RESUMEN
AIMS: Although mitochondrial abnormalities have been reported within paraspinal muscles in patients with axial weakness and neuromuscular disease as well as with ageing, the basis of respiratory deficiency in paraspinal muscles is not known. This study aimed to determine the extent and basis of respiratory deficiency in paraspinal muscles from cases undergoing surgery for degenerative spinal disease and post mortem cases without a history of spinal disease, where age-related histopathological changes were previously reported. METHODS: Cervical and lumbar paraspinal muscles were obtained peri-operatively from 13 patients and from six post mortem control cases (age range 18-82 years) without a neurological disease. Sequential COX/SDH (mitochondrial respiratory chain complex IV/complex II) histochemistry was performed to identify respiratory-deficient muscle fibres (lacking complex IV with intact complex II activity). Real-time polymerase chain reaction, long-range polymerase chain reaction and sequencing were used to identify and characterize mitochondrial DNA (mtDNA) deletions and determine mtDNA copy number status. Mitochondrial respiratory chain complex subunits were detected by immunohistochemistry. RESULTS: The density of respiratory-deficient fibres increased with age. On average, 3.96% of fibres in paraspinal muscles were respiratory-deficient (range 0-10.26). Respiratory deficiency in 36.8% of paraspinal muscle fibres was due to clonally expanded mtDNA deletions. MtDNA depletion accounted for further 13.5% of respiratory deficiency. The profile of immunohistochemically detected subunits of complexes was similar in respiratory-deficient fibres with and without mtDNA deletions or mtDNA depletion. CONCLUSIONS: Paraspinal muscles appeared to be particularly susceptible to age-related mitochondrial respiratory chain defects. Clonally expanded mtDNA deletions and focal mtDNA depletion may contribute towards the development of age-related postural abnormalities.
Asunto(s)
ADN Mitocondrial/genética , Eliminación de Gen , Músculos Respiratorios/patología , Adolescente , Adulto , Anciano , Anciano de 80 o más Años , Envejecimiento/genética , Envejecimiento/fisiología , Autopsia , Ciclooxigenasa 1/metabolismo , Femenino , Dosificación de Gen , Humanos , Inmunohistoquímica , Degeneración del Disco Intervertebral/patología , Captura por Microdisección con Láser , Masculino , Persona de Mediana Edad , Enfermedades Mitocondriales/patología , Fibras Musculares Esqueléticas/patología , Enfermedades Neuromusculares/patología , Reacción en Cadena de la Polimerasa , Postura/fisiología , Escoliosis/patología , Escoliosis/cirugía , Análisis de Secuencia de ADN , Adulto JovenRESUMEN
Multiple sclerosis (MS) is an inflammatory demyelinating disease of the central nervous system. Recent evidence suggests that dysfunction of surviving demyelinated axons and axonal degeneration contribute to the progression of MS. We review the evidence for and potential mechanisms of degeneration as well as dysfunction of chronically demyelinated axons in MS with particular reference to mitochondria, the main source of adenosine-5'-triphosphate in axons. Besides adenosine-5'-triphosphate production, mitochondria play an important role in calcium handling and produce reactive oxygen species. The mitochondrial changes in axons lacking healthy myelin sheaths as well as redistribution of sodium channels suggest that demyelinated axons would be more vulnerable to energy deficit than myelinated axons. A dysfunction of mitochondria in lesions as well as in the normal-appearing white and grey matter is increasingly recognized in MS and could be an important determinant of axonal dysfunction and degeneration. Mitochondria are a potential therapeutic target in MS.
Asunto(s)
Axones/fisiología , Mitocondrias/fisiología , Esclerosis Múltiple/fisiopatología , Adenosina Trifosfato/biosíntesis , Animales , Axones/patología , Axones/ultraestructura , Enfermedades Desmielinizantes/fisiopatología , Progresión de la Enfermedad , Metabolismo Energético , Humanos , Esclerosis Múltiple/patología , Esclerosis Múltiple/terapia , Vaina de Mielina/patología , Degeneración NerviosaRESUMEN
We present a case of cerebral fat embolism (CFE) that demonstrated evidence of diffuse white matter cytotoxic edema on diffusion-weighted magnetic resonance imaging, in addition to punctate hyperintensities on T2-weighted and diffusion-weighted imaging. The case suggests that CFE represents a combination of occlusive arteriolar disease and secondary neurotoxicity.
Asunto(s)
Edema Encefálico/diagnóstico , Edema Encefálico/etiología , Imagen de Difusión por Resonancia Magnética , Embolia Grasa/complicaciones , Embolia Grasa/diagnóstico , Embolia Intracraneal/complicaciones , Embolia Intracraneal/diagnóstico , Adolescente , Femenino , HumanosRESUMEN
Mitochondrial defects have been implicated in the degeneration of axons in a number of CNS disorders, including multiple sclerosis. Uniquely, mitochondria harbor the only non-nuclear DNA (mitochondrial DNA or mtDNA), which encodes functionally important subunits of the respiratory chain. The pattern of mitochondrial respiratory chain subunit expression provides important clues to the underlying mechanism of mitochondrial injury. In snap frozen tissue mitochondrial respiratory chain complex IV or cytochrome c oxidase (COX) activity may be determined using a well-established histochemical technique, COX histochemistry. Lack of COX activity may be the result of mtDNA mutations, degradation of transcripts of subunits, modification of subunits or inhibition of complexes. Mitochondria lacking complex IV activity, however, have not been further explored within axons in CNS disorders. By combining COX histochemistry with immunofluorescent labeling of mitochondrial proteins we describe a method to identify mitochondria lacking complex IV activity in CNS tissue and locate inactive mitochondria to axons using confocal microscopy. Inactive axonal mitochondria may then be further investigated using confocal microscopy to define the pattern of mitochondrial respiratory chain complex subunit expression. Our technique may be used to gain important clues to the underlying mechanisms of mitochondrial injury within axons in a number of CNS disorders and relevant animal models.
Asunto(s)
Axones/ultraestructura , Complejo IV de Transporte de Electrones/metabolismo , Mitocondrias/enzimología , Médula Espinal/ultraestructura , Análisis de Varianza , Axones/enzimología , Cianatos/farmacología , Inhibidores Enzimáticos/farmacología , Humanos , Mitocondrias/efectos de los fármacos , Esclerosis Múltiple/patología , Músculo Esquelético/enzimología , Músculo Esquelético/patología , Músculo Esquelético/ultraestructura , Cambios Post Mortem , Prostaglandina-Endoperóxido Sintasas/metabolismo , Azida Sódica/farmacologíaAsunto(s)
Leucocitos Mononucleares/metabolismo , Esclerosis Múltiple/metabolismo , Fagocitos/metabolismo , Receptores CCR5/biosíntesis , Receptores de Quimiocina/biosíntesis , Adulto , Autopsia , Femenino , Humanos , Inmunohistoquímica , Masculino , Persona de Mediana Edad , Receptores CCR1 , Receptores CCR8RESUMEN
Cytochrome c oxidase or mitochondrial respiratory chain complex IV is where over 90% of oxygen is consumed. The relationship between complex IV activity and mitochondrial proteins, which provides a guide to understanding the mechanisms in primary mitochondrial disorders, has been determined by histochemistry (activity) and immunohistochemistry in serial sections. In the central nervous system (CNS), mitochondrial activity and immunoreactivity have been determined in populations of cells in serial sections as capturing cells in more than one section is difficult. In this report we describe a method to determine complex IV activity in relation to mitochondrial proteins at a single cell level in the CNS. We performed complex IV histochemistry and immunohistochemistry consecutively in snap frozen sections. Although the product of complex IV histochemistry reduces the sensitivity of standard immunohistochemistry (secondary antibody and ABC method) the biotin-free Menapath polymer detection system (A. Menarini Diagnostics, Wokingham, UK) enables mitochondrial proteins to be detected following complex IV histochemistry. The co-occurring chromogens may then be separately visualised and analysed using multi-spectral imaging (Nuance system CRi, Woburn, MA). Our technique is applicable for exploring mitochondrial defects within single cells in a variety of CNS disorders and animal models of those diseases.
Asunto(s)
Complejo IV de Transporte de Electrones/análisis , Histocitoquímica/métodos , Inmunohistoquímica/métodos , Proteínas Mitocondriales/análisis , Neuroquímica/métodos , Adulto , Anciano , Animales , Respiración de la Célula/fisiología , Compuestos Cromogénicos/análisis , Compuestos Cromogénicos/metabolismo , Complejo IV de Transporte de Electrones/metabolismo , Femenino , Técnica del Anticuerpo Fluorescente/métodos , Humanos , Microtomía/métodos , Enfermedades Mitocondriales/diagnóstico , Enfermedades Mitocondriales/metabolismo , Enfermedades Mitocondriales/fisiopatología , Proteínas Mitocondriales/metabolismo , Polímeros , Valor Predictivo de las Pruebas , Ratas , Sensibilidad y Especificidad , Espectrofotometría/métodosRESUMEN
The interaction between chemokines and their receptors leads to selective recruitment of cells to foci of inflammation. Cross-sectional studies have reported significantly different expression of chemokine receptors CXCR3, CCR5 and CCR2 on peripheral blood lymphocytes in multiple sclerosis (MS) compared with controls. Cells expressing these receptors are likely to play a pathogenic role as suggested by studies of experimental autoimmune encephalomyelitis. Also, immunogenetic studies of nonfunctional CCR5 receptors in MS patients, due to 32delta deletion, demonstrated a delay in time to next relapse. The aims of this study were to detect any changes in the serial expression of chemokine receptors CCR2, CCR3, CCR5 and CXCR3 on peripheral blood CD4+ lymphocytes from patients with MS and to correlate the changes with relapses. Upregulation of CXCR3 expression on peripheral blood CD4+ lymphocytes was associated with all relapses and CCR5 expression was significantly affected with all relapses. Clinical recovery, with or without intravenous methylprednisolone treatment, coincided with the return of CXCR3 towards baseline in all but one case. Fluctuation in the expression of CXCR3 and CCR5 was also significantly greater in clinically stable patients with MS compared with controls, which may be due to subclinical disease activity. These findings provide further support for the view that CXCR3 and CCR5 antagonists could have a therapeutic value in MS.
Asunto(s)
Linfocitos/metabolismo , Esclerosis Múltiple Recurrente-Remitente/inmunología , Esclerosis Múltiple Recurrente-Remitente/metabolismo , Receptores de Quimiocina/metabolismo , Adulto , Anciano , Antiinflamatorios/administración & dosificación , Biomarcadores , Femenino , Citometría de Flujo , Humanos , Interferón gamma/metabolismo , Interleucina-4/metabolismo , Estudios Longitudinales , Masculino , Metilprednisolona/administración & dosificación , Persona de Mediana Edad , Esclerosis Múltiple Recurrente-Remitente/tratamiento farmacológico , Receptores CCR2 , Receptores CCR5/metabolismo , Receptores CXCR3 , Recurrencia , Regulación hacia Arriba/inmunologíaRESUMEN
Chemokines are likely to contribute to the pathogenesis of chronic inflammatory demyelinating polyneuropathy (CIDP), as evidenced by data from experimental autoimmune neuritis. The alpha and beta chemokines in the cerebrospinal fluid (CSF) and serum from patients with CIDP were analysed using an enzyme linked immunosorbent assay. CXCL9, CXCL10, and CCL3 were raised in the CSF in CIDP compared with controls and non-demyelinating neuropathies (p < 0.001). Although the CSF levels of CCL2 were significantly higher than the serum levels for all groups, the difference between groups was not significant. CXCL9, CXCL10, and CCL3 may contribute to the pathogenesis of CIDP by recruiting inflammatory T cells and monocytes to spinal nerve roots, while CCL2 is likely to play a physiological role.
Asunto(s)
Quimiocinas/sangre , Quimiocinas/líquido cefalorraquídeo , Polirradiculoneuropatía Crónica Inflamatoria Desmielinizante/sangre , Polirradiculoneuropatía Crónica Inflamatoria Desmielinizante/líquido cefalorraquídeo , Adulto , Anciano , Anciano de 80 o más Años , Progresión de la Enfermedad , Ensayo de Inmunoadsorción Enzimática , Femenino , Humanos , Masculino , Persona de Mediana EdadRESUMEN
OBJECTIVE: To define the chemokine profile in the CSF of patients with multiple sclerosis (MS) and compare it with three control groups; patients with benign headache (headache), non-inflammatory neurological diseases (NIND), and other inflammatory neurological diseases (IND). In addition, the correlations of CSF chemokine concentrations with chemokine receptor expression on CSF CD4(+) T cells and with clinical disease activity were assessed. METHODS: Forty three patients with MS, 24 with IND, 44 with NIND, and 12 with benign headache undergoing diagnostic or therapeutic lumbar puncture were included. Supernatant fluid from CSF was analysed for four beta (CCL2, CCL3, CCL4, CCL5) and two alpha (CXCL9, CXCL10)chemokines by enzyme linked immunosorbent assay (ELISA). Chemokine receptors CCR3, CCR5, and CXCR3 on CD4(+) T cells from eight patients with MS were analysed using directly conjugated fluorescent labelled monoclonal antibodies and flow cytometry. RESULTS: CXCL10, formerly interferon-gamma inducible protein-10 (IP-10), was significantly increased and CCL2, formerly monocyte chemoattractant protein-1 (MCP-1), was significantly reduced in the CSF of patients with MS and IND compared with those with benign headache and NIND. Concentrations of CXCL10 were significantly greater in patients with relapsing-remitting compared with secondary progressive MS and correlated significantly with CXCR3 expression on CSF CD4(+) T cells from patients with MS. Concentrations of CXCL10 decreased and CCL2 concentrations increased as time from the last relapse increased in patients with MS. CONCLUSION: Increased CXCL10 and decreased CCL2 concentrations in the CSF are associated with relapses in MS. Although serial values from individual patients were not available, this study suggests that CXCL10 and CCL2 may return towards baseline concentrations after a relapse. Correlation of CXCL10 with CD4(+) T cell expression of CXCR3 was consistent with its chemoattractant role for activated lymphocytes. Thus CXCL10 neutralising agents and CXCR3 receptor antagonists may be therapeutic targets in MS.