RESUMEN
Purpose: Upregulation of type I interferon (IFN) signaling has been increasingly detected in inflammatory diseases. Recently, upregulation of the IFN signature has been suggested as a potential biomarker of IFN-driven inflammatory diseases. Yet, it remains unclear to what extent type I IFN is involved in the pathogenesis of undifferentiated inflammatory diseases. This study aimed to quantify the type I IFN signature in clinically undiagnosed patients and assess clinical characteristics in those with a high IFN signature. Methods: The type I IFN signature was measured in patients' whole blood cells. Clinical and biological data were collected retrospectively, and an intensive genetic analysis was performed in undiagnosed patients with a high IFN signature. Results: A total of 117 samples from 94 patients with inflammatory diseases, including 37 undiagnosed cases, were analyzed. Increased IFN signaling was observed in 19 undiagnosed patients, with 10 exhibiting clinical features commonly found in type I interferonopathies. Skin manifestations, observed in eight patients, were macroscopically and histologically similar to those found in proteasome-associated autoinflammatory syndrome. Genetic analysis identified novel mutations in the PSMB8 gene of one patient, and rare variants of unknown significance in genes linked to type I IFN signaling in four patients. A JAK inhibitor effectively treated the patient with the PSMB8 mutations. Patients with clinically quiescent idiopathic pulmonary hemosiderosis and A20 haploinsufficiency showed enhanced IFN signaling. Conclusions: Half of the patients examined in this study, with undifferentiated inflammatory diseases, clinically quiescent A20 haploinsufficiency, or idiopathic pulmonary hemosiderosis, had an elevated type I IFN signature.
Asunto(s)
Interferón Tipo I , Inhibidores de las Cinasas Janus , Biomarcadores , Humanos , Interferón Tipo I/genética , Japón , Complejo de la Endopetidasa Proteasomal/genética , Estudios RetrospectivosRESUMEN
Familial Mediterranean fever (FMF) is a hereditary, autoinflammatory disease that causes recurrent fever, arthritis, and serositis. The diagnosis of FMF is based on the presentation of typical clinical symptoms and the Mediterranean fever gene (MEFV) test. However, the challenge lies in diagnosing atypical cases. In this report, we have described a pediatric patient with complex FMF whose diagnosis required trio-whole exome sequencing (WES) and functional validation of a rare MEFV variant. A 3-year-old boy presented with recurrent episodes of elevated liver enzymes and arthralgia. He was diagnosed with autoimmune hepatitis (AIH), and his liver enzymes improved rapidly with steroid treatment. However, he exhibited recurrent arthralgia and severe abdominal attacks. Trio-WES identified compound heterozygous mutations in MEFV (V726A and I692del). Ex vivo functional assays of the patient's monocytes and macrophages, which had been pre-treated with Clostridium difficile toxin A (TcdA) and colchicine, were comparable to those of typical FMF patients, thereby confirming the diagnosis of FMF. Although he was intolerant to colchicine because of liver toxicity, subsequent administration of canakinumab successfully ameliorated his abdominal attacks. However, it was ineffective against liver injury, which recurred after steroid tapering. Therefore, in this case, the pathogenesis of AIH was probably interleukin-1ß (IL-1ß)-independent. In fact, AIH might have been a concurrent disease with FMF, rather than being one of its complications. Nevertheless, further studies are necessary to determine whether FMF-induced inflammasome activation contributes to AIH development. Moreover, we must consider the possibility of mixed phenotypes in such atypical patients who present distinct pathologies simultaneously.
Asunto(s)
Fiebre Mediterránea Familiar , Hepatitis Autoinmune , Artralgia , Niño , Colchicina/uso terapéutico , Fiebre Mediterránea Familiar/complicaciones , Fiebre Mediterránea Familiar/diagnóstico , Fiebre Mediterránea Familiar/tratamiento farmacológico , Hepatitis Autoinmune/complicaciones , Hepatitis Autoinmune/diagnóstico , Hepatitis Autoinmune/tratamiento farmacológico , Humanos , Masculino , Mutación , Pirina/genéticaRESUMEN
The concept of autoinflammation, first proposed in 1999, refers to a seemingly unprovoked episode of sterile inflammation manifesting as unexplained fever, skin rashes, and arthralgia. Autoinflammatory diseases are caused mainly by hereditary abnormalities of innate immunity, without the production of autoantibodies or autoreactive T cells. The revolutionary discovery of induced pluripotent stem cells (iPSCs), whereby a patient's somatic cells can be reprogrammed into an embryonic pluripotent state by forced expression of a defined set of transcription factors, has the transformative potential to enable in vitro disease modeling and drug candidate screening, as well as to provide a resource for cell replacement therapy. Recent reports demonstrate that recapitulating a disease phenotype in vitro is feasible for numerous monogenic diseases, including autoinflammatory diseases. In this review, we provide a comprehensive overview of current advances in research into autoinflammatory diseases involving iPSC-derived monocytes/macrophages. This review may aid in the planning of new studies of autoinflammatory diseases.
Asunto(s)
Enfermedades Autoinflamatorias Hereditarias , Células Madre Pluripotentes Inducidas , Enfermedades Autoinflamatorias Hereditarias/genética , Humanos , Inflamación , Macrófagos , MonocitosRESUMEN
PURPOSE: Pathogenic MEFV variants cause pyrin-associated autoinflammatory diseases (PAADs), which include familial Mediterranean fever (FMF), FMF-like disease, and pyrin-associated autoinflammation with neutrophilic dermatosis (PAAND). The diagnosis of PAADs is established by clinical phenotypic and genetic analyses. However, the pathogenicity of most MEFV variants remains controversial, as they have not been functionally evaluated. This study aimed to establish and validate a new functional assay to evaluate the pathogenicity of MEFV variants. METHODS: We transfected THP-1 monocytes with 32 MEFV variants and analyzed their effects on cell death with or without stimulation with Clostridium difficile toxin A (TcdA) or UCN-01. These variants were classified using hierarchical cluster analysis. Macrophages were obtained from three healthy controls and two patients with a novel homozygous MEFVP257L variant, for comparison of IL-1ß secretion using a cell-based assay and a novel THP-1-based assay. RESULTS: Disease-associated MEFV variants induced variable degrees of spontaneous or TcdA/UCN-01-induced cell death in THP-1. Cell death was caspase-1 dependent and was accompanied by ASC speck formation and IL-1ß secretion, indicating that pathogenic MEFV variants induced abnormal pyrin inflammasome activation and subsequent pyroptotic cell deaths in this assay. The MEFV variants (n = 32) exhibiting distinct response signatures were classified into 6 clusters, which showed a good correlation with the clinical phenotypes. Regarding the pathogenicity of MEFVP257L variants, the results were consistent between the cell-based assay and the THP-1-based assay. CONCLUSION: Our assay facilitates a rapid and comprehensive assessment of the pathogenicity of MEFV variants and contributes to a refined definition of PAAD subtypes.
Asunto(s)
Citometría de Flujo/métodos , Variación Genética/genética , Pirina/genética , Muerte Celular/genética , Línea Celular , Femenino , Predisposición Genética a la Enfermedad/genética , Humanos , Inflamasomas/genética , Masculino , Persona de Mediana Edad , Monocitos/metabolismo , Fenotipo , Células THP-1RESUMEN
BACKGROUND: Deficiency of adenosine deaminase 2 (DADA2) is an autosomal recessive inflammatory disease caused by loss-of-function mutations in both alleles of the ADA2 gene. Most patients with DADA2 exhibit systemic vasculopathy consistent with polyarteritis nodosa, but large phenotypic variability has been reported, and the pathogenesis of DADA2 remains unclear. OBJECTIVES: This study sought to assess the clinical and genetic characteristics of Japanese patients with DADA2 and to gain insight into the pathogenesis of DADA2 by multi-omics analysis. METHODS: Clinical and genetic data were collected from 8 Japanese patients with DADA2 diagnosed between 2016 and 2019. ADA2 variants in this cohort were functionally analyzed by in vitro overexpression analysis. PBMCs from 4 patients with DADA2 were subjected to transcriptome and proteome analyses. Patient samples were collected before and after introduction of anti- TNF-α therapies. Transcriptome data were compared with those of normal controls and patients with other autoinflammatory diseases. RESULTS: Five novel ADA2 variants were identified in these 8 patients and were confirmed pathogenic by in vitro analysis. Anti-TNF-α therapy controlled inflammation in all 8 patients. Transcriptome and proteome analyses showed that upregulation of type II interferon signaling was characteristic of DADA2. Network analysis identified STAT1 as a key regulator and a hub molecule in DADA2 pathogenesis, a finding supported by the hyperactivation of STAT1 in patients' monocytes and B cells after IFN-γ stimulation. CONCLUSIONS: Type II interferon signaling and STAT1 are associated with the pathogenesis of DADA2.
Asunto(s)
Adenosina Desaminasa/deficiencia , Agammaglobulinemia/inmunología , Péptidos y Proteínas de Señalización Intercelular/deficiencia , Interferón gamma/inmunología , Leucocitos Mononucleares/inmunología , Factor de Transcripción STAT1/inmunología , Inmunodeficiencia Combinada Grave/inmunología , Adenosina Desaminasa/inmunología , Adolescente , Adulto , Agammaglobulinemia/genética , Agammaglobulinemia/patología , Pueblo Asiatico , Niño , Preescolar , Femenino , Perfilación de la Expresión Génica , Humanos , Lactante , Péptidos y Proteínas de Señalización Intercelular/inmunología , Interferón gamma/genética , Japón , Leucocitos Mononucleares/patología , Masculino , Proteómica , Factor de Transcripción STAT1/genética , Inmunodeficiencia Combinada Grave/genética , Inmunodeficiencia Combinada Grave/patologíaRESUMEN
In 2013, a drug-coated balloon catheter (DCB) (SeQuent Please) for the treatment of coronary in-stent restenosis (ISR) was approved in Japan. The pre-marketing Japan domestic NP001 study demonstrated better outcomes of the DCB (n = 138) compared to plain balloon angioplasty (n = 72). After the introduction to marketing, a post-marketing surveillance (PMS) (n = 396) was conducted to evaluate the safety and efficacy of the DCB in Japanese routine clinical practice. The aim of this paper was to assess differences between the pre-marketing NP001 study and the PMS. Compared to the NP001 study, more complex lesions were treated in the PMS (type B2/C: 69.0% vs 20.4%, total occlusion: 11.2% vs 0%, p < 0.001, respectively) and target lesion was more frequently ISR related to drug-eluting stent (DES) (79.5% vs 39.4%, p < 0.001). Regarding clinical outcomes, the rate of target lesion revascularization (TLR) was higher in the PMS than in the NP001 study (TLR: 12.9% at 7 months and 17.6% at 12 months vs 2.8% at 6 months, p = 0.001, p < 0.001, respectively). Multivariable logistic regression analysis revealed that DES-ISR was a risk factor of TLR after DCB treatment for ISR (odds ratio: 5.77, 95% CI 1.75-18.95, p = 0.004). Among representative published trials using DCB for ISR, clinical outcomes are often worse in DES-ISR trials than those in bare metal stent-ISR trials. The rates of TLR in previous DES-ISR trials are similar to that in the current PMS (TLR at 12 months: 22.1% for ISAR-DESIRE 3, 15.3% for PEPCAD-DES, and 13.0% for RIBS IV). The effectiveness and safety of DCB for coronary ISR have been confirmed in the Japanese real-world survey. PMS would be useful to evaluate the safety and effectiveness of medical products throughout their total life cycles.
Asunto(s)
Angioplastia Coronaria con Balón/métodos , Materiales Biocompatibles Revestidos , Reestenosis Coronaria/cirugía , Stents Liberadores de Fármacos , Anciano , Angiografía Coronaria , Reestenosis Coronaria/diagnóstico , Femenino , Humanos , Japón , Masculino , Paclitaxel , Diseño de Prótesis , Factores de Riesgo , Factores de Tiempo , Resultado del TratamientoAsunto(s)
Fiebre Mediterránea Familiar/genética , Genotipo , Células Madre Pluripotentes Inducidas/fisiología , Inflamasomas/metabolismo , Macrófagos/fisiología , Prurito/genética , Pirina/genética , Autofagia/genética , Diferenciación Celular , Células Cultivadas , Predisposición Genética a la Enfermedad , Humanos , Linaje , Polimorfismo Genético , ARN Interferente Pequeño/genéticaRESUMEN
X-linked dominant incontinentia pigmenti (IP) and X-linked recessive anhidrotic ectodermal dysplasia with immunodeficiency (EDA-ID) are caused by loss-of-function and hypomorphic IKBKG (also known as NEMO) mutations, respectively. We describe a European mother with mild IP and a Japanese mother without IP, whose 3 boys with EDA-ID died from ID. We identify the same private variant in an intron of IKBKG, IVS4+866 C>T, which was inherited from and occurred de novo in the European mother and Japanese mother, respectively. This mutation creates a new splicing donor site, giving rise to a 44-nucleotide pseudoexon (PE) generating a frameshift. Its leakiness accounts for NF-κB activation being impaired but not abolished in the boys' cells. However, aberrant splicing rates differ between cell types, with WT NEMO mRNA and protein levels ranging from barely detectable in leukocytes to residual amounts in induced pluripotent stem cell-derived (iPSC-derived) macrophages, and higher levels in fibroblasts and iPSC-derived neuronal precursor cells. Finally, SRSF6 binds to the PE, facilitating its inclusion. Moreover, SRSF6 knockdown or CLK inhibition restores WT NEMO expression and function in mutant cells. A recurrent deep intronic splicing mutation in IKBKG underlies a purely quantitative NEMO defect in males that is most severe in leukocytes and can be rescued by the inhibition of SRSF6 or CLK.
Asunto(s)
Displasia Ectodérmica , Mutación del Sistema de Lectura , Quinasa I-kappa B , Incontinencia Pigmentaria , Intrones , Displasia Ectodérmica/genética , Displasia Ectodérmica/metabolismo , Displasia Ectodérmica/patología , Femenino , Fibroblastos/metabolismo , Fibroblastos/patología , Humanos , Quinasa I-kappa B/deficiencia , Quinasa I-kappa B/metabolismo , Síndromes de Inmunodeficiencia/genética , Síndromes de Inmunodeficiencia/metabolismo , Síndromes de Inmunodeficiencia/patología , Incontinencia Pigmentaria/genética , Incontinencia Pigmentaria/metabolismo , Incontinencia Pigmentaria/patología , Macrófagos/metabolismo , Macrófagos/patología , MasculinoRESUMEN
Herein we describe the case of a 7-month-old girl with Kawasaki disease (KD) in whom status epilepticus with fever developed on day 3 and cluster of seizures on day 6 of illness, followed by severe disturbance of consciousness afterward. Diffusion-weighted magnetic resonance imaging on day 6 of illness showed diffuse high signals in the bilateral subcortical white matter, while electroencephalogram indicated low-voltage slow waves. This indicated acute encephalopathy with biphasic seizures and late reduced diffusion (AESD); severe neurological sequelae remained. This is the first report of AESD as a complication of KD.
Asunto(s)
Encefalopatías/etiología , Síndrome Mucocutáneo Linfonodular/complicaciones , Convulsiones/etiología , Enfermedad Aguda , Encefalopatías/diagnóstico por imagen , Imagen de Difusión por Resonancia Magnética , Femenino , Humanos , Lactante , Síndrome Mucocutáneo Linfonodular/diagnóstico , Convulsiones/diagnóstico por imagen , Estado Epiléptico/diagnóstico por imagen , Estado Epiléptico/etiologíaRESUMEN
Parkinson's disease (PD) is a common neurodegenerative disorder that is characterized by the degeneration of dopaminergic neurons in the substantia nigra and dopamine depletion in the striatum. Although the motor symptoms are still regarded as the main problem, non-motor symptoms in PD also markedly impair the quality of life. Several non-motor symptoms, such as sleep disturbances and depression, are suggested to be implicated in the alteration in circadian clock function. In this study, we investigated circadian disruption and the mechanism in a 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) mouse model of PD. MPTP-treated mice exhibited altered 24-h rhythms in body temperature and locomotor activity. In addition, MPTP treatment also affected the circadian clock system at the genetic level. The exposure of human neuroblastoma cells (SH-SY5Y) to 1-metyl-4-phenylpyridinium (MPP(+)) increased or decreased the mRNA levels of several clock genes in a dose-dependent manner. MPP(+)-induced changes in clock genes expression were reversed by Compound C, an inhibitor of AMP-activated protein kinase (AMPK). Most importantly, addition of ATP to the drinking water of MPTP-treated mice attenuated neurodegeneration in dopaminergic neurons, suppressed AMPK activation and prevented circadian disruption. The present findings suggest that the activation of AMPK caused circadian dysfunction, and ATP may be a novel therapeutic strategy based on the molecular clock in PD.
Asunto(s)
Trastornos Cronobiológicos/inducido químicamente , Intoxicación por MPTP/fisiopatología , 1-Metil-4-fenil-1,2,3,6-Tetrahidropiridina/antagonistas & inhibidores , Proteínas Quinasas Activadas por AMP/antagonistas & inhibidores , Factores de Transcripción ARNTL/biosíntesis , Factores de Transcripción ARNTL/genética , Adenosina Trifosfato/uso terapéutico , Animales , Temperatura Corporal/efectos de los fármacos , Temperatura Corporal/fisiología , Dominio Catalítico/efectos de los fármacos , Línea Celular Tumoral , Trastornos Cronobiológicos/genética , Criptocromos/biosíntesis , Criptocromos/genética , Relación Dosis-Respuesta a Droga , Regulación de la Expresión Génica/efectos de los fármacos , Humanos , Intoxicación por MPTP/tratamiento farmacológico , Intoxicación por MPTP/genética , Masculino , Ratones , Ratones Endogámicos C57BL , Actividad Motora/efectos de los fármacos , Actividad Motora/fisiología , Neostriado/efectos de los fármacos , Neostriado/fisiología , Neuroblastoma/patología , Miembro 1 del Grupo D de la Subfamilia 1 de Receptores Nucleares/biosíntesis , Miembro 1 del Grupo D de la Subfamilia 1 de Receptores Nucleares/genética , Fosforilación/efectos de los fármacos , Procesamiento Proteico-Postraduccional/efectos de los fármacos , Pirazoles/farmacología , Pirimidinas/farmacologíaRESUMEN
Tumor necrosis factor-α (TNF-α) is one of the main mediators of inflammatory response activated by fatty acids in obesity, and this signaling through TNF-α receptor (TNFR) is responsible for obesity-associated insulin resistance. Recently, TNF-α has shown to affect lipid metabolism including the regulation of lipase activity and bile acid synthesis. However, there is scanty in vivo evidence for the involvement of TNF-α in this process, and the mechanistic role of TNFR remains unclear. In this study, TNFR2 knockout mice (R2KO) and wild-type (WT) mice were fed commercial normal diet (ND) or high-fat diet (HFD) for 8 weeks. In R2KO/HFD mice, the increase in body weight and the accumulation of fat were significantly ameliorated compared with WT/HFD mice in association with the decrease in plasma total cholesterol (137.7±3.1 vs. 98.6±3.1 mg/dL, P<0.005), glucose (221.9±14.7 vs. 167.3±8.1 mg/dL, P<0.01), and insulin (5.1±0.3 vs. 3.4±0.3 ng/mL, P<0.05). Fecal excretion of lipid contents was significantly increased in R2KO mice. In R2KO/HFD mice, the decrease in hepatic cholesterol-7a-hydroxylase activity, the rate-limiting enzyme in bile acid synthesis, was inhibited (1.7±0.2 vs. 8.1±1.0 pmol/min/mg protein, P<0.01). These results suggested that HFD-induced obesity with metabolic derangements could be ameliorated in mice lacking TNF-α receptor 2 via increasing fecal bile acid and lipid content excretion. Therefore, TNF-α signaling through TNFR2 is essentially involved in the bile acid synthesis and excretion of lipids, resulting in its beneficial effects.
Asunto(s)
Ácidos y Sales Biliares/fisiología , Dieta Alta en Grasa , Resistencia a la Insulina , Obesidad , Receptores Tipo II del Factor de Necrosis Tumoral/deficiencia , Animales , Ácidos y Sales Biliares/metabolismo , Heces/química , Metabolismo de los Lípidos , Lípidos/biosíntesis , Ratones , Ratones Noqueados , Factor de Necrosis Tumoral alfaRESUMEN
Oxidative stress may be involved in the dopaminergic neurodegenerations seen in 6-OHDA-lesioned rats through its production of free radicals and through mitochondrial dysfunction. In this study, we noninvasively demonstrate brain redox alterations in 6-OHDA-lesioned rats using Overhauser-enhanced magnetic resonance imaging (OMRI). The reduction rate of 3-methoxycarbonyl-2,2,5,5-tetramethylpyrrolidine-l-oxyl (methoxycarbonyl-PROXYL), a redox-sensitive contrast agent, was used as an index of the redox status in vivo. The methoxycarbonyl-PROXYL reduction rate, calculated from continuous images, decreased significantly in lesioned hemispheres compared to their corresponding contralateral hemispheres. The reduction rates in cellular fractions obtained from the striatum were estimated by X-band electron spin resonance (ESR) and calculated by assuming first-order kinetics for their time-dependent decreases. When methoxycarbonyl-PROXYL was mixed with cytoplasm fractions, the reduction rates were the same in both hemispheres. However, the ESR signal of methoxycarbonyl-PROXYL in the mitochondrial fraction of the lesioned hemispheres decayed more slowly than that of the corresponding contralateral hemispheres. Concordantly, biochemical assays showed that the activity of mitochondrial complex I also decreased more slowly in lesioned hemispheres. Thus, this method of noninvasively imaging brain redox alterations faithfully reflects changes in mitochondrial complex I activity in 6-OHDA-lesioned rats.
Asunto(s)
Cuerpo Estriado/metabolismo , Modelos Animales de Enfermedad , Metabolismo Energético/fisiología , Imagen por Resonancia Magnética/métodos , Mitocondrias/patología , Estrés Oxidativo/fisiología , Oxidopamina/administración & dosificación , Trastornos Parkinsonianos/metabolismo , Trastornos Parkinsonianos/patología , Animales , Cuerpo Estriado/efectos de los fármacos , Cuerpo Estriado/fisiopatología , Metabolismo Energético/efectos de los fármacos , Masculino , Mitocondrias/fisiología , Oxidación-Reducción , Estrés Oxidativo/efectos de los fármacos , Trastornos Parkinsonianos/inducido químicamente , Ratas , Ratas Sprague-Dawley , Fracciones SubcelularesRESUMEN
Abuse of the powerfully addictive psychostimulant, methamphetamine, occurs worldwide. Recent studies have suggested that methamphetamine-induced dopaminergic neurotoxicity is related to oxidative stress. In response to nerve activation, the mitochondrial respiratory chain is rapidly activated. The enhancement of mitochondrial respiratory chain activation may induce oxidative stress in the brain. However, there is little experimental evidence regarding the mitochondrial function after methamphetamine administration in vivo. Here, we evaluated whether a single administration of methamphetamine induces ATP consumption and overactivation of mitochondria. We measured mitochondrial function in two different ways: by monitoring oxygen partial pressure using an oxygen-selective electrode, and by imaging of redox reactions using a nitroxyl radical (i.e., nitroxide) coupled with Overhauser-enhanced magnetic resonance imaging (OMRI). A single administration of methamphetamine to Wistar rats induced dopaminergic nerve activation, ATP consumption and an increase in mitochondrial respiratory chain function in both the striatum and cortex. Furthermore, antioxidant TEMPOL prevented the increase in mitochondrial oxidative damage and methamphetamine-induced sensitization. These findings suggest that energy-supplying reactions after dopaminergic nerve activation are associated with oxidative stress in both the striatum and cortex, leading to abnormal behavior.
Asunto(s)
Encéfalo/efectos de los fármacos , Estimulantes del Sistema Nervioso Central/farmacología , Metanfetamina/farmacología , Mitocondrias/efectos de los fármacos , Estrés Oxidativo/efectos de los fármacos , Adenosina Trifosfato/metabolismo , Animales , Encéfalo/metabolismo , Transporte de Electrón/efectos de los fármacos , Transporte de Electrón/fisiología , Procesamiento de Imagen Asistido por Computador , Imagen por Resonancia Magnética , Masculino , Mitocondrias/metabolismo , Ratas , Ratas WistarRESUMEN
Stroke is a major cause of mortality and morbidity in hypertensive patients. This study investigated the effects of nifedipine, an L-type voltage-gated Ca(2+) channel blocker, on ischemic lesion volume after focal cerebral ischemia and reperfusion in rats. Rats were subjected to 1 h of transient middle cerebral artery occlusion (MCAO). At 2 days after MCAO, the rats were randomized into two groups that were fed either a normal control diet (n=10) or a nifedipine (0.001%) containing diet (n=11) for 2 weeks. Nifedipine treatment significantly reduced ischemic lesion volume (116.5 ± 10.8 vs. 80.0 ± 8.2 mm(3), P < 0.05) without affecting body weight or blood pressure. It also decreased thiobarbituric-reactive substances, an index of lipid peroxide, (2.6 ± 0.4 vs. 1.7 ± 0.1 µmol g(-1) tissue, P < 0.05) and increased glutathione peroxidase (54.9 ± 4.7 vs. 70.9 ± 6.4 U g(-1) protein, P < 0.05) and glutathione reductase activities (32.4 ± 1.4 vs. 39.9 ± 2.7 U g(-1) protein, P < 0.05) in the mitochondria from the ischemic hemispheres. These results suggest that nifedipine treatment can reduce ischemic lesion volume after focal cerebral ischemia, possibly because of the decrease in oxidative stress with an increase in antioxidant activities within the ischemic area.
Asunto(s)
Antioxidantes/uso terapéutico , Isquemia Encefálica/tratamiento farmacológico , Encéfalo/efectos de los fármacos , Nifedipino/uso terapéutico , Accidente Cerebrovascular/tratamiento farmacológico , Animales , Antioxidantes/farmacología , Encéfalo/metabolismo , Encéfalo/patología , Isquemia Encefálica/metabolismo , Isquemia Encefálica/patología , Masculino , Mitocondrias/efectos de los fármacos , Mitocondrias/metabolismo , Mitocondrias/patología , Nifedipino/farmacología , Estrés Oxidativo/efectos de los fármacos , Ratas , Ratas Wistar , Daño por Reperfusión/tratamiento farmacológico , Daño por Reperfusión/metabolismo , Daño por Reperfusión/patología , Accidente Cerebrovascular/metabolismo , Accidente Cerebrovascular/patologíaRESUMEN
Oxidative stress is believed to be an important mechanism underlying dopamine-induced neuronal damage. This study provides X-band electron spin resonance (ESR) spectroscopic evidence for reactive oxygen species (ROS) generation during dopamine metabolism. The authors induced excess dopamine metabolism in the mouse striatum by bathing it in tyramine-containing perfusate using microdialysis. The addition of tyramine to the perfusate raised the levels of extracellular dopamine and hydrogen peroxide significantly. The ESR signal from hydroxy-TEMPO decayed during tyramine perfusion and treatment with a monoamine-oxidase inhibitor or radical scavenger suppressed the signal decay. Decreases in the number of tyrosine hydroxylase-immunopositive fibres and in dopamine concentration after tyramine perfusion were observed. Moreover, the tyramine-perfused mice showed a marked methamphetamine-induced rotational response. Notably, these effects of tyramine were suppressed by the simultaneous perfusion of hydroxy-TEMPO. These findings indicate that the ROS generation, which was monitored by hydroxy-TEMPO, caused oxidative damage to the dopaminergic neurons.
Asunto(s)
Dopamina/metabolismo , Degeneración Nerviosa/metabolismo , Neuronas/metabolismo , Estrés Oxidativo/fisiología , Especies Reactivas de Oxígeno/metabolismo , Inhibidores de Captación Adrenérgica/farmacología , Animales , Antioxidantes/farmacología , Encéfalo/efectos de los fármacos , Encéfalo/metabolismo , Encéfalo/patología , Óxidos N-Cíclicos/farmacología , Espectroscopía de Resonancia por Spin del Electrón , Inmunohistoquímica , Masculino , Ratones , Ratones Endogámicos C57BL , Microdiálisis , Neuronas/efectos de los fármacos , Neuronas/patología , Tiramina/farmacologíaRESUMEN
Free radicals are believed to be key factors that promote ischemia reperfusion injury in the brain. This study used the characteristic spectrum of methoxycarbonyl-PROXYL to detect free radical reactions in hydrophilic and lipophilic compartments in a transient middle cerebral artery occlusion (MCAO) mouse model. Methoxycarbonyl-PROXYL, which has a high water/octanol partition coefficient, allows the detection of nitroxyl radical in both compartments simultaneously. Free radicals generation was analysed from the enhanced ESR signal decay rate of methoxycarbonyl-PROXYL. The signal decay rate in the lipidic compartment was significantly enhanced 1 h after reperfusion following MCAO. The enhanced signal decay rate was significantly suppressed by Trolox. The accumulation of lipid peroxidation products increased by 6 h post-reperfusion and was suppressed by methoxycarbonyl-PROXYL or Trolox. These results demonstrate that information pertaining to different sites of free radical generation in vivo can be obtained simultaneously and that lipid-derived radicals are generated in transient MCAO mice.
Asunto(s)
Encéfalo/metabolismo , Espectroscopía de Resonancia por Spin del Electrón , Infarto de la Arteria Cerebral Media/metabolismo , Peroxidación de Lípido , Óxidos de Nitrógeno/metabolismo , Animales , Antioxidantes/farmacología , Encéfalo/efectos de los fármacos , Cromanos/farmacología , Óxidos N-Cíclicos/química , Modelos Animales de Enfermedad , Infarto de la Arteria Cerebral Media/tratamiento farmacológico , Peroxidación de Lípido/efectos de los fármacos , Masculino , Ratones , Octanoles/química , Pirrolidinas/química , Solventes/química , Marcadores de Spin , Factores de Tiempo , Agua/químicaRESUMEN
Oxidative stress has been implicated in the cell death that occurs after ischemia-reperfusion of the brain, which causes the production of reactive oxygen species and a decrease in antioxidants, leading to mitochondrial dysfunction. However, the invasive methods used to collect much of this evidence are themselves stress inducing, which could skew the results. In this study, we aimed at demonstrating brain redox alterations after ischemia-reperfusion noninvasively, using Overhauser-enhanced magnetic resonance imaging. The reduction rate of 3-methoxycarbonyl-2,2,5,5-tetramethylpyrrolidine-L-oxyl (methoxycarbonyl-PROXYL), a redox-sensitive contrast agent, was used as an index of the redox status in vivo. No changes were observed in the antioxidant concentration, the mitochondrial complex activity, or in the redox status image intensity after 3 h of reperfusion, following transient middle cerebral artery occlusion; however, after 24 h of reperfusion, the methoxycarbonyl-PROXYL reduction rate, calculated from continuous images, had decreased significantly. Concordantly, biochemical assays showed that the concentration of ascorbic acid in the ischemic hemisphere and the activity of mitochondrial complex II had also decreased. Thus, the noninvasive imaging of the brain redox alterations faithfully reflected changes in antioxidant levels and in mitochondrial complex II activity after ischemia-reperfusion.
Asunto(s)
Encéfalo/metabolismo , Infarto de la Arteria Cerebral Media/metabolismo , Imagen por Resonancia Magnética/métodos , Animales , Antioxidantes/análisis , Ácido Ascórbico/análisis , Medios de Contraste , Cinética , Masculino , Proteínas Mitocondriales/análisis , Oxidación-Reducción , Ratas , Ratas Wistar , Reperfusión , Factores de TiempoRESUMEN
Nitroxyl radicals are important antioxidants that have been used to protect animal tissues from oxidative damage. Their reaction with hydroxyl radical ((*)OH) is generally accepted to be the mechanism of antioxidant function. However, the direct interaction of nitroxyl radicals with (*)OH does not always provide a satisfactory explanation in various pH, because the concentration of hydrogen ion may affect the generation of secondary (*)OH-derived radicals. In the present study, it was confirmed that the reaction between 4-hydroxy-2,2,6,6-tetramethylpiperidine-N-oxyl (TEMPOL) and (*)OH generated TEMPOL-hydroxylamine, 4-oxo-2,2,6,6-tetramethylpiperidine-N-oxyl (TEMPON) and TEMPON-hydroxylamine using HPLC coupled with electrochemical detection. In the absence of NADH, TEMPOL-H may be generated by the reaction with secondary (*)OH-derived radicals in acidic condition. In the presence of NADH, a large proportion of the non-paramagnetic products was TEMPOL-H. Finally, it was clarified that TEMPOL-H was generated during dopamine metabolism, which is believed to be one of the (*)OH sources in pathological processes such as Parkinson's disease.