ABSTRACT
Myelination of axons facilitates the rapid propagation of electrical signals and the long-term integrity of axons. The ubiquitin-proteasome system is essential for proper protein homeostasis, which is particularly crucial for interactions of postmitotic cells. In our study, we examined how the E3 ubiquitin ligase FBXO7-SCF (SKP1, Cul1, F-box protein) expressed in myelinating cells affects the axon-myelin unit. Deletion of Fbxo7 in oligodendrocytes and Schwann cells in mice using the Cnp1-Cre driver line led to motor impairment due to hindlimb paresis. It did not result in apoptosis of myelinating cells, nor did it affect the proper myelination of axons or lead to demyelination. It however triggered axonal degeneration in the CNS and resulted in the severe degeneration of axons in the PNS, inducing a full-blown neuropathy. Both the CNS and PNS displayed inflammation, while the PNS was also characterized by fibrosis, massive infiltration of macrophages, and edema. Tamoxifen-induced deletion of Fbxo7, after myelination using the Plp1-CreERT2 line, led to a small number of degenerated axons and hence a very mild peripheral neuropathy. Interestingly, loss of Fbxo7 also resulted in reduced proteasome activity in Schwann cells but not in cerebellar granule neurons, indicating a specific sensitivity of the former cell type. Together, our results demonstrate an essential role for FBXO7 in myelinating cells to support associated axons, which is fundamental to the proper developmental establishment and the long-term integrity of the axon-myelin unit.SIGNIFICANCE STATEMENT The myelination of axons facilitates the fast propagation of electrical signals and the trophic support of the myelin-axon unit. Here, we report that deletion of Fbxo7 in myelinating cells in mice triggered motor impairment but had no effect on myelin biogenesis. Loss of Fbxo7 in myelinating glia, however, led to axonal degeneration in the CNS and peripheral neuropathy of the axonal type. In addition, we found that Schwann cells were particularly sensitive to Fbxo7 deficiency reflected by reduced proteasome activity. Based on these findings, we conclude that Fbxo7 is essential for the support of the axon-myelin unit and long-term axonal health.
Subject(s)
Axons/metabolism , F-Box Proteins/genetics , Myelin Sheath/metabolism , Peripheral Nervous System Diseases/metabolism , Animals , Apoptosis , Axons/pathology , Cells, Cultured , Central Nervous System/metabolism , Central Nervous System/pathology , F-Box Proteins/metabolism , Female , Gene Deletion , Male , Mice , Mice, Inbred C57BL , Myelin Sheath/pathology , Peripheral Nervous System Diseases/genetics , Peripheral Nervous System Diseases/pathology , Proteasome Endopeptidase Complex/metabolismABSTRACT
Charcot-Marie-Tooth disease 1 A (CMT1A) is caused by an intrachromosomal duplication of the gene encoding for PMP22 leading to peripheral nerve dysmyelination, axonal loss, and progressive muscle weakness. No therapy is available. PXT3003 is a low-dose combination of baclofen, naltrexone, and sorbitol which has been shown to improve disease symptoms in Pmp22 transgenic rats, a bona fide model of CMT1A disease. However, the superiority of PXT3003 over its single components or dual combinations have not been tested. Here, we show that in a dorsal root ganglion (DRG) co-culture system derived from transgenic rats, PXT3003 induced myelination when compared to its single and dual components. Applying a clinically relevant ("translational") study design in adult male CMT1A rats for 3 months, PXT3003, but not its dual components, resulted in improved performance in behavioral motor and sensory endpoints when compared to placebo. Unexpectedly, we observed only a marginally increased number of myelinated axons in nerves from PXT3003-treated CMT1A rats. However, in electrophysiology, motor latencies correlated with increased grip strength indicating a possible effect of PXT3003 on neuromuscular junctions (NMJs) and muscle fiber pathology. Indeed, PXT3003-treated CMT1A rats displayed an increased perimeter of individual NMJs and a larger number of functional NMJs. Moreover, muscles of PXT3003 CMT1A rats displayed less neurogenic atrophy and a shift toward fast contracting muscle fibers. We suggest that ameliorated motor function in PXT3003-treated CMT1A rats result from restored NMJ function and muscle innervation, independent from myelination.
Subject(s)
Baclofen/administration & dosage , Charcot-Marie-Tooth Disease/drug therapy , Demyelinating Diseases/drug therapy , Naltrexone/administration & dosage , Neuromuscular Junction/drug effects , Sorbitol/administration & dosage , Animals , Charcot-Marie-Tooth Disease/genetics , Charcot-Marie-Tooth Disease/physiopathology , Coculture Techniques , Demyelinating Diseases/genetics , Demyelinating Diseases/physiopathology , Drug Synergism , Drug Therapy, Combination , Female , Male , Myelin Proteins/genetics , Neural Conduction/drug effects , Neural Conduction/physiology , Neuromuscular Junction/physiology , Rats , Rats, Sprague-Dawley , Rats, TransgenicABSTRACT
Oligodendrocytes, the myelin-forming glial cells of the central nervous system, maintain long-term axonal integrity. However, the underlying support mechanisms are not understood. Here we identify a metabolic component of axon-glia interactions by generating conditional Cox10 (protoheme IX farnesyltransferase) mutant mice, in which oligodendrocytes and Schwann cells fail to assemble stable mitochondrial cytochrome c oxidase (COX, also known as mitochondrial complex IV). In the peripheral nervous system, Cox10 conditional mutants exhibit severe neuropathy with dysmyelination, abnormal Remak bundles, muscle atrophy and paralysis. Notably, perturbing mitochondrial respiration did not cause glial cell death. In the adult central nervous system, we found no signs of demyelination, axonal degeneration or secondary inflammation. Unlike cultured oligodendrocytes, which are sensitive to COX inhibitors, post-myelination oligodendrocytes survive well in the absence of COX activity. More importantly, by in vivo magnetic resonance spectroscopy, brain lactate concentrations in mutants were increased compared with controls, but were detectable only in mice exposed to volatile anaesthetics. This indicates that aerobic glycolysis products derived from oligodendrocytes are rapidly metabolized within white matter tracts. Because myelinated axons can use lactate when energy-deprived, our findings suggest a model in which axon-glia metabolic coupling serves a physiological function.
Subject(s)
Axons/physiology , Glycolysis , Myelin Sheath/metabolism , Oligodendroglia/metabolism , Action Potentials , Alkyl and Aryl Transferases/deficiency , Alkyl and Aryl Transferases/genetics , Alkyl and Aryl Transferases/metabolism , Animals , Brain/cytology , Brain/metabolism , Cell Respiration , Cell Survival , Demyelinating Diseases/enzymology , Demyelinating Diseases/genetics , Demyelinating Diseases/metabolism , Demyelinating Diseases/pathology , Electron Transport Complex IV/antagonists & inhibitors , Electron Transport Complex IV/genetics , Electron Transport Complex IV/metabolism , Lactic Acid/metabolism , Magnetic Resonance Spectroscopy , Membrane Proteins/deficiency , Membrane Proteins/genetics , Membrane Proteins/metabolism , Mice , Mitochondria/enzymology , Mitochondria/genetics , Mitochondria/metabolism , Mitochondria/pathology , Mutant Proteins/genetics , Mutant Proteins/metabolism , Oligodendroglia/cytology , Oligodendroglia/drug effects , Oligodendroglia/enzymology , Protons , Schwann Cells/enzymology , Schwann Cells/metabolism , Time FactorsABSTRACT
Pelizaeus-Merzbacher disease (PMD) is a severe hypomyelinating disease, characterized by ataxia, intellectual disability, epilepsy, and premature death. In the majority of cases, PMD is caused by duplication of PLP1 that is expressed in myelinating oligodendrocytes. Despite detailed knowledge of PLP1, there is presently no curative therapy for PMD. We used a Plp1 transgenic PMD mouse model to test the therapeutic effect of Lonaprisan, an antagonist of the nuclear progesterone receptor, in lowering Plp1 mRNA overexpression. We applied placebo-controlled Lonaprisan therapy to PMD mice for 10 weeks and performed the grid slip analysis to assess the clinical phenotype. Additionally, mRNA expression and protein accumulation as well as histological analysis of the central nervous system were performed. Although Plp1 mRNA levels are increased 1.8-fold in PMD mice compared to wild-type controls, daily Lonaprisan treatment reduced overexpression at the RNA level to about 1.5-fold, which was sufficient to significantly improve the poor motor phenotype. Electron microscopy confirmed a 25% increase in the number of myelinated axons in the corticospinal tract when compared to untreated PMD mice. Microarray analysis revealed the upregulation of proapoptotic genes in PMD mice that could be partially rescued by Lonaprisan treatment, which also reduced microgliosis, astrogliosis, and lymphocyte infiltration.
Subject(s)
Estrenes/therapeutic use , Hormone Antagonists/therapeutic use , Pelizaeus-Merzbacher Disease/drug therapy , Progesterone/antagonists & inhibitors , Animals , Disease Models, Animal , Estrenes/pharmacokinetics , Estrenes/pharmacology , Gene Expression Regulation/drug effects , Hormone Antagonists/pharmacokinetics , Hormone Antagonists/pharmacology , Male , Mice , Mice, Inbred C57BL , Mice, Transgenic , Myelin Proteolipid Protein/genetics , Phenotype , RNA, Messenger/geneticsABSTRACT
BACKGROUND: Charcot-Marie-Tooth disease type 1A (CMT1A) is the most common inherited neuropathy, a debilitating disease without known cure. Among patients with CMT1A, disease manifestation, progression and severity are strikingly variable, which poses major challenges for the development of new therapies. Hence, there is a strong need for sensitive outcome measures such as disease and progression biomarkers, which would add powerful tools to monitor therapeutic effects in CMT1A. METHODS: We established a pan-European and American consortium comprising nine clinical centres including 311 patients with CMT1A in total. From all patients, the CMT neuropathy score and secondary outcome measures were obtained and a skin biopsy collected. In order to assess and validate disease severity and progression biomarkers, we performed qPCR on a set of 16 animal model-derived potential biomarkers in skin biopsy mRNA extracts. RESULTS: In 266 patients with CMT1A, a cluster of eight cutaneous transcripts differentiates disease severity with a sensitivity and specificity of 90% and 76.1%, respectively. In an additional cohort of 45 patients with CMT1A, from whom a second skin biopsy was taken after 2-3 years, the cutaneous mRNA expression of GSTT2, CTSA, PPARG, CDA, ENPP1 and NRG1-Iis changing over time and correlates with disease progression. CONCLUSIONS: In summary, we provide evidence that cutaneous transcripts in patients with CMT1A serve as disease severity and progression biomarkers and, if implemented into clinical trials, they could markedly accelerate the development of a therapy for CMT1A.
Subject(s)
Charcot-Marie-Tooth Disease/therapy , Disease Progression , Genetic Markers/genetics , Skin/pathology , Treatment Outcome , Adult , Aged , Biopsy , Cathepsin A/genetics , Charcot-Marie-Tooth Disease/blood , Charcot-Marie-Tooth Disease/genetics , Female , Glutathione Transferase/genetics , Glycoproteins/genetics , Humans , Male , Middle Aged , Neuregulin-1/genetics , Nuclear Proteins , PPAR gamma/genetics , Phosphoric Diester Hydrolases/genetics , Prognosis , Pyrophosphatases/genetics , RNA, Messenger/genetics , Real-Time Polymerase Chain Reaction , Transcription, Genetic/geneticsABSTRACT
Protein zero (P0) is the major structural component of peripheral myelin. Lack of this adhesion protein from Schwann cells causes a severe dysmyelinating neuropathy with secondary axonal degeneration in humans with the neuropathy Dejerine-Sottas syndrome (DSS) and in the corresponding mouse model (P0(null)-mice). In the mammalian CNS, the tetraspan-membrane protein PLP is the major structural myelin constituent and required for the long-term preservation of myelinated axons, which fails in hereditary spastic paraplegia (SPG type-2) and the relevant mouse model (Plp(null)-mice). The Plp-gene is also expressed in Schwann cells but PLP is of very low abundance in normal peripheral myelin; its function has thus remained enigmatic. Here we show that the abundance of PLP but not of other tetraspan myelin proteins is strongly increased in compact peripheral myelin of P0(null)-mice. To determine the functional relevance of PLP expression in the absence of P0, we generated P0(null)*Plp(null)-double-mutant mice. Compared with either single-mutant, P0(null)*Plp(null)-mice display impaired nerve conduction, reduced motor functions, and premature death. At the morphological level, axonal segments were frequently non-myelinated but in a one-to-one relationship with a hypertrophic Schwann cell. Importantly, axonal numbers were reduced in the vital phrenic nerve of P0(null)*Plp(null)-mice. In the absence of P0, thus, PLP also contributes to myelination by Schwann cells and to the preservation of peripheral axons. These data provide a link between the Schwann cell-dependent support of peripheral axons and the oligodendrocyte-dependent support of central axons.
Subject(s)
Axons/metabolism , Myelin P0 Protein/metabolism , Myelin Proteolipid Protein/metabolism , Phrenic Nerve/metabolism , Sciatic Nerve/metabolism , Animals , Axons/pathology , Brain/metabolism , Brain/pathology , Female , Kaplan-Meier Estimate , Mice, Inbred C57BL , Mice, Knockout , Mortality, Premature , Motor Activity/physiology , Myelin P0 Protein/genetics , Myelin Proteolipid Protein/genetics , Myelin Sheath/metabolism , Myelin Sheath/pathology , Myelin-Associated Glycoprotein/metabolism , Neural Conduction/physiology , Optic Nerve/metabolism , Optic Nerve/pathology , Phrenic Nerve/pathology , Sciatic Nerve/pathologyABSTRACT
Charcot-Marie-Tooth 1A (CMT1A) is a demyelinating hereditary neuropathy for which pharmacological treatments are not yet available. An abnormally high intracellular Ca(2+) concentration was observed in Schwann cells (SC) from CMT1A rats, caused by the PMP22-mediated overexpression of the P2X7 purinoceptor. The purpose of this study was to investigate the tolerability and therapeutic potential of a pharmacological antagonist of the P2X7 receptor (A438079) in CMT1A. A438079 ameliorated in vitro myelination of organotypic DRG cultures from CMT1A rats. Furthermore, we performed an experimental therapeutic trial in PMP22 transgenic and in wild-type rats. A preliminary dose-escalation trial showed that 3mg/kg A438079 administered via intraperitoneal injection every 24h for four weeks was well tolerated by wild type and CMT1A rats. Affected rats treated with 3mg/kg A438079 revealed a significant improvement of the muscle strength, when compared to placebo controls. Importantly, histologic analysis revealed a significant increase of the total number of myelinated axons in tibial nerves. Moreover, a significant decrease of the hypermyelination of small caliber axons and a significant increase of the frequency and diameter of large caliber myelinated axons was highlighted. An improved distal motor latencies was recorded, whereas compound muscle action potentials (CMAP) remained unaltered. A438079 reduced the SC differentiation defect in CMT1A rats. These results show that pharmacological inhibition of the P2X7 receptor is well tolerated in CMT1A rats and represents a proof-of-principle that antagonizing this pathway may correct the molecular derangements and improve the clinical phenotype in the CMT1A neuropathy.
Subject(s)
Axons/pathology , Charcot-Marie-Tooth Disease/pathology , Demyelinating Diseases/pathology , Myelin Proteins/metabolism , Receptors, Purinergic P2X7/metabolism , Schwann Cells/metabolism , Animals , Animals, Genetically Modified , Charcot-Marie-Tooth Disease/physiopathology , Demyelinating Diseases/genetics , Disease Models, Animal , Myelin Proteins/genetics , Phenotype , Rats, Sprague-Dawley , Rats, TransgenicABSTRACT
PURPOSE: Charcot-Marie-Tooth (CMT) disease is a heterogeneous group of genetic disorders of the peripheral nervous system. Copy-number variants (CNVs) contribute significantly to CMT, as duplication of PMP22 underlies the majority of CMT1 cases. We hypothesized that CNVs and/or single-nucleotide variants (SNVs) might exist in patients with CMT with an unknown molecular genetic etiology. METHODS: Two hundred patients with CMT, negative for both SNV mutations in several CMT genes and for CNVs involving PMP22, were screened for CNVs by high-resolution oligonucleotide array comparative genomic hybridization. Whole-exome sequencing was conducted on individuals with rare, potentially pathogenic CNVs. RESULTS: Putatively causative CNVs were identified in five subjects (~2.5%); four of the five map to known neuropathy genes. Breakpoint sequencing revealed Alu-Alu-mediated junctions as a predominant contributor. Exome sequencing identified MFN2 SNVs in two of the individuals. CONCLUSION: Neuropathy-associated CNV outside of the PMP22 locus is rare in CMT. Nevertheless, there is potential clinical utility in testing for CNVs and exome sequencing in CMT cases negative for the CMT1A duplication. These findings suggest that complex phenotypes including neuropathy can potentially be caused by a combination of SNVs and CNVs affecting more than one disease-associated locus and contributing to a mutational burden.Genet Med 18 5, 443-451.
Subject(s)
Charcot-Marie-Tooth Disease/genetics , GTP Phosphohydrolases/genetics , Mitochondrial Proteins/genetics , Myelin Proteins/genetics , Polyneuropathies/genetics , Adult , Age of Onset , Charcot-Marie-Tooth Disease/physiopathology , Child, Preschool , Comparative Genomic Hybridization , DNA Copy Number Variations/genetics , Exome/genetics , Female , Genetic Predisposition to Disease , High-Throughput Nucleotide Sequencing/methods , Humans , Male , Motor Neurons/metabolism , Motor Neurons/pathology , Myelin P0 Protein/genetics , Neural Conduction/genetics , Polymorphism, Single Nucleotide/genetics , Polyneuropathies/physiopathologyABSTRACT
The transient receptor potential A1 (TRPA1) channel is essential for vertebrate pain. Even though TRPA1 activation by ligands has been studied extensively, the molecular machinery regulating TRPA1 is only poorly understood. Using an unbiased proteomics-based approach we uncovered the physical association of Annexin A2 (AnxA2) with native TRPA1 in mouse sensory neurons. AnxA2 is enriched in a subpopulation of sensory neurons and coexpressed with TRPA1. Furthermore, we observe an increase of TRPA1 membrane levels in cultured sensory neurons from AnxA2-deficient mice. This is reflected by our calcium imaging experiments revealing higher responsiveness upon TRPA1 activation in AnxA2-deficient neurons. In vivo these findings are associated with enhanced nocifensive behaviors specifically in TRPA1-dependent paradigms of acute and inflammatory pain, while heat and mechanical sensitivity as well as TRPV1-mediated pain are preserved in AnxA2-deficient mice. Our results support a model whereby AnxA2 limits the availability of TRPA1 channels to regulate nociceptive signaling in vertebrates.
Subject(s)
Annexin A2/metabolism , Calcium Channels/metabolism , Nerve Tissue Proteins/metabolism , Nociception/physiology , Nociceptors/metabolism , Pain/metabolism , Transient Receptor Potential Channels/metabolism , Animals , Behavior, Animal/physiology , HEK293 Cells , Hot Temperature , Humans , Mice , Pain Measurement , Physical Stimulation , Rats , TRPA1 Cation ChannelABSTRACT
Glycogen synthase kinase 3ß (GSK3ß) inhibitors, especially the mood stabilizer lithium chloride, are also used as neuroprotective or anti-inflammatory agents. We studied the influence of LiCl on the remyelination of peripheral nerves. We showed that the treatment of adult mice with LiCl after facial nerve crush injury stimulated the expression of myelin genes, restored the myelin structure, and accelerated the recovery of whisker movements. LiCl treatment also promoted remyelination of the sciatic nerve after crush. We also demonstrated that peripheral myelin gene MPZ and PMP22 promoter activities, transcripts, and protein levels are stimulated by GSK3ß inhibitors (LiCl and SB216763) in Schwann cells as well as in sciatic and facial nerves. LiCl exerts its action in Schwann cells by increasing the amount of ß-catenin and provoking its nuclear localization. We showed by ChIP experiments that LiCl treatment drives ß-catenin to bind to T-cell factor/lymphoid-enhancer factor response elements identified in myelin genes. Taken together, our findings open perspectives in the treatment of nerve demyelination by administering GSK3ß inhibitors such as lithium.
Subject(s)
Glycogen Synthase Kinase 3/metabolism , Lithium Chloride/pharmacology , Myelin Sheath/chemistry , Peripheral Nerves/metabolism , Animals , Cell Nucleus/metabolism , Glycogen Synthase Kinase 3 beta , Male , Mice , Mice, Inbred C57BL , Myelin P0 Protein/metabolism , Peripheral Nerves/drug effects , Placebos , Proto-Oncogene Proteins c-akt/metabolism , Schwann Cells/metabolism , Sciatic Nerve/injuries , Signal TransductionABSTRACT
Disease progression in amyotrophic lateral sclerosis (ALS) is characterized by degeneration of motoneurons (MN) and their axons, but is also influenced by neighboring cells such as astrocytes and microglial cells. The role of microglia in ALS is complex as it switches from an anti-inflammatory and neuroprotective phenotype in early disease to a proinflammatory and neurotoxic phenotype in later stages. Our previous studies in models of neurodegeneration identified rho kinase (ROCK) as a target, which can be manipulated to beneficially influence disease progression. Here, we examined the neuroprotective potential of the ROCK inhibitor Fasudil to target the central pathogenic features of ALS. Application of Fasudil to kainic acid-lesioned primary MN in vitro resulted in a strong prosurvival effect. In vivo, SOD1(G93A) mice benefited from oral treatment with Fasudil showing prolonged survival and improved motor function. These findings were correlated to an improved survival of motor neurons and a pronounced alteration of astroglial and microglial cell infiltration of the spinal cord under Fasudil treatment. Modeling a proinflammatory microglial phenotype by stimulation with LPS in vitro, Fasudil decreased the release of proinflammatory cytokines and chemokines TNFα, Il6, CCL2, CCL3, and CCL5 while CXCL1 release was only transiently suppressed. In sciatic nerve motor axons, neuromuscular junction remodeling processes were increased. In conclusion, we provide preclinical and neurobiological evidence that inhibition of ROCK by the clinically approved small molecule inhibitor Fasudil may be a novel therapeutic approach in ALS combining both neuroprotection and immunomodulation for the cure of this devastating disease.
Subject(s)
1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine/analogs & derivatives , Amyotrophic Lateral Sclerosis/enzymology , Microglia/drug effects , Protein Kinase Inhibitors/pharmacology , rho-Associated Kinases/antagonists & inhibitors , 1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine/pharmacology , Amyotrophic Lateral Sclerosis/genetics , Amyotrophic Lateral Sclerosis/pathology , Animals , Astrocytes/cytology , Astrocytes/drug effects , Axons/drug effects , Axons/pathology , Cell Survival/drug effects , Cells, Cultured , Disease Models, Animal , Mice , Mice, Inbred C57BL , Mice, Transgenic , Microglia/pathology , Spinal Cord/drug effectsABSTRACT
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.
Subject(s)
Arthrogryposis , Charcot-Marie-Tooth Disease , Hereditary Sensory and Motor Neuropathy , Phosphatidylinositol 3-Kinases , Mice , Animals , Proto-Oncogene Proteins c-akt , Rodentia/metabolism , Charcot-Marie-Tooth Disease/genetics , Charcot-Marie-Tooth Disease/pathology , Myelin Proteins/genetics , Myelin Proteins/metabolism , TOR Serine-Threonine KinasesABSTRACT
Charcot-Marie-Tooth disease is the most common inherited neuropathy and a duplication of the peripheral myelin protein 22 gene causes the most frequent subform Charcot-Marie-Tooth 1A. Patients develop a slowly progressive dysmyelinating and demyelinating peripheral neuropathy and distally pronounced muscle atrophy. The amount of axonal loss determines disease severity. Although patients share an identical monogenetic defect, the disease progression is strikingly variable and the impending disease course can not be predicted in individual patients. Despite promising experimental data, recent therapy trials have failed. Established clinical outcome measures are thought to be too insensitive to detect amelioration within trials. Surrogate biomarkers of disease severity in Charcot-Marie-Tooth 1A are thus urgently needed. Peripheral myelin protein 22 transgenic rats harbouring additional copies of the peripheral myelin protein 22 gene ('Charcot-Marie-Tooth rats'), which were kept on an outbred background mimic disease hallmarks and phenocopy the variable disease severity of patients with Charcot-Marie-Tooth 1A. Hence, we used the Charcot-Marie-Tooth rat to dissect prospective and surrogate markers of disease severity derived from sciatic nerve and skin tissue messenger RNA extracts. Gene set enrichment analysis of sciatic nerve transcriptomes revealed that dysregulation of lipid metabolism associated genes such as peroxisome proliferator-activated receptor gamma constitutes a modifier of present and future disease severity. Importantly, we directly validated disease severity markers from the Charcot-Marie-Tooth rats in 46 patients with Charcot-Marie-Tooth 1A. Our data suggest that the combination of age and cutaneous messenger RNA levels of glutathione S-transferase theta 2 and cathepsin A composes a strong indicator of disease severity in patients with Charcot-Marie-Tooth 1A, as quantified by the Charcot-Marie-Tooth Neuropathy Score. This translational approach, utilizing a transgenic animal model, demonstrates that transcriptional analysis of skin biopsy is suitable to identify biomarkers of Charcot-Marie-Tooth 1A.
Subject(s)
Axons/pathology , Charcot-Marie-Tooth Disease/pathology , Myelin Proteins/genetics , Sciatic Nerve/pathology , Animals , Axons/physiology , Charcot-Marie-Tooth Disease/genetics , Charcot-Marie-Tooth Disease/physiopathology , Disease Models, Animal , Glutathione Transferase/genetics , Myelin P0 Protein/genetics , Neural Conduction/physiology , PPAR gamma/genetics , Pain Measurement , Phenotype , Rats , Rats, Transgenic , Sciatic Nerve/physiopathology , Severity of Illness IndexABSTRACT
The peripheral nervous system harbors a remarkable potential to regenerate after acute nerve trauma. Full functional recovery, however, is rare and critically depends on peripheral nerve Schwann cells that orchestrate breakdown and resynthesis of myelin and, at the same time, support axonal regrowth. How Schwann cells meet the high metabolic demand required for nerve repair remains poorly understood. We here report that nerve injury induces adipocyte to glial signaling and identify the adipokine leptin as an upstream regulator of glial metabolic adaptation in regeneration. Signal integration by leptin receptors in Schwann cells ensures efficient peripheral nerve repair by adjusting injury-specific catabolic processes in regenerating nerves, including myelin autophagy and mitochondrial respiration. Our findings propose a model according to which acute nerve injury triggers a therapeutically targetable intercellular crosstalk that modulates glial metabolism to provide sufficient energy for successful nerve repair.
Subject(s)
Myelin Sheath , Peripheral Nerves , Myelin Sheath/metabolism , Neuroglia , Schwann Cells/metabolism , Nerve Regeneration/physiologyABSTRACT
Ascorbic acid (vitamin C) is necessary for myelination of Schwann cell/neuron cocultures and has shown beneficial effects in the treatment of a Charcot-Marie-Tooth neuropathy 1A (CMT1A) mouse model. Although clinical studies revealed that ascorbic acid treatment had no impact on CMT1A, it is assumed to have an important function in peripheral nerve myelination and possibly in remyelination. However, the transport pathway of ascorbic acid into peripheral nerves and the mechanism of ascorbic acid function in peripheral nerves in vivo remained unclear. In this study, we used sodium-dependent vitamin C transporter 2-heterozygous (SVCT2(+/-)) mice to elucidate the functions of SVCT2 and ascorbic acid in the murine peripheral nervous system. SVCT2 and ascorbic acid levels were reduced in SVCT2(+/-) peripheral nerves. Morphometry of sciatic nerve fibers revealed a decrease in myelin thickness and an increase in G-ratios in SVCT2(+/-) mice. Nerve conduction velocities and sensorimotor performance in functional tests were reduced in SVCT2(+/-) mice. To investigate the mechanism of ascorbic acid function, we studied the expression of collagens in the extracellular matrix of peripheral nerves. Here, we show that expression of various collagen types was reduced in sciatic nerves of SVCT2(+/-) mice. We found that collagen gene transcription was reduced in SVCT2(+/-) mice but hydroxyproline levels were not, indicating that collagen formation was regulated on the transcriptional and not the posttranslational level. These results help to clarify the transport pathway and mechanism of action of ascorbic acid in the peripheral nervous system and may lead to novel therapeutic approaches to peripheral neuropathies by manipulation of SVCT2 function.
Subject(s)
Charcot-Marie-Tooth Disease/genetics , Extracellular Matrix/genetics , Nerve Fibers, Myelinated/pathology , Peripheral Nervous System/pathology , Sodium-Coupled Vitamin C Transporters/deficiency , Animals , Ascorbic Acid/genetics , Ascorbic Acid/metabolism , Cells, Cultured , Charcot-Marie-Tooth Disease/metabolism , Charcot-Marie-Tooth Disease/pathology , Coculture Techniques , Extracellular Matrix/metabolism , Extracellular Matrix/pathology , Female , Male , Mice , Mice, Knockout , Nerve Fibers, Myelinated/metabolism , Peripheral Nervous System/metabolism , Sodium-Coupled Vitamin C Transporters/geneticsABSTRACT
INTRODUCTION OR BACKGROUND: Charcot-Marie-Tooth (CMT) disease represents a broad group of inherited motor and sensory neuropathies which can originate from various genetic aberrations, e.g. mutations, deletions and duplications. SOURCES OF DATA: We performed a literature review on murine animal models of CMT disease with regard to experimental therapeutic approaches. Hereby, we focussed on the demyelinating subforms of CMT (CMT1). PubMed items were CMT, animal model, demyelination and therapy. AREAS OF AGREEMENT: Patients affected by CMT suffer from slowly progressive, distally pronounced muscle atrophy caused by an axonal loss. The disease severity is highly variable and impairments may result in wheelchair boundness. No therapy is available yet. AREAS OF CONTROVERSY: Numerous rodent models for the various CMT subtypes are available today. The selection of the correct animal model for the specific CMT subtype provides an important prerequisite for the successful translation of experimental findings in patients. GROWING POINTS: Despite more than 20 years of remarkable progress in CMT research, the disease is still left untreatable. There is a growing number of experimental therapeutic strategies that may be translated into future clinical trials in patients with CMT. AREAS TIMELY FOR DEVELOPING RESEARCH: The slow disease progression and insensitive outcome measures hamper clinical therapy trials in CMT. Biomarkers may provide powerful tools to monitor therapeutic efficacy. Recently, we have shown that transcriptional profiling can be utilized to assess and predict the disease severity in a transgenic rat model and in affected humans.
Subject(s)
Charcot-Marie-Tooth Disease/therapy , Disease Models, Animal , Animals , Animals, Genetically Modified , Charcot-Marie-Tooth Disease/genetics , Genetic Predisposition to Disease , Mice , Molecular Targeted Therapy/methods , RatsABSTRACT
Hexokinase (HK)-1 mitochondrial-binding mechanisms and consequential physiological relevance remain unclear. Recently, De Jesus et al. studied myeloid cells with HK1 carrying mutated mitochondrial-binding domains (MBDs) and provided evidence that HK1 localization controls glucose metabolic fate. Increases in cytosolic HK1 may also contribute to the inflammation associated with diabetes and aging.
Subject(s)
Hexokinase , Mitochondria , Carbohydrate Metabolism , Glucose/metabolism , Hexokinase/genetics , Hexokinase/metabolism , Humans , Inflammation/metabolism , Mitochondria/metabolismABSTRACT
To maintain homeostasis, the body, including the brain, reprograms its metabolism in response to altered nutrition or disease. However, the consequences of these challenges for the energy metabolism of the different brain cell types remain unknown. Here, we generated a proteome atlas of the major central nervous system (CNS) cell types from young and adult mice, after feeding the therapeutically relevant low-carbohydrate, high-fat ketogenic diet (KD) and during neuroinflammation. Under steady-state conditions, CNS cell types prefer distinct modes of energy metabolism. Unexpectedly, the comparison with KD revealed distinct cell type-specific strategies to manage the altered availability of energy metabolites. Astrocytes and neurons but not oligodendrocytes demonstrated metabolic plasticity. Moreover, inflammatory demyelinating disease changed the neuronal metabolic signature in a similar direction as KD. Together, these findings highlight the importance of the metabolic cross-talk between CNS cells and between the periphery and the brain to manage altered nutrition and neurological disease.
Subject(s)
Brain , Diet, Ketogenic , Animals , Brain/metabolism , Carbohydrates , Ketone Bodies/metabolism , Mice , Proteome/metabolismABSTRACT
Charcot-Marie-Tooth disease (CMT) is the most common inherited neuropathy and a duplication of the peripheral myelin protein of 22 kDa (PMP22) gene causes the most frequent subform CMT1A. Clinical impairments are determined by the amount of axonal loss. Axons of the spontaneous mouse mutant Wallerian degeneration slow (Wlds) show markedly reduced degeneration following various types of injuries. Protection is conferred by a chimeric Wlds gene encoding an N-terminal part of ubiquitination factor Ube4b and full length nicotinamide mononucleotide adenylyl transferase 1 (Nmnat1). Nmnat1 enzyme generates nicotinamide adenine dinucleotide (NAD) from nicotinamide mononucleotide. Here, in a Pmp22 transgenic animal model of Charcot-Marie-Tooth disease type 1A (CMT rat), the Wlds transgene reduced axonal loss and clinical impairments without altering demyelination. Furthermore, nicotinamide - substrate precursor of the Nmnat1 enzyme - transiently delayed posttraumatic axonal degeneration in an in vivo model of acute peripheral nerve injury, but to a lower extent than Wlds. In contrast, 8 weeks of nicotinamide treatment did not influence axonal loss or clinical manifestations in the CMT rat. Therefore, nicotinamide can partially substitute for the protective Wlds effect in acute traumatic, but not in chronic secondary axonal injury. Future studies are needed to develop axon protective therapy in CMT1A which may be combined with therapeutic strategies aimed at downregulation of toxic PMP22 overexpression.
Subject(s)
Axons/pathology , Charcot-Marie-Tooth Disease/genetics , Nerve Tissue Proteins/genetics , Neuroprotective Agents/therapeutic use , Niacinamide/therapeutic use , Sciatic Neuropathy/genetics , Wallerian Degeneration/genetics , Wallerian Degeneration/prevention & control , Animals , Axons/metabolism , Charcot-Marie-Tooth Disease/drug therapy , Charcot-Marie-Tooth Disease/pathology , Disease Models, Animal , Female , Rats , Rats, Sprague-Dawley , Rats, Transgenic , Sciatic Neuropathy/complications , Sciatic Neuropathy/pathology , Wallerian Degeneration/pathologyABSTRACT
Charcot-Marie-Tooth disease (CMT) is the most common inherited neuropathy. The predominant subtype, CMT-1A, accounts for more than 50% of all cases and is associated with an interstitial chromosomal duplication of 17p12 (refs. 2,3). We have generated a model of CMT-1A by introducing extra copies of the responsible disease gene, Pmp22 (encoding the peripheral myelin protein of 22 kDa), into transgenic rats. Here, we used this model to test whether progesterone, a regulator of the myelin genes Pmp22 and myelin protein zero (Mpz) in cultured Schwann cells, can modulate the progressive neuropathy caused by moderate overexpression of Pmp22. Male transgenic rats (n = 84) were randomly assigned into three treatment groups: progesterone, progesterone antagonist (onapristone) and placebo control. Daily administration of progesterone elevated the steady-state levels of Pmp22 and Mpz mRNA in the sciatic nerve, resulting in enhanced Schwann cell pathology and a more progressive clinical neuropathy. In contrast, administration of the selective progesterone receptor antagonist reduced overexpression of Pmp22 and improved the CMT phenotype, without obvious side effects, in wild-type or transgenic rats. Taken together, these data provide proof of principle that the progesterone receptor of myelin-forming Schwann cells is a promising pharmacological target for therapy of CMT-1A.