RESUMO
BACKGROUND: Neuropathic pain (NP), which results from injury or lesion of the somatosensory nervous system, is intimately associated with glial cells. The roles of microglia and astrocytes in NP have been broadly described, while studies on oligodendrocytes have largely focused on axonal myelination. The mechanisms of oligodendrocytes and their interactions with other glial cells in NP development remain uncertain. METHODS: To explore the function of the interaction of the three glial cells and their interactions on myelin development in NP, we evaluated changes in NP and myelin morphology after a chronic constriction injury (CCI) model in mice, and used single-cell sequencing to reveal the subpopulations characteristics of oligodendrocytes, microglia, and astrocytes in the spinal cord tissues, as well as their relationship with myelin lesions; the proliferation and differentiation trajectories of oligodendrocyte subpopulations were also revealed using pseudotime cell trajectory and RNA velocity analysis. In addition, we identified chemokine ligand-receptor pairs between glial cells by cellular communication and verified them using immunofluorescence. RESULTS: Our study showed that NP peaked on day 7 after CCI in mice, a time at which myelin lesions were present in both the spinal cord and sciatic nerve. Oligodendrocytes, microglia, and astrocytes subpopulations in spinal cord tissue were heterogeneous after CCI and all were involved in suppressing the process of immune defense and myelin production. In addition, the differentiation trajectory of oligodendrocytes involved a unidirectional lattice process of OPC-1-Oligo-9, which was arrested at the Oligo-2 stage under the influence of microglia and astrocytes. And the CADM1-CADM1, NRP1-VEGFA interactions between glial cells are enhanced after CCI and they had a key role in myelin lesions and demyelination. CONCLUSIONS: Our study reveals the close relationship between the differentiation block of oligodendrocytes after CCI and their interaction with microglia and astrocytes-mediated myelin lesions and NP. CADM1/CADM1 and NRP-1/VEGFA may serve as potential therapeutic targets for use in the treatment of NP.
Assuntos
Camundongos Endogâmicos C57BL , Bainha de Mielina , Neuralgia , Neuroglia , Medula Espinal , Animais , Camundongos , Medula Espinal/patologia , Medula Espinal/metabolismo , Bainha de Mielina/patologia , Bainha de Mielina/metabolismo , Neuralgia/patologia , Neuralgia/metabolismo , Neuroglia/patologia , Neuroglia/metabolismo , Masculino , Análise de Célula ÚnicaRESUMO
Microglia are highly plastic immune cells which exist in a continuum of activation states. By shaping the function of oligodendrocyte precursor cells (OPCs), the brain cells which differentiate to myelin-forming cells, microglia participate in both myelin injury and remyelination during multiple sclerosis. However, the mode(s) of action of microglia in supporting or inhibiting myelin repair is still largely unclear. Here, we analysed the effects of extracellular vesicles (EVs) produced in vitro by either pro-inflammatory or pro-regenerative microglia on OPCs at demyelinated lesions caused by lysolecithin injection in the mouse corpus callosum. Immunolabelling for myelin proteins and electron microscopy showed that EVs released by pro-inflammatory microglia blocked remyelination, whereas EVs produced by microglia co-cultured with immunosuppressive mesenchymal stem cells promoted OPC recruitment and myelin repair. The molecular mechanisms responsible for the harmful and beneficial EV actions were dissected in primary OPC cultures. By exposing OPCs, cultured either alone or with astrocytes, to inflammatory EVs, we observed a blockade of OPC maturation only in the presence of astrocytes, implicating these cells in remyelination failure. Biochemical fractionation revealed that astrocytes may be converted into harmful cells by the inflammatory EV cargo, as indicated by immunohistochemical and qPCR analyses, whereas surface lipid components of EVs promote OPC migration and/or differentiation, linking EV lipids to myelin repair. Although the mechanisms through which the lipid species enhance OPC maturation still remain to be fully defined, we provide the first demonstration that vesicular sphingosine 1 phosphate stimulates OPC migration, the first fundamental step in myelin repair. From this study, microglial EVs emerge as multimodal and multitarget signalling mediators able to influence both OPCs and astrocytes around myelin lesions, which may be exploited to develop novel approaches for myelin repair not only in multiple sclerosis, but also in neurological and neuropsychiatric diseases characterized by demyelination.
Assuntos
Astrócitos/fisiologia , Doenças Desmielinizantes/fisiopatologia , Vesículas Extracelulares/fisiologia , Microglia/fisiologia , Bainha de Mielina/fisiologia , Remielinização/fisiologia , Animais , Astrócitos/patologia , Diferenciação Celular/fisiologia , Movimento Celular/fisiologia , Técnicas de Cocultura , Corpo Caloso/patologia , Corpo Caloso/fisiopatologia , Doenças Desmielinizantes/patologia , Modelos Animais de Doenças , Vesículas Extracelulares/patologia , Inflamação/patologia , Inflamação/fisiopatologia , Lisofosfatidilcolinas , Masculino , Células-Tronco Mesenquimais/fisiologia , Camundongos Endogâmicos C57BL , Microglia/patologia , Bainha de Mielina/patologia , Neuroproteção/fisiologia , Células Precursoras de Oligodendrócitos/patologia , Células Precursoras de Oligodendrócitos/fisiologia , Ratos Sprague-DawleyRESUMO
A 70-year-old woman was admitted to our hospital due to bilateral optic neuritis and left facial sensory disturbance that became exacerbated over 10 days. Both serum and cerebrospinal fluid (CSF) were negative for aquaporin 4 antibody and myelin oligodendrocyte glycoprotein antibody. A high level of myelin basic protein (MBP) in her CSF was observed. Brain MRI showed a high T2 signal and contrast enhancement of the bilateral optic nerve, intramedullary tract and central myelin lesion in the trigeminal nerve. After intravenous methylprednisolone pulse therapy, her visual impairment and facial sensory disturbance gradually improved. She was diagnosed with clinically isolated syndrome, based on 2017 McDonald criteria. A diagnosis of multiple sclerosis (MS) was suspected due to the trigeminal myelin lesion confined to the central myelin portion and high level of MBP in the CSF. Treatment with dimethyl fumarate has been effective for preventing recurrence in 13 months in this patient. The central-peripheral myelin transitional zone at the trigeminal nerve is located 1-6 mm from the pons, where central myelin changes to the peripheral myelin. This patient showed a high T2 signal at the trigeminal nerve 3 mm from the pons on MRI, suggesting the involvement of a central trigeminal myelin lesion. Findings of a central trigeminal myelin lesion on MRI may aid in differentiating between MS and seronegative neuromyelitis optica spectrum disorder.
Assuntos
Esclerose Múltipla , Neuromielite Óptica , Neurite Óptica , Idoso , Aquaporina 4/imunologia , Autoanticorpos/sangue , Feminino , Humanos , Esclerose Múltipla/imunologia , Bainha de Mielina , Glicoproteína Mielina-Oligodendrócito/imunologia , Neurite Óptica/diagnóstico , Neurite Óptica/etiologiaRESUMO
Oligodendrocytes are the myelinating cells of the central nervous system. The physiological importance of oligodendrocytes is highlighted by diseases such as multiple sclerosis, in which the myelin sheaths are degraded and the axonal signal transmission is compromised. In a healthy brain, spontaneous remyelination is rare, and newly formed myelin sheaths are thinner and shorter than the former ones. The myelination process requires the migration, proliferation, and differentiation of oligodendrocyte precursor cells (OPCs) and is influenced by proteins of the extracellular matrix (ECM), which consists of a network of glycoproteins and proteoglycans. In particular, the glycoprotein tenascin-C (Tnc) has an inhibitory effect on the differentiation of OPCs and the remyelination efficiency of oligodendrocytes. The structurally similar tenascin-R (Tnr) exerts an inhibitory influence on the formation of myelin membranes in vitro. When Tnc knockout oligodendrocytes were applied to an in vitro myelination assay using artificial fibers, a higher number of sheaths per single cell were obtained compared to the wild-type control. This effect was enhanced by adding brain-derived neurotrophic factor (BDNF) to the culture system. Tnr-/- oligodendrocytes behaved differently in that the number of formed sheaths per single cell was decreased, indicating that Tnr supports the differentiation of OPCs. In order to study the functions of tenascin proteins in vivo Tnc-/- and Tnr-/- mice were exposed to Cuprizone-induced demyelination for a period of 10 weeks. Both Tnc-/- and Tnr-/- mouse knockout lines displayed a significant increase in the regenerating myelin sheath thickness after Cuprizone treatment. Furthermore, in the absence of either tenascin, the number of OPCs was increased. These results suggest that the fine-tuning of myelin regeneration is regulated by the major tenascin proteins of the CNS.
Assuntos
Doenças Desmielinizantes , Células Precursoras de Oligodendrócitos , Tenascina , Animais , Cuprizona/efeitos adversos , Doenças Desmielinizantes/induzido quimicamente , Doenças Desmielinizantes/metabolismo , Modelos Animais de Doenças , Matriz Extracelular/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Bainha de Mielina/metabolismo , Células Precursoras de Oligodendrócitos/metabolismo , Tenascina/metabolismoRESUMO
Oligodendrocytes form myelin membranes and thereby secure the insulation of axons and the rapid conduction of action potentials. Diseases such as multiple sclerosis highlight the importance of this glial cell population for brain function. In the adult brain, efficient remyelination following the damage to oligodendrocytes is compromised. Myelination is characterized by proliferation, migration, and proper integration of oligodendrocyte precursor cells (OPCs). These processes are among others controlled by proteins of the extracellular matrix (ECM). As a prominent representative ECM molecule, tenascin-C (Tnc) exerts an inhibitory effect on the migration and differentiation of OPCs. The structurally similar paralogue tenascin-R (Tnr) is known to promote the differentiation of oligodendrocytes. The model of lysolecithin-induced demyelination of cerebellar slice cultures represents an important tool for the analysis of the remyelination process. Ex vivo cerebellar explant cultures of Tnc -/- and Tnr -/- mouse lines displayed enhanced remyelination by forming thicker myelin membranes upon exposure to lysolecithin. The inhibitory effect of tenascins on remyelination could be confirmed when demyelinated wildtype control cultures were exposed to purified Tnc or Tnr protein. In that approach, the remyelination efficiency decreased in a dose-dependent manner with increasing concentrations of ECM molecules added. In order to examine potential roles in a complex in vivo environment, we successfully established cuprizone-based acute demyelination to analyze the remyelination behavior after cuprizone withdrawal in SV129, Tnc -/- , and Tnr -/- mice. In addition, we documented by immunohistochemistry in the cuprizone model the expression of chondroitin sulfate proteoglycans that are inhibitory for the differentiation of OPCs. In conclusion, inhibitory properties of Tnc and Tnr for myelin membrane formation could be demonstrated by using an ex vivo approach.