RESUMEN
Axon initial segment (AIS) cell surface proteins mediate key biological processes in neurons including action potential initiation and axo-axonic synapse formation. However, few AIS cell surface proteins have been identified. Here, we use antibody-directed proximity biotinylation to define the cell surface proteins in close proximity to the AIS cell adhesion molecule Neurofascin. To determine the distributions of the identified proteins, we use CRISPR-mediated genome editing for insertion of epitope tags in the endogenous proteins. We identify Contactin-1 (Cntn1) as an AIS cell surface protein. Cntn1 is enriched at the AIS through interactions with Neurofascin and NrCAM. We further show that Cntn1 contributes to assembly of the AIS extracellular matrix, and regulates AIS axo-axonic innervation by inhibitory basket cells in the cerebellum and inhibitory chandelier cells in the cortex.
Asunto(s)
Segmento Inicial del Axón , Fenómenos Biológicos , Segmento Inicial del Axón/metabolismo , Contactina 1/metabolismo , Biotinilación , Sinapsis/metabolismo , Axones/metabolismo , Proteínas de la Membrana/metabolismo , Anticuerpos/metabolismoRESUMEN
Methamphetamine (Meth), a commonly used central nervous system stimulant, is highly addictive. Currently, there is no effective treatment for Meth dependence and abuse, although cell adhesion molecules (CAMs) have been shown to play an important role in the formation and remodeling of synapses in the nervous system while also being involved in addictive behavior. Contactin 1 (CNTN1) is a CAM that is widely expressed in the brain; nevertheless, its role in Meth addiction remains unclear. Therefore, in the present study, we established mouse models of single and repeated Meth exposure and subsequently determined that CNTN1 expression in the nucleus accumbens (NAc) was upregulated in mice following single or repeated Meth exposure, whereas CNTN1 expression in the hippocampus was not significantly altered. Intraperitoneal injection of the dopamine receptor 2 antagonist haloperidol reversed Meth-induced hyperlocomotion and upregulation of CNTN1 expression in the NAc. Additionally, repeated Meth exposure also induced conditioned place preference (CPP) in mice and upregulated the expression levels of CNTN1, NR2A, NR2B, and PSD95 in the NAc. Using an AAV-shRNA-based approach to specifically silence CNTN1 expression in the NAc via brain stereotaxis reversed Meth-induced CPP and decreased the expression levels of NR2A, NR2B, and PSD95 in the NAc. These findings suggest that CNTN1 expression in the NAc plays an important role in Meth-induced addiction, and the underlying mechanism may be related to the expression of synapse-associated proteins in the NAc. The results of this study improved our understanding of the role of cell adhesion molecules in Meth addiction.
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Trastornos Relacionados con Anfetaminas , Estimulantes del Sistema Nervioso Central , Metanfetamina , Ratones , Animales , Metanfetamina/farmacología , Núcleo Accumbens , Contactina 1/metabolismo , Estimulantes del Sistema Nervioso Central/farmacología , Encéfalo/metabolismo , Trastornos Relacionados con Anfetaminas/metabolismoRESUMEN
Diabetes is frequently accompanied by cognitive impairment with insidious onset, and progressive cognitive and behavioral changes. ß-1, 3-galactosyltransferase 2 (B3galt2) contributes to glycosylation, showing a clue for neuronal apoptosis, proliferation and differentiation. However, the role of B3galt2 in diabetic cognitive dysfunction (DCD) has not been investigated. In the present study, we aimed to explore the role of B3galt2 in DCD. Additionally, the potential therapeutic effects of salidroside on DCD was also explored. Diabetic C57BL/6J mice showed cognitive dysfunction together with down-regulated B3galt2. Overexpression of B3galt2 reversed the cognitive decline of diabetic C57BL/6J. Moreover, cognitive impairment was aggravated in B3galt2+/- diabetic mice compared with C57BL/6J diabetic mice. Immunohistochemistry fluorescence indicated that B3galt2 and F3/Contactin were co-localized in the hippocampal regions. Importantly, the expression of F3/Contactin can be regulated by the manipulation of B3galt2, overexpression of which assuaged hippocampal neuronal damage, protected the synapsin, and reduced neuronal apoptosis in diabetic mice. Interestingly, SAL alleviated DCD and reversed the expression of B3galt2 in diabetic C57BL/6J mice. These findings indicate that inhibition of B3galt2/F3/Contactin pathway contributes to DCD, and participates in SAL reversed DCD.
Asunto(s)
Disfunción Cognitiva , Diabetes Mellitus Experimental , Ratones , Animales , Contactina 1/metabolismo , Ratones Endogámicos C57BL , Contactinas , Transducción de SeñalRESUMEN
BACKGROUND: Despite highly effective treatment strategies for patients with relapsing-remitting multiple sclerosis (RRMS), long-term neurodegeneration and disease progression are often considerable. Accurate blood-based biomarkers that predict long-term neurodegeneration are lacking. OBJECTIVE: To assess the predictive value of serum neurofilament-light (sNfL) and serum contactin-1 (sCNTN1) for long-term magnetic resonance imaging (MRI)-derived neurodegeneration in natalizumab-treated patients with RRMS. METHODS: sNfL and sCNTN1 were measured in an observational cohort of natalizumab-treated patients with RRMS at baseline (first dose) and at 3 months, Year 1, Year 2, and last follow-up (median = 5.2 years) of treatment. Disability progression was quantified using "EDSS-plus" criteria. Neurodegeneration was measured by calculating annualized percentage brain, ventricular, and thalamic volume change (PBVC, VVC, and TVC, respectively). Linear regression analysis was performed to identify longitudinal predictors of neurodegeneration. RESULTS: In total, 88 patients (age = 37 ± 9 years, 75% female) were included, of whom 48% progressed. Year 1 sNfL level (not baseline or 3 months) was associated with PBVC (standardized (std.) ß = -0.26, p = 0.013), VVC (standardized ß = 0.36, p < 0.001), and TVC (standardized ß = -0.24, p = 0.02). For sCNTN1, only 3-month level was associated with VVC (standardized ß = -0.31, p = 0.002). CONCLUSION: Year 1 (but not baseline) sNfL level was predictive for long-term brain atrophy in patients treated with natalizumab. sCNTN1 level did not show a clear predictive value.
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Encéfalo , Contactina 1 , Esclerosis Múltiple Recurrente-Remitente , Adulto , Femenino , Humanos , Masculino , Persona de Mediana Edad , Atrofia , Encéfalo/diagnóstico por imagen , Encéfalo/patología , Esclerosis Múltiple Recurrente-Remitente/diagnóstico por imagen , Esclerosis Múltiple Recurrente-Remitente/tratamiento farmacológico , Natalizumab/efectos adversos , Contactina 1/metabolismoRESUMEN
Influenza A viruses (IAVs) continuously challenge the poultry industry and human health. Elucidation of the host factors that modulate the IAV lifecycle is vital for developing antiviral drugs and vaccines. In this study, we infected A549 cells with IAVs and found that host protein contactin-1 (CNTN1), a member of the immunoglobulin superfamily, enhanced viral replication. Bioinformatic prediction and experimental validation indicated that the expression of CNTN1 was reduced by microRNA-200c (miR-200c) through directly targeting. We further showed that CNTN1-modulated viral replication in A549 cells is dependent on type I interferon signaling. Co-immunoprecipitation experiments revealed that CNTN1 specifically interacts with MAVS and promotes its proteasomal degradation by removing its K63-linked ubiquitination. Moreover, we discovered that the deubiquitinase USP25 is recruited by CNTN1 to catalyze the deubiquitination of K63-linked MAVS. Consequently, the CNTN1-induced degradation cascade of MAVS blocked RIG-I-MAVS-mediated interferon signaling, leading to enhanced viral replication. Taken together, our data reveal novel roles of CNTN1 in the type I interferon pathway and regulatory mechanism of IAV replication.
Asunto(s)
Proteínas Adaptadoras Transductoras de Señales/metabolismo , Contactina 1/metabolismo , Proteína 58 DEAD Box/metabolismo , Virus de la Influenza A/metabolismo , Gripe Humana/virología , MicroARNs/metabolismo , Receptores Inmunológicos/metabolismo , Ubiquitina Tiolesterasa/metabolismo , Células A549 , Interacciones Microbiota-Huesped , Humanos , Interferón Tipo I/metabolismo , Transducción de Señal , Ubiquitinación , Replicación ViralRESUMEN
Since the potential roles of extracellular vesicles secreted by adipose-derived mesenchymal stem cells (ADSCs) are not well understood in collagen metabolism, the purpose of this research was to evaluate the effects of ADSCs-extracellular vesicles in stress urinary incontinence and the regulatory mechanism of delivered microRNA-93 (miR-93). ADSCs were isolated and cultured, and ADSCs-extracellular vesicles were extracted and identified. Stress urinary incontinence primary fibroblasts or satellite cells were treated with ADSCs-extracellular vesicles to detect the expression of Elastin, Collagen I, and Collagen III in fibroblasts and Pax7 and MyoD in satellite cells. After transfecting ADSCs with miR-93 mimics or inhibitors, extracellular vesicles were isolated and treated with stress urinary incontinence primary fibroblasts or satellite cells to observe cell function changes. The online prediction and luciferase activity assay confirmed the targeting relationship between miR-93 and coagulation factor III (F3). The rescue experiment verified the role of ADSCs-extracellular vesicles carrying miR-93 in stress urinary incontinence primary fibroblasts and satellite cells by targeting F3. ADSCs-extracellular vesicles treatment upregulated expression of Elastin, Collagen I, and Collagen III in stress urinary incontinence primary fibroblasts and expression of Pax7 and MyoD in stress urinary incontinence primary satellite cells. miR-93 expression was increased in stress urinary incontinence primary fibroblasts or satellite cells treated with ADSCs-extracellular vesicles. Extracellular vesicles secreted by ADSCs could deliver miR-93 to fibroblasts and then negatively regulate F3 expression; ADSCs-extracellular vesicles could reverse the effect of F3 on extracellular matrix remodeling in stress urinary incontinence fibroblasts. miR-93 expression was also increased in stress urinary incontinence primary satellite cells treated by ADSCs-extracellular vesicles. Extracellular vesicles secreted by ADSCs were delivered to satellite cells through miR-93, which directly targets F3 expression and upregulates Pax7 and MyoD expression in satellite cells. Our study indicates that miR-93 delivered by ADSCs-extracellular vesicles could regulate extracellular matrix remodeling of stress urinary incontinence fibroblasts and promote activation of stress urinary incontinence satellite cells through targeting F3.
Asunto(s)
Vesículas Extracelulares/fisiología , Células Madre Mesenquimatosas/ultraestructura , Incontinencia Urinaria de Esfuerzo/prevención & control , Tejido Adiposo/citología , Animales , Células Cultivadas , Contactina 1/genética , Contactina 1/metabolismo , Fibroblastos/metabolismo , Fibroblastos/patología , Regulación de la Expresión Génica , Masculino , Células Madre Mesenquimatosas/citología , MicroARNs/genética , MicroARNs/metabolismo , Ratas , Ratas Sprague-Dawley , Transducción de Señal/genética , Incontinencia Urinaria de Esfuerzo/genética , Incontinencia Urinaria de Esfuerzo/metabolismo , Incontinencia Urinaria de Esfuerzo/patologíaRESUMEN
BACKGROUND AND OBJECTIVES: As autoantibodies to contactin-1 from patients with chronic inflammatory demyelinating polyradiculoneuropathy not only bind to the paranodes where they are supposed to cause conduction failure but also bind to other neuronal cell types, we aimed to investigate the effect of anti-contactin-1 autoantibodies on contactin-1 surface expression in cerebellar granule neurons, dorsal root ganglion neurons, and contactin-1-transfected human embryonic kidney 293 cells. METHODS: Immunocytochemistry including structured illumination microscopy and immunoblotting was used to determine expression levels of contactin-1 and/or sodium channels after long-term exposure to autoantibodies from 3 seropositive patients. For functional analysis of sodium channels, whole-cell recordings of sodium currents were performed on dorsal root ganglion neurons incubated with anti-contactin-1 autoantibodies. RESULTS: We found a reduction in contactin-1 expression levels on dorsal root ganglion neurons, cerebellar granule neurons, and contactin-1-transfected human embryonic kidney 293 cells and decreased dorsal root ganglion sodium currents after long-term exposure to anti-contactin-1 autoantibodies. Sodium channel density did not decrease. DISCUSSION: Our results demonstrate a direct effect of anti-contactin-1 autoantibodies on the surface expression of contactin-1 and sodium currents in dorsal root ganglion neurons. This may be the pathophysiologic correlate of sensory ataxia reported in these patients.
Asunto(s)
Autoanticuerpos/inmunología , Contactina 1/inmunología , Contactina 1/metabolismo , Ganglios Espinales/metabolismo , Ganglios Espinales/fisiopatología , Canales de Sodio/fisiología , Ganglios Espinales/inmunología , Células HEK293 , Humanos , Polineuropatías/inmunología , Sodio/metabolismo , Canales de Sodio/metabolismoRESUMEN
BACKGROUND: Exosomes have emerged as a vital player in cell-cell communication; however, whether airway epithelial cell (AEC)-generated exosomes participate in asthma development remains unknown. OBJECTIVE: Our aims were to characterize the AEC-secreted exosomes and the potentially functional protein(s) that may contribute to the proinflammatory effects of AEC exosomes in the dendritic cell (DC)-dominant airway allergic models and to confirm their clinical significance in patients with asthma. METHODS: Mice were treated with exosomes derived from house dust mite (HDM)-stimulated AECs (HDM-AEC-EXOs) or monocyte-derived DCs primed by HDM and/or contactin-1 (CNTN1). The numbers of DCs in the lung were determined by flow cytometry. Proteomic analysis of purified HDM-AEC-EXOs was performed. CNTN1 small interfering RNA was designed to probe its role in airway allergy, and γ-secretase inhibitor was used to determine involvement of the Notch pathway. RESULTS: HDM-AEC-EXOs facilitate the recruitment, proliferation, migration, and activation of monocyte-derived DCs in cell culture and in mice. CNTN1 in exosomes is a critical player in asthma pathology. RNA interference-mediated silencing and pharmaceutical inhibitors characterize Notch2 receptor as necessary for relaying the CNTN1 signal to activate TH2 cell/TH17 cell immune response. Studies of patients with asthma also support existence of the CNTN1-Notch2 axis that has been observed in cell and mouse models. CONCLUSION: This study's findings reveal a novel role for CNTN1 in asthma pathogenesis mediated through exosome secretion, indicating a potential strategy for the treatment of allergic airway inflammation.
Asunto(s)
Asma/inmunología , Contactina 1/metabolismo , Células Dendríticas/inmunología , Exosomas/metabolismo , Hipersensibilidad/inmunología , Mucosa Respiratoria/metabolismo , Células Th2/inmunología , Animales , Antígenos Dermatofagoides/inmunología , Movimiento Celular , Proliferación Celular , Células Cultivadas , Contactina 1/genética , Humanos , Ratones , Ratones Endogámicos C57BL , Monocitos/citología , ARN Interferente Pequeño/genética , Receptor Notch2/genética , Receptor Notch2/metabolismoRESUMEN
Contactin 1 (CNTN1) is a new oncogenic protein of prostate cancer (PC); its impact on PC remains incompletely understood. We observed CNTN1 upregulation in LNCaP cell-derived castration-resistant PCs (CRPC) and CNTN1-mediated enhancement of LNCaP cell proliferation. CNTN1 overexpression in LNCaP cells resulted in enrichment of the CREIGHTON_ENDOCRINE_THERAPY_RESISTANCE_3 gene set that facilitates endocrine resistance in breast cancer. The leading-edge (LE) genes (n = 10) of this enrichment consist of four genes with limited knowledge on PC and six genes novel to PC. These LE genes display differential expression during PC initiation, metastatic progression, and CRPC development, and they predict PC relapse following curative therapies at hazard ratio (HR) 2.72, 95% confidence interval (CI) 1.96-3.77, and p = 1.77 × 10-9 in The Cancer Genome Atlas (TCGA) PanCancer cohort (n = 492) and HR 2.72, 95% CI 1.84-4.01, and p = 4.99 × 10-7 in Memorial Sloan Kettering Cancer Center (MSKCC) cohort (n = 140). The LE gene panel classifies high-, moderate-, and low-risk of PC relapse in both cohorts. Additionally, the gene panel robustly predicts poor overall survival in clear cell renal cell carcinoma (ccRCC, p = 1.13 × 10-11), consistent with ccRCC and PC both being urogenital cancers. Collectively, we report multiple CNTN1-related genes relevant to PC and their biomarker values in predicting PC relapse.
Asunto(s)
Carcinogénesis/genética , Carcinoma de Células Renales/genética , Contactina 1/genética , Neoplasias Renales/genética , Proteínas de Neoplasias/genética , Recurrencia Local de Neoplasia/genética , Neoplasias de la Próstata Resistentes a la Castración/genética , Atlas como Asunto , Carcinogénesis/metabolismo , Carcinogénesis/patología , Carcinoma de Células Renales/diagnóstico , Carcinoma de Células Renales/mortalidad , Carcinoma de Células Renales/patología , Línea Celular Tumoral , Proliferación Celular , Estudios de Cohortes , Contactina 1/metabolismo , Bases de Datos Genéticas , Regulación Neoplásica de la Expresión Génica , Humanos , Neoplasias Renales/diagnóstico , Neoplasias Renales/mortalidad , Neoplasias Renales/patología , Masculino , Familia de Multigenes , Proteínas de Neoplasias/metabolismo , Recurrencia Local de Neoplasia/diagnóstico , Recurrencia Local de Neoplasia/mortalidad , Recurrencia Local de Neoplasia/patología , Pronóstico , Modelos de Riesgos Proporcionales , Próstata/metabolismo , Próstata/patología , Neoplasias de la Próstata Resistentes a la Castración/diagnóstico , Neoplasias de la Próstata Resistentes a la Castración/mortalidad , Neoplasias de la Próstata Resistentes a la Castración/patologíaRESUMEN
Molecular profiling of small extracellular vesicles (sEV) isolated from plasma of cancer patients emerges as promising strategy for biomarkers discovery. We investigated the proteomic profiles of sEV immunoselected using anti-CSPG4 antibodies from 15 melanoma patients' plasma. The proteomes of sEV separated into melanoma cell-derived (MTEX) and non-malignant cell-derived (NMTEX) were compared using high-resolution mass spectrometry. Paired analysis identified the MTEX-associated profile of 16 proteins that discriminated MTEX from NMETEX. We also identified the MTEX profile that discriminated between seven patients with no evidence of melanoma (NED) after therapy and eight with progressive disease (PD). Among 75 MTEX proteins overexpressed in PD patients, PDCD6IP (ALIX) had the highest discriminating value, while CNTN1 (contactin-1) was upregulated only in MTEX of NED patients. This is the first report documenting that proteomes of tumour-derived sEV in patients' plasma discriminate cancer from non-cancer and identify proteins with potential to serve as prognostic biomarkers in melanoma.
Asunto(s)
Exosomas/metabolismo , Vesículas Extracelulares/metabolismo , Melanoma/metabolismo , Plasma/metabolismo , Proteoma/metabolismo , Adulto , Anciano , Biomarcadores de Tumor/sangre , Proteoglicanos Tipo Condroitín Sulfato/inmunología , Proteoglicanos Tipo Condroitín Sulfato/metabolismo , Contactina 1/metabolismo , Progresión de la Enfermedad , Exosomas/química , Vesículas Extracelulares/química , Femenino , Humanos , Masculino , Espectrometría de Masas , Melanoma/química , Proteínas de la Membrana/inmunología , Proteínas de la Membrana/metabolismo , Persona de Mediana Edad , Plasma/química , Proteínas/metabolismoRESUMEN
BACKGROUND: Contactin1 (CNTN1), a member of the immunoglobulin superfamily, is known to correlate with tumor development and progression. Although recent studies have found that elevated CNTN1 has been demonstrated in some types of cancers, the expression and prognosis of CNTN1 in colorectal cancer (CRC) are unclear. Here, we aimed to determine the clinicopathological characteristics and prognostic role of CNTN1 in CRC patients. METHODS: The protein expression of CNTN1 in tumor tissues was evaluated by immunohistochemistry. In addition, the mRNA and protein expressions of CNTN1 were examined by qRT-PCR and Western blotting analysis in 40 matched adjacent normal mucosa samples. The relationships of CNTN1 with clinicopathological data and prognosis significance were analyzed. RESULTS: Immunohistochemical consequence suggested that the protein level of CNTN1 was obviously raised in CRC compared with adjacent normal mucosa tissues (56.9% vs 10.3%, P< 0.05). In addition, we detected a significant increase in CNTN1 mRNA and protein levels in CRC tissues compared with the matched adjacent normal mucosa tissues. Moreover, increased CNTN1 exprssion was significantly associated with tumor size, lymph node metastasis (LNM), tumor node-metastasis (TNM) stage and carcino-embryonic antigen (CEA) in clinical analysis. Kaplan-Meier analysis suggested that patients with CNTN1 over-expression showed worse overall survival (OS) (P= 0.001). Multivariate analysis indicated that high CNTN1 expression was an independent predictor for poor OS in CRC patients (P= 0.028). Further analysis revealed that patients with high CNTN1 combined with LNM present accurately predicted poorer outcome. CONCLUSION: Taken together, the findingsindicate that CNTN1 plays a significant role and serve as a potential biomarker for the prediction of adverse prognosis in CRC. Intriguingly, high express of CNTN1 + LNM-present combination may improve the accuracy of prognosis.
Asunto(s)
Neoplasias Colorrectales/metabolismo , Contactina 1/metabolismo , Adulto , Anciano , Anciano de 80 o más Años , Neoplasias Colorrectales/genética , Neoplasias Colorrectales/patología , Contactina 1/genética , Femenino , Humanos , Metástasis Linfática , Masculino , Persona de Mediana Edad , Pronóstico , Regulación hacia ArribaRESUMEN
Even with recent progress, cancer remains the second leading cause of death, outlining a need to widen the current understanding on oncogenic factors. Accumulating evidence from recent years suggest Contactin 1 (CNTN1)'s possession of multiple oncogenic activities in a variety of cancer types. CNTN1 is a cell adhesion molecule that is dysregulated in many human carcinomas and plays important roles in cancer progression and metastases. Abnormalities in CNTN1 expression associate with cancer progression and poor prognosis. Mechanistically, CNTN1 functions in various signaling pathways frequently altered in cancer, such as the vascular endothelial growth factor C (VEGFC)-VEGF receptor 3 (VEFGR3)/fms-related tyrosine kinase 4 (Flt4) axis, phosphatidylinositol 3-kinase (PI3K)/protein kinase B (AKT), Notch signaling pathway and epithelial-mesenchymal transition (EMT) process. These oncogenic events are resulted via interactions between tumor and stroma, which can be contributed by CNTN1, an adhesion protein. CNTN1 expression in breast cancer correlates with the expression of genes functioning in cancer-stroma interactions and skeletal system development. Evidence supports that CNTN1 promotes cancer-stromal interaction, resulting in activation of a complex network required for cancer progression and metastasis (bone metastasis for breast cancer). CNTN1 inhibitions has been proven to be effective in experimental models to reduce oncogenesis. In this paper, we will review CNTN1's alterations in cancer, its main biochemical mechanisms and interactions with its relevant cancer pathways.
Asunto(s)
Carcinogénesis/metabolismo , Contactina 1/metabolismo , Proteínas Oncogénicas/metabolismo , Animales , Carcinogénesis/genética , Carcinogénesis/patología , Contactina 1/genética , Humanos , Neurogénesis , Proteínas Oncogénicas/genética , Transducción de SeñalAsunto(s)
Contactina 1/metabolismo , Glomerulonefritis Membranosa/diagnóstico , Glomerulonefritis Membranosa/metabolismo , Polirradiculoneuropatía Crónica Inflamatoria Desmielinizante/diagnóstico , Polirradiculoneuropatía Crónica Inflamatoria Desmielinizante/metabolismo , Adulto , Glomerulonefritis Membranosa/complicaciones , Humanos , Masculino , Polirradiculoneuropatía Crónica Inflamatoria Desmielinizante/complicacionesRESUMEN
White matter injury caused by perinatal hypoxia-ischemia is characterized by myelination disorders; however, its pathophysiological mechanisms are not fully elucidated. The neurofascin 155 (NF155) protein, expressed in oligodendrocytes, is critical for myelination. Previous findings suggest that NF155 participates in the pathological mechanisms of developmental myelination disorders in hypoxic-ischemic cerebral white matter lesions, and it might regulate cytoskeletal changes. Therefore, we hypothesized that increased NF155 expression during the early stages of hypoxic oligodendrocyte injury helps normalize myelin sheath development and consequently improves neural function by repairing paranodal structures of myelin sheaths and regulating cytoskeletal changes. To test this hypothesis, we established a hypoxic-ischemic, mixed neonatal rat forebrain cell culture model. When NF155 expression was upregulated, synergistic effects occurred between this protein and the paranodal proteins CASPR and contactin. In addition, the expression of Rho GTPase family proteins that regulate key cytoskeletal pathways, myelin sheath structures, and functions were restored, and axonal structures acquired a clear and transparent appearance. These results suggest that NF155 may enable myelin sheath repair by repairing paranodal region structures and regulating oligodendrocyte cytoskeletal mechanisms. Overall, the present study provides new insights into the pathogenesis of hypoxic-ischemic cerebral white matter lesions.
Asunto(s)
Moléculas de Adhesión Celular/metabolismo , Hipoxia/metabolismo , Isquemia/metabolismo , Fibras Nerviosas Mielínicas/metabolismo , Factores de Crecimiento Nervioso/metabolismo , Remielinización , Animales , Animales Recién Nacidos , Moléculas de Adhesión Celular/genética , Moléculas de Adhesión Celular Neuronal/metabolismo , Contactina 1/metabolismo , Vaina de Mielina , Factores de Crecimiento Nervioso/genética , Neuroglía , Oligodendroglía , Cultivo Primario de Células , Ratas , Ratas Sprague-Dawley , Proteína de Unión al GTP cdc42/metabolismo , Proteína de Unión al GTP rac1/metabolismo , Proteínas de Unión al GTP rho/metabolismoRESUMEN
Central nervous system myelin is a multilayered membrane produced by oligodendrocytes to increase neural processing speed and efficiency, but the molecular mechanisms underlying axonal selection and myelin wrapping are unknown. Here, using combined morphological and molecular analyses in mice and zebrafish, we show that adhesion molecules of the paranodal and the internodal segment work synergistically using overlapping functions to regulate axonal interaction and myelin wrapping. In the absence of these adhesive systems, axonal recognition by myelin is impaired with myelin growing on top of previously myelinated fibers, around neuronal cell bodies and above nodes of Ranvier. In addition, myelin wrapping is disturbed with the leading edge moving away from the axon and in between previously formed layers. These data show how two adhesive systems function together to guide axonal ensheathment and myelin wrapping, and provide a mechanistic understanding of how the spatial organization of myelin is achieved.
Asunto(s)
Axones/fisiología , Sistema Nervioso Central/fisiología , Vaina de Mielina/fisiología , Moléculas de Adhesión de Célula Nerviosa/metabolismo , Animales , Animales Modificados Genéticamente , Adhesión Celular/fisiología , Moléculas de Adhesión Celular Neuronal/genética , Moléculas de Adhesión Celular Neuronal/metabolismo , Contactina 1/genética , Contactina 1/metabolismo , Femenino , Larva , Masculino , Ratones Endogámicos C57BL , Ratones Transgénicos , Vaina de Mielina/patología , Glicoproteína Asociada a Mielina/genética , Glicoproteína Asociada a Mielina/metabolismo , Moléculas de Adhesión de Célula Nerviosa/genética , Nervio Óptico/metabolismo , Nervio Óptico/patología , Pez Cebra/genética , Proteínas de Pez Cebra/genética , Proteínas de Pez Cebra/metabolismoRESUMEN
The fast and reliable propagation of action potentials along myelinated fibers relies on the clustering of voltage-gated sodium channels at nodes of Ranvier. Axo-glial communication is required for assembly of nodal proteins in the central nervous system, yet the underlying mechanisms remain poorly understood. Oligodendrocytes are known to support node of Ranvier assembly through paranodal junction formation. In addition, the formation of early nodal protein clusters (or prenodes) along axons prior to myelination has been reported, and can be induced by oligodendrocyte conditioned medium (OCM). Our recent work on cultured hippocampal neurons showed that OCM-induced prenodes are associated with an increased conduction velocity (Freeman et al., 2015). We here unravel the nature of the oligodendroglial secreted factors. Mass spectrometry analysis of OCM identified several candidate proteins (i.e., Contactin-1, ChL1, NrCAM, Noelin2, RPTP/Phosphacan, and Tenascin-R). We show that Contactin-1 combined with RPTP/Phosphacan or Tenascin-R induces clusters of nodal proteins along hippocampal GABAergic axons. Furthermore, Contactin-1-immunodepleted OCM or OCM from Cntn1-null mice display significantly reduced clustering activity, that is restored by addition of soluble Contactin-1. Altogether, our results identify Contactin-1 secreted by oligodendrocytes as a novel factor that may influence early steps of nodal sodium channel cluster formation along specific axon populations.
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Contactina 1/metabolismo , Hipocampo/metabolismo , Proteína Nodal/metabolismo , Oligodendroglía/metabolismo , Animales , Células Cultivadas , Sistema Nervioso Central/citología , Sistema Nervioso Central/metabolismo , Contactina 1/genética , Neuronas GABAérgicas/metabolismo , Hipocampo/citología , Ratones , Ratones Noqueados , Ratones Transgénicos , Proteína Nodal/genética , Unión Proteica/fisiología , Ratas , Ratas Sprague-Dawley , Ratas WistarRESUMEN
Epithelial cells form sheets and tubules in various epithelial organs and establish apicobasal polarity and asymmetric vesicle transport to provide functionality in these structures. However, the molecular mechanisms that allow epithelial cells to establish polarity are not clearly understood. Here, we present evidence that the kinase activity of the receptor tyrosine kinase for collagen, discoidin domain receptor 1 (DDR1), is required for efficient establishment of epithelial polarity, proper asymmetric protein secretion, and execution of morphogenic programs. Lack of DDR1 protein or inhibition of DDR1 kinase activity disturbed tubulogenesis, cystogenesis, and the establishment of epithelial polarity and caused defects in the polarized localization of membrane-type 1 matrix metalloproteinase (MT1-MMP), GP135, primary cilia, laminin, and the Golgi apparatus. Disturbed epithelial polarity and cystogenesis upon DDR1 inhibition was caused by excess ROCK (rho-associated, coiled-coil-containing protein kinase)-driven actomyosin contractility, and pharmacological inhibition of ROCK was sufficient to correct these defects. Our data indicate that a DDR1-ROCK signaling axis is essential for the efficient establishment of epithelial polarity.
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Actomiosina/metabolismo , Polaridad Celular/fisiología , Receptor con Dominio Discoidina 1/metabolismo , Células Epiteliales/metabolismo , Animales , Células CACO-2 , Cilios/metabolismo , Contactina 1/metabolismo , Perros , Femenino , Aparato de Golgi/metabolismo , Humanos , Laminina/metabolismo , Células de Riñón Canino Madin Darby , Masculino , Metaloproteinasa 14 de la Matriz/metabolismo , Ratones , Ratones Endogámicos C57BL , Organoides , Quinasas Asociadas a rho/metabolismoRESUMEN
Optic neuritis is one of the first manifestations of multiple sclerosis. Its pathogenesis is incompletely understood, but considered to be initiated by an auto-immune response directed against myelin sheaths of the optic nerve. Here, we demonstrate in two frequently used and well-validated mouse models of optic neuritis that ribbon synapses in the myelin-free retina are targeted by an auto-reactive immune system even before alterations in the optic nerve have developed. The auto-immune response is directed against two adhesion proteins (CASPR1/CNTN1) that are present both in the paranodal region of myelinated nerves as well as at retinal ribbon synapses. This occurs in parallel with altered synaptic vesicle cycling in retinal ribbon synapses and altered visual behavior before the onset of optic nerve demyelination. These findings indicate that early synaptic dysfunctions in the retina contribute to the pathology of optic neuritis in multiple sclerosis.
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Autoinmunidad , Esclerosis Múltiple/patología , Células Fotorreceptoras de Vertebrados/metabolismo , Sinapsis/metabolismo , Animales , Anticuerpos/metabolismo , Bovinos , Activación de Complemento , Contactina 1/metabolismo , Modelos Animales de Enfermedad , Encefalomielitis Autoinmune Experimental/sangre , Encefalomielitis Autoinmune Experimental/líquido cefalorraquídeo , Encefalomielitis Autoinmune Experimental/metabolismo , Encefalomielitis Autoinmune Experimental/patología , Células HEK293 , Humanos , Ratones Endogámicos C57BL , Esclerosis Múltiple/sangre , Esclerosis Múltiple/líquido cefalorraquídeo , Nervio Óptico/metabolismo , Nervio Óptico/patología , Neuritis Óptica/metabolismo , Neuritis Óptica/patología , Células Fotorreceptoras de Vertebrados/ultraestructura , Retina/metabolismo , Sinapsis/ultraestructura , Vesículas Sinápticas/metabolismoRESUMEN
BACKGROUND: Contactin1 (CNTN1) has been shown to play an important role in the invasion and metastasis of several tumors; however, the role of CNTN1 in breast cancer has not been fully studied. The purpose of this study is to investigate the role of CNTN1 in regulating tumor growth, migration and invasion in breast cancer. RESULTS: To investigate its function, CNTN1 was expressed in Hs578T cells. CNTN1 expression was confirmed by western blot, immunohistochemistry and real-time RT-PCR. The effect of CNTN1 overexpression on proliferation, migration and invasion of Hs578T breast cancer cells was assessed in vitro and in vivo. Our results showed that CNTN1 overexpression promoted Hs578T cell proliferation, cell cycle progression, colony formation, invasion and migration. Notably, overexpression of CNTN1 in Hs578T cells enhanced the growth of mouse xenograft tumors. CONCLUSIONS: CNTN1 promotes growth, metastasis and invasion of Hs578T breast cancer cell line. Thus, therapies targeting CNTN1 may prove efficacious for breast cancer. However, further investigation is required to understand the mechanism by which CNTN1 influences proliferation, metastasis and invasion in breast cancer.
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Neoplasias de la Mama/patología , Movimiento Celular , Contactina 1/metabolismo , Animales , Línea Celular Tumoral , Proliferación Celular , Femenino , Humanos , Masculino , Ratones Desnudos , Invasividad Neoplásica , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Purpose: Successful immunotherapies for IDHmut gliomas require better knowledge of T-cell target antigens. Here, we elucidated their antigen repertoire recognized by spontaneous T-cell responses using an unbiased proteomic approach.Experimental Design: Protein fractionations of tissue lysates from IDHmut gliomas (n = 4) were performed. Fractions were tested by IFNγ ELISpot assay for recognition through patients' T cells. Proteins of immunogenic fractions were identified by mass spectrometry and validated by in silico-predicted synthetic long peptides in patients of origin, additional IDHmut glioma patients (n = 16), and healthy donors (n = 13). mRNA and protein expression of immunogenic antigens was analyzed in tumor tissues and IDHmut glioma stem-like cells (GSC). HLA-A*02-restricted T-cell epitopes were functionally determined by short peptides and numbers of antigen-specific T cells by HLA-peptide tetramer analysis.Results: A total of 2,897 proteins were identified in immunogenic tumor fractions. Based on a thorough filter process, 79 proteins were selected as potential T-cell antigens. Twenty-six of these were recognized by the patients' T cells, and five of them (CRKII, CFL1, CNTN1, NME2, and TKT) in up to 56% unrelated IDHmut glioma patients. Most immunogenic tumor-associated antigens (TAA) were expressed in IDHmut gliomas and GSCs, while being almost absent in normal brain tissues. Finally, we identified HLA-A*02-restricted epitopes for CRKII, NME2, and TKT that were recognized by up to 2.82% of antigen-specific peripheral cytotoxic T cells in IDHmut glioma patients.Conclusions: By analyzing the repertoire of T-cell target antigens in IDHmut glioma patients, we identified five novel immunogenic TAAs and confirmed their expression on IDHmut tumors and GSCs. Clin Cancer Res; 24(12); 2951-62. ©2018 AACR.