Schwann Cell Dedifferentiation Research Articles

Neuritin accelerates Schwann cell dedifferentiation via PI3K/Akt/mTOR signalling pathway during Wallerian degeneration.

Neuritin, also known as candidate plasticity gene 15 (CPG15), was first identified as one of the activity-dependent gene products in the brain. Previous studies have been reported that Neuritin induces neuritogenesis, neurite arborization, neurite outgrowth and synapse formation, which are involved in the development and functions of the central nervous system. However, the role of Neuritin in peripheral nerve injury is still unknown. Given the importance and necessity of Schwann cell dedifferentiation response to peripheral nerve injury, we aim to investigate the molecular mechanism of Neuritin steering Schwann cell dedifferentiation during Wallerian degeneration (WD) in injured peripheral nerve. Herein, using the explants of sciatic nerve, an exvivo model of nerve degeneration, we provided evidences indicating that Neuritin vividly accelerates Schwann cell dedifferentiation. Moreover, we found that Neuritin promotes Schwann cell demyelination as well as axonal degeneration, phagocytosis, secretion capacity. In summary, we first described Neuritin acts as a positive regulator for Schwann cell dedifferentiation and WD after peripheral nerve injury.

Targeting PI3K/Akt/mTOR signaling in rodent models of PMP22 gene-dosage diseases

Haplo-insufficiency of the gene encoding the myelin protein PMP22 leads to focal myelin overgrowth in the peripheral nervous system and hereditary neuropathy with liability to pressure palsies (HNPP). Conversely, duplication of PMP22 causes Charcot-Marie-Tooth disease type 1A (CMT1A), characterized by hypomyelination of medium to large caliber axons. The molecular mechanisms of abnormal myelin growth regulation by PMP22 have remained obscure. Here, we show in rodent models of HNPP and CMT1A that the PI3K/Akt/mTOR-pathway inhibiting phosphatase PTEN is correlated in abundance with PMP22 in peripheral nerves, without evidence for direct protein interactions. Indeed, treating DRG neuron/Schwann cell co-cultures from HNPP mice with PI3K/Akt/mTOR pathway inhibitors reduced focal hypermyelination. When we treated HNPP mice in vivo with the mTOR inhibitor Rapamycin, motor functions were improved, compound muscle amplitudes were increased and pathological tomacula in sciatic nerves were reduced. In contrast, we found Schwann cell dedifferentiation in CMT1A uncoupled from PI3K/Akt/mTOR, leaving partial PTEN ablation insufficient for disease amelioration. For HNPP, the development of PI3K/Akt/mTOR pathway inhibitors may be considered as the first treatment option for pressure palsies.

A materiobiology-inspired sericin nerve guidance conduit extensively activates regeneration-associated genes of Schwann cells for long-gap peripheral nerve repair

Peripheral nerve injury (PNI) is one of the most common causes of adult disability. Following PNI, Schwann cells (SCs) dedifferentiate, proliferate, and upregulate a variety of regeneration-associated genes (RAGs), including neurotrophic factors, ultimately promoting nerve regeneration. Even though certain reported biomaterials could upregulate one or several neurotrophic factors of SCs, none of them was able to activate these RAGs extensively. Herein, this work represents a novel manganese organic framework material (Mn-MOF) capable of effectively upregulating RAGs of SCs. In vitro cell coculture assay revealed that these Mn-MOF treated SCs enhanced the axon extension of PC12 cells, further confirming the axon regeneration-promoting activity of this framework. STAT3 and C-JUN signaling pathways were involved in Mn-MOF-induced RAGs activation. Using sericin, a well-proved biomaterial with neuroprotective properties, a Mn-MOF modified sericin conduit (MOF-SS) was prepared via π-π interactions between the cyclobenzene of framework materials and the aromatic acid of natural proteins. The microstructures and physiochemical properties of MOF-SS were comprehensively characterized, and its RAGs-activating effect on SCs was well confirmed. When applied in vivo to bridge 13-mm sciatic nerve defect, this conduit significantly promoted nerve microstructural regeneration, enhanced electrical conduction performance of regenerated nerve, improved functional recovery, and more importantly, alleviated muscle atrophy. In summary, this work represents a novel Mn-MOF modified sericin conduit capable of extensively activating RAGs of SCs, significantly promoting long-gap PNI repair.

Perspectives on the Novel Multifunctional Nerve Guidance Conduits: From Specific Regenerative Procedures to Motor Function Rebuilding.

Peripheral nerve injury potentially destroys the quality of life by inducing functional movement disorders and sensory capacity loss, which results in severe disability and substantial psychological, social, and financial burdens. Autologous nerve grafting has been commonly used as treatment in the clinic; however, its rare donor availability limits its application. A series of artificial nerve guidance conduits (NGCs) with advanced architectures are also proposed to promote injured peripheral nerve regeneration, which is a complicated process from axon sprouting to targeted muscle reinnervation. Therefore, exploring the interactions between sophisticated NGC complexes and versatile cells during each process including axon sprouting, Schwann cell dedifferentiation, nerve myelination, and muscle reinnervation is necessary. This review highlights the contribution of functional NGCs and the influence of microscale biomaterial architecture on biological processes of nerve repair. Progressive NGCs with chemical molecule induction, heterogenous topographical morphology, electroactive, anisotropic assembly microstructure, and self-powered electroactive and magnetic-sensitive NGCs are also collected, and they are expected to be pioneering features in future multifunctional and effective NGCs.

Iron role paradox in nerve degeneration and regeneration.

Iron accumulates in the neural tissue during peripheral nerve degeneration. Some studies have already been suggested that iron facilitates Wallerian degeneration (WD) events such as Schwann cell de-differentiation. On the other hand, intracellular iron levels remain elevated during nerve regeneration and gradually decrease. Iron enhances Schwann cell differentiation and axonal outgrowth. Therefore, there seems to be a paradox in the role of iron during nerve degeneration and regeneration. We explain this contradiction by suggesting that the increase in intracellular iron concentration during peripheral nerve degeneration is likely to prepare neural cells for the initiation of regeneration. Changes in iron levels are the result of changes in the expression of iron homeostasis proteins. In this review, we will first discuss the changes in the iron/iron homeostasis protein levels during peripheral nerve degeneration and regeneration and then explain how iron is related to nerve regeneration. This data may help better understand the mechanisms of peripheral nerve repair and find a solution to prevent or slow the progression of peripheral neuropathies.

Leprosy-specific subsets of macrophages and Schwann cells identified by single-cell RNA-sequencing

In Mycobacterium leprae (M. leprae)-infection, inflammatory cells’ subsets and dynamics as well as the interactions with Schwann cells have remained elusive. We investigated individual cells in M. leprae-inoculated nude mice by single-cell RNA-sequencing (scRNA-seq). For macrophages, we dissected two M1-like subsets and five M2-like subsets, where lipid-associated signatures were pervasive in both M1-like and M2-like subsets. There were four macrophage trajectories showing: (i) pro-inflammatory (M1), (ii) lipid metabolism-related (M2), (iii) anti-inflammatory (M2), and (iv) interferon-stimulated gene-related (M2) fates. They displayed early divergence without ever rejoining along the paths, suggesting simultaneous or continuous stimuli for macrophage activation in leprosy. The scRNA-seq predicted Schwann cell-macrophage interactions (Notch1-Jag1, Plxnb1-Sema4d interactions). An immature Schwann cell subset showing Tfap2a expression was identified, indicating Schwann cell dedifferentiation in leprosy tissues. Expressions of Notch1, Jag1, Plxnb1, Sema4d, and Tfap2a were validated in mouse or human leprosy tissues by immunohistochemistry. We identified both pro-inflammatory and inflammation-resolution signatures, where lipid-associated signatures were pervasive to the macrophages, representing leprosy-specific macrophage states for prolonged and repeated episodes of inflammation and resolution. Our study identified refined molecular states and interactions of macrophages and Schwann cells, suggesting novel insights into the pathogenesis of unhealed inflammation with neuropathy and potential therapeutic targets for leprosy.

PGC-1α Inhibits Schwann Cell Dedifferentiation and Delays Peripheral Nerve Degeneration by Targeting PON1.

PPARγ coactivator-1 alpha (PGC-1α) is an essential transcription factor co-activator that regulates gene transcription and neural regeneration. Schwann cells, which are unique glial cells in peripheral nerves that dedifferentiate after peripheral nerve injury (PNI) and are released from degenerative nerves. Wallerian degeneration is a series of stereotypical events that occurs in response to nerve fibers after PNI. The role of PGC-1α in Schwann cell dedifferentiation and Wallerian degeneration is not yet clear. As Wallerian degeneration plays a crucial role in PNI, we conducted a study to determine whether PGC-1α has an effect on peripheral nerve degeneration after injury. We examined the expression of PGC-1α after sciatic nerve crush or transection using Western blotting and found that PGC-1α expression increased after PNI. Then we utilized ex vivo and in vitro models to investigate the effects of PGC-1α inhibition and activation on Schwann cell dedifferentiation and nerve degeneration. Our findings indicate that PGC-1α negatively regulates Schwann cell dedifferentiation and nerve degeneration. Through the use of RNA-seq, siRNA/plasmid transfection and reversal experiments, we identified that PGC-1α targets inhibit the expression of paraoxonase 1 (PON1) during Schwann cell dedifferentiation in degenerated nerves. In summary, PGC-1α plays a crucial role in preventing Schwann cell dedifferentiation and its activation can reduce peripheral nerve degeneration by targeting PON1. PGC-1α inhibits Schwann cell dedifferentiation and peripheral nerve degeneration. PGC-1α negatively regulates Schwann cell dedifferentiation and peripheral nerve degeneration after injury by targeting PON1.

Transcription Factor BCL11A Regulates Schwann Cell Behavior During Peripheral Nerve Regeneration.

Nerve injury-induced Schwann cell dedifferentiation helps to construct a favorable microenvironment for axon growth. Transcription factors regulate cell reprogramming and thus may be critical for Schwann cell phenotype switch during peripheral nerve regeneration. Here, we show that transcription factor B-cell lymphoma/leukemia 11A (BCL11A) is up-regulated in Schwann cells of injured peripheral nerves. Bcl11a silencing suppresses Schwann cell viability, decreases Schwann cell proliferation and migration rates, and impairs the debris clearance ability of Schwann cells. Reduced Bcl11a in injured peripheral nerves results in restricted axon elongation and myelin wrapping, leading to recovery failure. Mechanistically, we demonstrate that BCL11A may mediate Schwann cell activity through binding to the promoter of nuclear receptor subfamily 2 group F member 2 (Nr2f2) and regulating Nr2f2 expression. Collectively, we conclude that BCL11A is essential for Schwann cell activation and peripheral nerve regeneration, providing a potential therapeutic target for the treatment of peripheral nerve injury.

Deferoxamine Promotes Peripheral Nerve Regeneration by Enhancing Schwann Cell Function and Promoting Axon Regeneration of Dorsal Root Ganglion

Deferoxamine (DFO) is a potent iron chelator for clinical treatment of various diseases. Recent studies have also shown its potential to promote vascular regeneration during peripheral nerve regeneration. However, the effect of DFO on the Schwann cell function and axon regeneration remains unclear. In this study, we investigated the effects of different concentrations of DFO on Schwann cell viability, proliferation, migration, expression of key functional genes, and axon regeneration of dorsal root ganglia (DRG) through a series of in vitro experiments. We found that DFO improves Schwann cell viability, proliferation, and migration in the early stages, with an optimal concentration of 25 μM. DFO also upregulates the expression of myelin-related genes and nerve growth-promoting factors in Schwann cells, while inhibiting the expression of Schwann cell dedifferentiation genes. Moreover, the appropriate concentration of DFO promotes axon regeneration in DRG. Our findings demonstrate that DFO, with suitable concentration and duration of action, can positively affect multiple stages of peripheral nerve regeneration, thereby improving the effectiveness of nerve injury repair. This study also enriches the theory of DFO promoting peripheral nerve regeneration and provides a basis for the design of sustained-release DFO nerve grafts.

Schwann cell-specific RhoA knockout accelerates peripheral nerve regeneration via promoting Schwann cell dedifferentiation.

Our previous studies indicated that RhoA knockdown or inhibition could alleviate the proliferation, migration, and differentiation of Schwann cells. However, the role of RhoA in Schwann cells during nerve injury and repair is still unknown. Herein, we developed two lines of Schwann cells conditional RhoA knockout (cKO) mice by breeding RhoAflox / flox mice with PlpCre -ERT2 or DhhCre mice. Our results indicate that RhoA cKO in Schwann cells accelerates axonal regrowth and remyelination after sciatic nerve injury, which enhances the recovery of nerve conduction and hindlimb gait, and alleviates the amyotrophy in gastrocnemius muscle. Mechanistic studies in both in vivo and in vitro models revealed that RhoA cKO could facilitate Schwann cell dedifferentiation via JNK pathway. Schwann cell dedifferentiation subsequently promotes Wallerian degeneration by enhancing phagocytosis and myelinophagy, as well as stimulating the production of neurotrophins (NT-3, NGF, BDNF, and GDNF). These findings shed light on the role of RhoA in Schwann cells during nerve injury and repair, indicating that cell type-specific RhoA targeting could serve as a promising molecular therapeutic strategy for peripheral nerve injury.

Regulation of the V-ATPase subunit ATP6V0D2 and its role in demyelination after peripheral nerve injury

After peripheral nerve injury, demyelinating Schwann cells discharge myelin debris and macrophages execute myelin degradation, leading to demyelination of degenerating axons, which is essential for efficient nerve regeneration. In this study, we show that vacuolar-type proton ATPase subunit d2 (Atp6v0d2) is among the most highly upregulated genes in degenerating mouse sciatic nerves after nerve injury using microarray analysis. ATP6V0D2 is mostly expressed in macrophages of injured nerves. Atp6v0d2 knockout mice display delayed peripheral nerve demyelination and significantly attenuated myelin lipid digestion after nerve injury. However, macrophage recruitment and Schwann cell dedifferentiation are unaffected by loss of Atp6v0d2 expression. Taken together, these data demonstrate that ATP6V0D2 in macrophages is specifically required for demyelination during Wallerian degeneration.

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.

Transection injury differentially alters the proteome of the human sural nerve.

Regeneration after severe peripheral nerve injury is often poor. Knowledge of human nerve regeneration and the growth microenvironment is greatly lacking. We aimed to identify the regenerative proteins in human peripheral nerve by comparing the proteome before and after a transection injury. In a unique study design, we collected closely matched samples of naïve and injured sural nerve. Naïve and injured (two weeks after injury) samples were analyzed using mass spectrometry and immunoassays. We found significantly altered levels following the nerve injury. Mass spectrometry revealed that injury samples had 568 proteins significantly upregulated and 471 significantly downregulated compared to naïve samples (q-value ≤ 0.05 and Z ≥ |2| (log2)). We used Gene Ontology (GO) pathway overrepresentation analysis to highlight groups of proteins that were significantly upregulated or downregulated with injury-induced degeneration and regeneration. Significant protein changes in key pathways were identified including growth factor levels, Schwann cell de-differentiation, myelination downregulation, epithelial-mesenchymal transition (EMT), and axonal regeneration pathways. The proteomes of the uninjured nerve compared to the degenerating/regenerating nerve may reveal biomarkers to aid in the development of repair strategies such as infusing supplemental trophic factors and in monitoring neural tissue regeneration.

Phenolic glycolipid-1 of Mycobacterium leprae is involved in human Schwann cell line ST8814 neurotoxic phenotype.

Leprosy is a chronic infectious disease caused by Mycobacterium leprae infection in Schwann cells. Axonopathy is considered a hallmark of leprosy neuropathy and is associated with the irreversible motor and sensory loss seen in infected patients. Although M. leprae is recognized to provoke Schwann cell dedifferentiation, the mechanisms involved in the contribution of this phenomenon to neural damage remain unclear. In the present work, we used live M. leprae to infect the immortalized human Schwann cell line ST8814. The neurotoxicity of infected Schwann cell-conditioned medium (SCCM) was then evaluated in a human neuroblastoma cell lineage and mouse neurons. ST8814 Schwann cells exposed to M. leprae affected neuronal viability by deviating glial 14 C-labeled lactate, important fuel of neuronal central metabolism, to de novo lipid synthesis. The phenolic glycolipid-1 (PGL-1) is a specific M. leprae cell wall antigen proposed to mediate bacterial-Schwann cell interaction. Therefore, we assessed the role of the PGL-1 on Schwann cell phenotype by using transgenic M. bovis (BCG)-expressing the M. leprae PGL-1. We observed that BCG-PGL-1 was able to induce a phenotype similar to M. leprae, unlike the wild-type BCG strain. We next demonstrated that this Schwann cell neurotoxic phenotype, induced by M. leprae PGL-1, occurs through the protein kinase B (Akt) pathway. Interestingly, the pharmacological inhibition of Akt by triciribine significantly reduced free fatty acid content in the SCCM from M. leprae- and BCG-PGL-1-infected Schwann cells and, hence, preventing neuronal death. Overall, these findings provide novel evidence that both M. leprae and PGL-1, induce a toxic Schwann cell phenotype, by modifying the host lipid metabolism, resulting in profound implications for neuronal loss. We consider this metabolic rewiring a new molecular mechanism to be the basis of leprosy neuropathy.

DDDR-06. NEUROFIBROMATOSIS TUMOR SUPPRESSORS COOPERATIVELY DRIVE TUMOR DE-DIFFERENTIATION AND MEK INHIBITOR RESISTANCE IN PERIPHERAL NERVOUS SYSTEM TUMORS

Abstract Schwann cell derived tumors comprising schwannomas, neurofibromas, and malignant peripheral nerve sheath tumors (MPNSTs) are the most common cancers of the peripheral nervous system and often arise in patients with neurofibromatosis type-1 (NF-1) or type-2 (NF-2). NF-1 is caused by loss of NF1, a negative regulator of Ras signaling, and NF-2 is caused by loss of NF2, a pleiotropic tumor suppressor with numerous functions including inhibition of PAK signaling. However, whether functional interactions exist between the NF1 and NF2 tumor suppressors remain unclear. More broadly, there are currently no effective molecular therapies for patients with Schwann cell tumors beyond the MEK inhibitor selumetinib to treat neurofibromas in patients with NF-1. Here, we integrate DNA methylation profiling, whole exome sequencing, bulk and single-cell RNA sequencing, biochemistry, and pharmacology across human samples, cell lines, and mouse xenografts to identify cellular de-differentiation as a driver of malignant transformation and selumetinib resistance. Single nuclear RNA-sequencing of human neurofibromas (n = 3) or MPNSTs (n = 3) revealed a total of 13 cell types with increased proliferating, de-differentiated tumor cell populations in MPNST samples. Single cell RNA-sequencing of MPNST mouse xenografts revealed persistence of de-differentiated cell populations in selumetinib treated samples compared to vehicle control, suggesting cellular de-differentiation underlies treatment resistance. A genome-wide CRISPRi screen for mediators of selumetinib response in NF1 deficient neurofibroma cells revealed NF2 loss drives selumetinib resistance. Consistently, NF2 suppression in NF1 deficient neurofibroma cells caused Schwann cell de-differentiation and activation of PAK, a serine threonine kinase. Translationally, a small molecule PAK inhibitor in combination with selumetinib formed an effective therapy in mouse MPNST xenografts. In sum, we elucidate a paradigm of de-differentiation driving malignant transformation and treatment resistance in Schwann cell tumors, uncovering a functional link between the NF1 and NF2 tumor suppressors that sheds light on a novel therapeutic vulnerability.

Investigation of the Hydrogen Sulfide Signaling Pathway in Schwann Cells during Peripheral Nerve Degeneration: Multi-Omics Approaches.

N-ethylmaleimide (NEM) inhibits peripheral nerve degeneration (PND) by targeting Schwann cells in a hydrogen sulfide (H2S)-pathway-dependent manner, but the underlying molecular and pharmacological mechanisms are unclear. We investigated the effect of NEM, an α,β-unsaturated carboxyl compound, on H2S signaling in in vitro- and ex vivo-dedifferentiated Schwann cells using global proteomics (LC-MS) and transcriptomics (whole-genome and small RNA-sequencing (RNA-seq)) methods. The multi-omics analyses identified several genes and proteins related to oxidative stress, such as Sod1, Gnao1, Stx4, Hmox2, Srxn1, and Edn1. The responses to oxidative stress were transcriptionally regulated by several transcription factors, such as Atf3, Fos, Rela, and Smad2. In a functional enrichment analysis, cell cycle, oxidative stress, and lipid/cholesterol metabolism were enriched, implicating H2S signaling in Schwann cell dedifferentiation, proliferation, and myelination. NEM-induced changes in the H2S signaling pathway affect oxidative stress, lipid metabolism, and the cell cycle in Schwann cells. Therefore, regulation of the H2S signaling pathway by NEM during PND could prevent Schwann cell demyelination, dedifferentiation, and proliferation.

Traumatic and Diabetic Schwann Cell Demyelination Is Triggered by a Transient Mitochondrial Calcium Release through Voltage Dependent Anion Channel 1.

A large number of peripheral neuropathies, among which are traumatic and diabetic peripheral neuropathies, result from the degeneration of the myelin sheath, a process called demyelination. Demyelination does not result from Schwann cell death but from Schwann cell dedifferentiation, which includes reprograming and several catabolic and anabolic events. Starting around 4 h after nerve injury, activation of MAPK/cJun pathways is the earliest characterized step of this dedifferentiation program. Here we show, using real-time in vivo imaging, that Schwann cell mitochondrial pH, motility and calcium content are altered as soon as one hour after nerve injury. Mitochondrial calcium release occurred through the VDAC outer membrane channel and mPTP inner membrane channel. This calcium influx in the cytoplasm induced Schwann-cell demyelination via MAPK/c-Jun activation. Blocking calcium release through VDAC silencing or VDAC inhibitor TRO19622 prevented demyelination. We found that the kinetics of mitochondrial calcium release upon nerve injury were altered in the Schwann cells of diabetic mice suggesting a permanent leak of mitochondrial calcium in the cytoplasm. TRO19622 treatment alleviated peripheral nerve defects and motor deficit in diabetic mice. Together, these data indicate that mitochondrial calcium homeostasis is instrumental in the Schwann cell demyelination program and that blocking VDAC constitutes a molecular basis for developing anti-demyelinating drugs for diabetic peripheral neuropathy.

Perioperative Suppression of Schwann Cell Dedifferentiation Reduces the Risk of Adenomyosis Resulting from Endometrial-Myometrial Interface Disruption in Mice.

We have recently demonstrated that endometrial–myometrial interface (EMI) disruption (EMID) can cause adenomyosis in mice, providing experimental evidence for the well-documented epidemiological finding that iatrogenic uterine procedures increase the risk of adenomyosis. To further elucidate its underlying mechanisms, we designed this study to test the hypothesis that Schwann cells (SCs) dedifferentiating after EMID facilitate the genesis of adenomyosis, but the suppression of SC dedifferentiation perioperatively reduces the risk. We treated mice perioperatively with either mitogen-activated protein kinase kinase (MEK)/extracellular-signal regulated protein kinase (ERK) or c-Jun N-terminal kinase (JNK) inhibitors or a vehicle 4 h before and 24 h, 48 h and 72 h after the EMID procedure. We found that EMID resulted in progressive SCs dedifferentiation, concomitant with an increased abundance of epithelial cells in the myometrium and a subsequent epithelial–mesenchymal transition (EMT). This EMID-induced change was abrogated significantly with perioperative administration of JNK or MEK/ERK inhibitors. Consistently, perioperative administration of a JNK or a MEK/ERK inhibitor reduced the incidence by nearly 33.5% and 14.3%, respectively, in conjunction with reduced myometrial infiltration of adenomyosis and alleviation of adenomyosis-associated hyperalgesia. Both treatments significantly decelerated the establishment of adenomyosis and progression of EMT, fibroblast-to-myofibroblast trans-differentiation and fibrogenesis in adenomyotic lesions. Thus, we provide the first piece of evidence strongly implicating the involvement of SCs in the pathogenesis of adenomyosis induced by EMID.

4‐Aminopyridine promotes cutaneous reinnervation and accelerates wound healing

Skin wound healing is a tightly orchestrated process between epithelial, dermal, neuronal and vascular cells, all working in concert to restore tissue integrity and skin barrier function. Here, we report the therapeutic regenerative effects of FDA‐approved 4‐aminopyridine (4‐AP) to promote skin wound healing by accelerating re‐epithelialization, wound induced hair neogenesis (WIHN), dermal collagen deposition, angiogenesis, and reinnervation. We demonstrate that 4‐AP enhanced wound closure by promoting keratinocyte proliferation and migration as well as collagen deposition through myofibroblast differentiation. 4‐AP treatment increased both epidermal thickness, as well as the number of hair follicles and blood vessels in healed wounds, suggesting better restoration of skin function and architecture compared to simple wound scar formation. Moreover, 4‐AP enhanced reinnervation of healed wounds by promoting Schwann cell (SC) de‐differentiation and secretion of neuromediators (NGF, SP) associated with regeneration. In‐vitro studies using human cells demonstrated that 4‐AP enhanced proliferation and migration of keratinocytes, and SCs and that 4‐AP enhances cellular interactions between neuronal and non‐neuronal cells to further accelerate wound healing. 4‐AP induced secretion of NGF from both SCs and keratinocytes. 4‐AP enhances many of the key attributes of successful wound healing and is a promising therapeutic adjuvant for skin wound healing and tissue regeneration.

Castration mediated schwann cell dedifferentiation leads to slower nerve conduction, decreased neuritogenesis, and nitrergic neuron loss

ABSTRACT Introduction/Objectives Low levels of androgens due to androgen deprivation therapy for prostate cancer treatment or due to aging lead to erectile dysfunction (ED). Castration, a preclinical model of low androgens, induces ED due to damage to pelvic nerves and a loss of erectogenic nitrergic nerves. Schwann cells (SC) release neurotrophic factors to promote pelvic nerve growth and survival. It is unknown how testosterone deficiency impacts SC function. This study determines how testosterone deficiency influences SC phenotype and trophic support in neuronal regeneration. We hypothesize testosterone deficiency will decrease myelinated pelvic SC populations, delay penile nerve conduction and reduce pelvic nitrergic neurons. Methods Male Sprague Dawley rats (12 wks) were divided into control (CON) and surgical castration (CAST) groups. All experiments were performed six weeks following castration. Dorsal penile nerves (DPN; 15-16 mm) were excised, placed in a sylgard dish containing oxygenated buffer and conduction velocity of myelinated fast fibers was assessed (n=8 per group). The major pelvic ganglia (MPG) were collected and dissociated into SC or neurons. Dissociated neurons were grown on laminin coated coverslips. To determine whether SCs could provide trophic support, we co-cultured neurons on top of confluent SCs. We assessed the following co-culture groups: CON SC + CON MPG, CON SC + CAST MPG, CAST SC + CON MPG (n=4 per group, cultured in triplicate). After 72 hours, all cultures were fixed and stained with beta tubulin class III, neuronal nitric oxide synthase (nNOS), and tyrosine hydroxylase (TH) to assess neurite length and branching, nitrergic and sympathetic neuron populations, respectively. Dissociated SC were characterized via qPCR for markers of all SC (Sox10), myelinated SC (Krox20), and dedifferentiated SC (glial fibrillary acidic protein; GFAP). Results CAST slows DPN myelinated fast fiber conduction velocity. In dissociated cultured MPG neurons, castration reduces both neurite length and number of branches. Interestingly, CAST MPG neurons co-cultured with CON SC restore neurite length and branching to same level as CON MPG neurons co-cultured with CON SC; however, CAST SC do not impact CON MPG neurite length. CAST MPG neurons co-cultured with CON SC and CON MPG neurons co-cultured with CAST SC both have less nitrergic neurons than CON SC + CON MPG neurons. CAST does not impact the number of TH neurons in single or co-cultured conditions. CAST reduces gene expression of Sox10, myelinated SC, and dedifferentiated SC. Conclusions CAST markedly impairs penile nerve conduction, reduces neurite outgrowth and the proportion of pro-erectile nitrergic neurons. Healthy SC are able to rescue neuritogenesis but not the survival of nitrergic neurons. Additionally, healthy MPG neurons plated on CAST SC have a reduced number of nitrergic neurons. We believe CAST SC are undergoing a phenotypic switch from a myelinated neurotrophic form into dedifferentiated SC promoting neuritogenesis and loss of nitrergic neurons. Preserving the myelinated SC phenotype may prevent impaired nerve damage and subsequent ED in patients with testosterone deficiency. Disclosure Work supported by industry: no.