonal density. This model provides experimental opportunities to study various aspects of peripheral myelination in vitro, which is crucial for understanding the pathology of and possible treatment opportunities for demyelination and neurodegeneration in inflammatory and neurodegenerative diseases of the peripheral nervous system.

Specific Cellular Effects of Low Bortezomib Concentrations on Purified Cultures of Schwann Cells, Satellite Glial Cells, Macrophages, and Dorsal Root Ganglion Neurons.

Peripheral neuropathy is one of the major adverse effects that limit the clinical application of bortezomib (BTZ). However, the underlying mechanisms of BTZ-induced peripheral neuropathy (BIPN) remain elusive. To examine cell types potentially involved in the development of BIPN, we used four purified cultures of cells of the peripheral nervous system: Schwann cells (SCs), satellite glial cells (SGCs), macrophages, and dorsal root ganglion (DRG) neurons. Administration of a low BTZ concentration (5 nM; similar to concentrations in clinical use) caused dedifferentiation of cultured SCs, returning mature SCs to an immature state. In cultured SGCs, BTZ increased glial fibrillary acidic protein (GFAP) levels without inducing the release of inflammatory cytokines or chemokines. In macrophages, BTZ caused little inflammatory response. Finally, in DRG neurons, BTZ strongly suppressed the expression levels of sensor and transducer ion channels without affecting cell morphology. Taken together, low concentrations of BTZ can cause SC dedifferentiation (i.e., demyelination), increased GFAP level in SGC, and decreased expression levels of sensor and transducer ion channels in DRG neurons (i.e., numbness feeling). Thus, we have reported, for the first time, specific effects of BTZ on peripheral nervous system cells, thereby contributing to a better understanding of the initiating mechanism of BIPN.

Using Human iPSC-Derived Peripheral Nervous System Disease Models for Drug Discovery.

Induced pluripotent stem cells (IPSCs), with their remarkable ability to differentiate into various cell types, including peripheral nervous system cells such as neurons and glial cells, offer an excellent platform for in vitro disease modeling. These iPSC-derived disease models have proven valuable in drug discovery, as they provide more precise simulations of a patient's disease state and allow for the assessment of potential therapeutic effectiveness and safety.

Single-cell RNA-seq reveals the communications between extracellular matrix-related components and Schwann cells contributing to the earlobe keloid formation.

Keloid is a major type of skin fibrotic disease, with one prominent feature of extensive accumulation of extracellular matrix (ECM) components, and another feature of pain/itching, which is closely related to the peripheral nervous system (PNS). However, the molecular pathogenesis of these two prominent features still needs to be further explored. In the present study, we performed single-cell RNA sequencing (scRNA-seq) on clinical earlobe keloid samples and adjacent normal skin samples and constructed a keloid atlas of 31,379 cells. All cells were clustered into 13 major cell types using cell-type-specific markers. Among them, fibroblast, vascular endothelial cells, and smooth muscle cells were defined as the ECM-related populations according to their ECM-associated functions. Also, we found that Schwann cells (SCs) were the main neuron cells of PNS in the skin. Interestingly, the cell proportions of ECM-related populations, as well as SC were increased significantly in the earlobe keloid compared to the adjacent normal tissues, suggesting an important role of these cell types in the development of the earlobe keloid. Comprehensive cell-cell interaction analysis at the single-cell level revealed a strong interaction between SC and ECM-related subgroups which might be mediated by SEMA3C signaling pathways and MK/PTN gene family, which are found to be mainly involved in promoting cell proliferation and migration. Moreover, further exploration of the interactions of ECM-related populations and SC in different keloids, including earlobe keloid, back keloid, and chest keloid revealed an increasing amount of TGFβ-TGFβ receptor interactions in chest/back keloids as compared to earlobe keloid, which suggested the anatomic site-specific pathogenesis in different keloids. Altogether, these findings suggested the interactions between ECM-related populations and SC contributing to the earlobe keloid formation and helped us to better understand the pathogenesis of keloids.

Mechanical stimulation of a bioactive, functionalized PVDF-TrFE scaffold provides electrical signaling for nerve repair applications.

Traumatic nerve injuries have limited success in achieving full functional recovery, with current clinical solutions often including implementation of nerve grafts or the use of nerve conduits to guide damaged axons across injury gaps. In search of alternative, and complimentary solutions, piezoelectric biomaterials demonstrate immense potential for tissue engineering applications. Piezoelectric poly(vinylidene fluoride-triflouroethylene) (PVFD-TrFE) scaffolds can be harnessed to non-invasively stimulate and direct function of key peripheral nervous system (PNS) cells in regeneration strategies. In this study, electrospun PVDF-TrFE was characterized, fabricated into a 3D scaffold, and finally rendered bioactive with the incorporation of a cell-secreted, decellularized extracellular matrix (dECM). PVDF-TrFE scaffolds were characterized extensively for piezoelectric capacity, mechanical properties, and cell-material interactions with fibroblasts and Schwann cells. Through functionalization of PVDF-TrFE scaffolds with a native, cell-assembled dECM, the ability to promote cell adhesion and enhanced viability was also demonstrated. Additionally, incorporation of bioactive functionalization improved the assembly of key regenerative ECM proteins and regenerative growth factors. PVDF-TrFE scaffolds were then fabricated into a conduit design that retained key physical, chemical, and piezoelectric properties necessary for PNS repair. This work shows great promise for multi-cue, electrospun biomaterials for regeneration of the PNS in traumatic injury.

P13. Transduction of the rat spinal cord using adenoassociated virus serotype 2.5

<h3>BACKGROUND CONTEXT</h3> Gene therapy is a method by which exogenous genes are introduced to treat disease or deliver drugs of interest. This is commonly done through either direct gene delivery in vivo or ex vivo through transduction of cells that are to be transplanted. There are several vectors for delivering genes, including viral vectors, naked plasmids, electroporation and lipid nanoparticles. Adeno-associated viruses (AAVs) are commonly used and considered to be well tolerated by the brain, muscles, nerves and multiple cell types. Rat Schwann cells are transduced efficiently by AAV1 whereas human Schwann cells are better transduced with AAV2 and 6. Rat nerves segments have demonstrated to be efficiently transduced by AAV1, 5, 7 and 9 whereas human nerves prefer AAV2. Many studies also agree that AAV1, 2, 4, 8 and 9 are optimal serotypes for CNS. We have utilized a novel viral vector, AAV serotype 2.5, that is a chimera of AAV2 vector modified with 5 amino acids from AAV1. AAV2.5 has shared characteristics of AAV1 and AAV2 with enhanced transduction for multiple cell types, such as in the muscle and articular tissue. Since AAV2.5 is not a naturally occurring virus, there is lower cross-reactivity to AAV2 antibodies. However, the ability of AAV2.5 to transduce tissues of the central (CNS) and peripheral nervous system (PNS) is not well understood. <h3>PURPOSE</h3> The purpose of this study is to determine the trophism and transduction efficiency of AAV2.5 for CNS and PNS tissue through in vitro transduction of dorsal root ganglion derived neurons, Schwann cells, and in vivo transduction of spinal cord. We predict that since AAV1 and AAV2 have known tropism for CNS and PNS cells to different extents depending on cell type, that the chimera AAV2.5 would be broadly effective. <h3>STUDY DESIGN/SETTING</h3> Cell culture and animal study. <h3>METHODS</h3> The cDNA encoding green fluorescent protein (GFP) was cloned into the conventional AAV packaging vector pTRUF2. For generation of self-complementing AAV vector plasmids, the cDNAs for GFP were directionally inserted into the SacII, NotI sites of pHpa-trs-SK plasmid. Transcription was driven by the cytomegalovirus promoter. Primary Schwann cells were isolated from the sciatic nerves of postnatal day 5 rat pups and dorsal root ganglion (DRG) neurons from embryonic day 15 rat pups. The cells were plated at a density of 125,000 cells per well and infected with 104 viral genomes per cell. For in vivo transduction of spinal cord tissue, 4 µL of 5 × 106– 5 × 1010vg/mL of AAV2.5-GFP was injected into 4 positions at T9. Immunostaining for markers of different spinal cord cell types colabeled with anti-GFP was done. <h3>RESULTS</h3> AAV2.5 was able to successfully infect DRG neurons and Schwann cells. AAV-2.5 vector was much more efficient than the commonly used AAV2 vector at transducing the Schwann cells. Cell survival and gene product production by AAV2.5 was greater than electroporation. When AAV2.5 -GFP was injected into the spinal cord there was a dose dependent increase in GFP expression by neurons and astrocytes up to 4 mm away from the injection site. <h3>CONCLUSIONS</h3> The study presents a novel vector for broad transduction in vivo or ex vivo of cells in the CNS and PNS. AAV2.5 was more efficient than the more commonly used AAV2 vector or electroporation. Since AAV2.5 is presently in phase 1 trials for osteoarthritis (NCT02790723), this vector type presents an interesting clinical translation opportunity for spinal cord and peripheral nerve injuries/diseases. <h3>FDA DEVICE/DRUG STATUS</h3> This abstract does not discuss or include any applicable devices or drugs.

Enhanced neurogenic differentiation on anisotropically conductive carbon nanotube reinforced polycaprolactone-collagen scaffold by applying direct coupling electrical stimulation

Electrical stimulation is conducive to neural regeneration. Different types of stimuli propagation patterns are required for regenerating cells in peripheral and central nervous systems. Modulation of the pattern of stimuli propagation cannot be achieved through external means. Reinforcing scaffolds, with suitably shaped conductive second phase materials, is a promising option in this regard. The present study has taken the effort of modulating the pattern (arrangement) of reinforced phase, namely multiwalled carbon nanotubes (MWCNT), in a biodegradable scaffold made of PCL-collagen mixture, by applying an external electric field during curing. Because of their extraordinary physical properties, MWCNTs have been selected as nano-reinforcement for this study. The nature of reinforcement affects the electrical conductivity of the scaffold and also determines the type of cell it can support for regeneration. Further, electrical stimulation, applied during incubation, was observed to have a positive influence on differentiating neural cells in vitro. However, the structure of the nano-reinforcement determined the differentiated morphology of the cells. Reinforced MWCNTs being tubes, imparted bipolarity to the cells. Therefore, these scaffolds, coupled with electrical stimulation possess significant potential to be used for directional regeneration of the nerves.

Spectrum of paired-like homeobox 2b immunoexpression in pediatric brain tumors with embryonal morphology.

Paired‐like homeobox 2b (PHOX2B) is an established immunomarker for peripheral neuroblastoma and autonomic nervous system cells. We aimed to evaluate the utility of PHOX2B immunostaining in central nervous system (CNS) tumors with embryonal morphology. Fifty‐one tumors were stained with PHOX2B and submitted for whole slide image analysis: 35 CNS tumors with embryonal morphology (31 CNS embryonal tumors and four gliomas); and 16 peripheral neuroblastomas were included for comparison. Diffuse nuclear immunopositivity was observed in all (16/16) neuroblastomas (primary and metastatic). Among CNS embryonal tumors, focal immunoreactivity for PHOX2B was observed in most (5/7) embryonal tumors with multilayered rosettes (ETMR) and a single high‐grade neuroepithelial tumor (HGNET) with PLAGL2 amplification; the remaining 27 CNS tumors were essentially immunonegative (<0.05% positive). Among ETMR, PHOX2B expression was observed in a small overall proportion (0.04%–4.94%) of neoplastic cells but focally reached up to 39% in 1 mm ‘hot spot’ areas. In the PLAGL2‐amplified case, 0.09% of the total neoplastic population was immunoreactive, with 0.53% in the ‘hot spot’ area. Care should be taken in interpreting PHOX2B immunopositivity in a differential diagnosis that includes metastatic neuroblastoma and CNS tumors; focal or patchy expression should not be considered definitively diagnostic of metastatic peripheral neuroblastoma.

Melatonin promotes peripheral nerve repair through Parkin-mediated mitophagy

Schwann cells (SCs) are the major glial cells in peripheral nervous system. They unsheathe and myelinate axons and play an essential role in peripheral nerve regeneration. Several studies report that Parkin-mediated mitophagy is associated with various diseases. Melatonin promotes proliferation of central glial cells. Little is known about the effect of melatonin and Parkin-mediated mitophagy on peripheral nerve repair.In this study, using a rat model of a peripheral nerve injury (PNI) and in vitro model established by RSC96 cells treated with tert-butyl hydroperoxide (TBHP), we found that Parkin-mediated mitophagy can effectively reduce the production of mitochondrial reactive oxygen species (ROS), maintain the balance of mitochondrial membrane potential, maintain autophagic flux, and inhibit mitochondrial apoptosis. At the same time, we found that the increase of Parkin under stress is a manifestation of the RSC96 cells' resistance to oxidative stress to maintain RSC96 cells' balance. In our experiment, melatonin is similar to a Parkin agonist, up-regulating the expression of Parkin, enhancing all the positive results of Parkin in a stress state, such as inhibiting active oxygen production, maintaining autophagic flux, and inhibiting mitochondrial apoptosis. In addition, we design in vivo experiments to verify in In vitro experiments. In in vivo, melatonin promotes the expression of Parkin, maintains autophagic flux, inhibits apoptosis, promotes myelin regeneration, reduces the regeneration of collagen fibers around damaged tissues, and promotes peripheral nerve repair. When adenovirus was used to down-regulate the expression of Parkin, we found that all the positive effects of melatonin were attenuated.Collectively, these findings indicate that melatonin upregulates Parkin-mediated mitophagy and promotes peripheral nerve repair. The results provide a basis for development of effective drugs for PNI treatment.

Glial regenerative response in the imaginal discs of Drosophila melanogaster.

Glial regenerative response in the imaginal discs of Drosophila melanogaster.

Hornerin deposits in neuronal intranuclear inclusion disease: direct identification of proteins with compositionally biased regions in inclusions

Neuronal intranuclear inclusion disease (NIID) is a neurodegenerative disorder, characterized by the presence of eosinophilic inclusions (NIIs) within nuclei of central and peripheral nervous system cells. This study aims to identify the components of NIIs, which have been difficult to analyze directly due to their insolubility. In order to establish a method to directly identify the components of NIIs, we first analyzed the huntingtin inclusion-rich fraction obtained from the brains of Huntington disease model mice. Although the sequence with expanded polyglutamine could not be identified by liquid-chromatography mass spectrometry, amino acid analysis revealed that glutamine of the huntingtin inclusion-rich fraction increased significantly. This is compatible with the calculated amino acid content of the transgene product. Therefore, we applied this method to analyze the NIIs of diseased human brains, which may have proteins with compositionally biased regions, and identified a serine-rich protein called hornerin. Since the analyzed NII-rich fraction was also serine-rich, we suggested hornerin as a major component of the NIIs. A specific distribution of hornerin in NIID was also investigated by Matrix-assisted laser desorption/ionization imaging mass spectrometry and immunofluorescence. Finally, we confirmed a variant of hornerin by whole-exome sequencing and DNA sequencing. This study suggests that hornerin may be related to the pathological process of this NIID, and the direct analysis of NIIs, especially by amino acid analysis using the NII-rich fractions, would contribute to a deeper understanding of the disease pathogenesis.

Dedifferentiated Schwann cell-derived TGF-β3 is essential for the neural system to promote wound healing.

Rationale: Wound healing is among the most complicated physiological processes and requires the synchronization of various cell types with distinct roles to re-establish the condition of the original skin. Patients affected by peripheral neuropathies often experience failure to heal. Loss of Schwann cells (SCs), a crucial population of peripheral nervous system cells in skin, may contribute to chronic wounds. However, the role of SCs in wound healing are poorly understood.Methods: The activity of SCs was investigated by using a cell atlas of the wound healing process, which was generated by integrating single-cell RNA sequencing (scRNA-seq) libraries covering different states of mouse back skin. The results of in silico analysis were validated by in vitro cell culture and in vivo mouse model. Selective inhibitors and conditional RNAi by virus transfection were utilized to investigate the role of SCs in wound healing. Findings from mouse experiments were further verified in scRNA-seq analysis of diabetic patients.Results: Our in silico analysis revealed the heterogeneous cellular components of skin and the dynamic interactions of neural crest derived cells (NCs) with other cell types. We found that SCs dedifferentiated at an early stage of wound repair with upregulated Wnt signaling. We also identified dedifferentiated SC (dSC) defect in diabetic wounds in both mouse and human. Wnt inhibition at the wound site repressed SC dedifferentiation, leading to defective repair. Furthermore, dSCs derived TGF-β3, which is context-dependent, promoted the migration of fibroblasts and keratinocytes. Moreover, TGF-β3 supplementation enhanced the healing of chronic wounds in diabetic mice with impaired SCs.Conclusion: Our study thus advances the understanding of the roles of neural-derived cells in skin regeneration and suggests a potential therapeutic strategy for wound healing disorders.

Neuron-fibrous scaffold interfaces in the peripheral nervous system: a perspective on the structural requirements.

The nerves of the peripheral nervous system are not able to effectively regenerate in cases of severe neural injury. This can result in debilitating consequences, including morbidity and lifelong impairments affecting the quality of the patient’s life. Recent findings in neural tissue engineering have opened promising avenues to apply fibrous tissue-engineered scaffolds to promote tissue regeneration and functional recovery. These scaffolds, known as neural scaffolds, are able to improve neural regeneration by playing two major roles, namely, by being a carrier for transplanted peripheral nervous system cells or biological cues and by providing structural support to direct growing nerve fibers towards the target area. However, successful implementation of scaffold-based therapeutic approaches calls for an appropriate design of the neural scaffold structure that is capable of up- and down-regulation of neuron-scaffold interactions in the extracellular matrix environment. This review discusses the main challenges that need to be addressed to develop and apply fibrous tissue-engineered scaffolds in clinical practice. It describes some promising solutions that, so far, have shown to promote neural cell adhesion and growth and a potential to repair peripheral nervous system injuries.

Anti-CD52 antibody treatment in murine experimental autoimmune encephalomyelitis induces dynamic and differential modulation of innate immune cells in peripheral immune and central nervous systems.

Anti‐CD52 antibody (anti‐CD52‐Ab) leads to a rapid depletion of T and B cells, followed by reconstitution of immune cells with tolerogenic characteristics. However, very little is known about its effect on innate immune cells. In this study, experimental autoimmune encephalomyelitis mice were administered murine anti‐CD52‐Ab to investigate its effect on dendritic cells and monocytes/macrophages in the periphery lymphoid organs and the central nervous system (CNS). Our data show that blood and splenic innate immune cells exhibited significantly increased expression of MHC‐II and costimulatory molecules, which was associated with increased capacity of activating antigen‐specific T cells, at first day but not three weeks after five daily treatment with anti‐CD52‐Ab in comparison with controls. In contrast to the periphery, microglia and infiltrating macrophages in the CNS exhibited reduced expression levels of MHC‐II and costimulatory molecules after antibody treatment at both time‐points investigated when compared to controls. Furthermore, the transit response of peripheral innate immune cells to anti‐CD52‐Ab treatment was also observed in the lymphocyte‐deficient SCID mice, suggesting the changes are not a direct consequence of the mass depletion of lymphocytes in the periphery. Our study demonstrates a dynamic and tissue‐specific modulation of the innate immune cells in their phenotype and function following the antibody treatment. The findings of differential modulation of the microglia and infiltrating macrophages in the CNS in comparison with the innate immune cells in the peripheral organs support the CNS‐specific beneficial effect of alemtuzumab treatment on inhibiting neuroinflammation in multiple sclerosis patients.