RESUMEN
We report on the development of a novel assay protocol for the separation and detection of charge isoforms of DJ-1 in biological samples by automated capillary isoelectric focusing followed by immunological detection. DJ-1 (PARK7) is considered as a biomarker candidate for Parkinson's disease and may potentially support the differentiation of clinical subtypes of the disease. The new method allows for separation and subsequent relative quantitative comparison of different isoforms of DJ-1 in biological samples. The assay was successfully applied to the analysis of DJ-1 isoform patterns in brains from mice subjected to normal or high-fat diet and revealed statistically significant group differences. Furthermore, in a pooled and concentrated sample of human cerebrospinal fluid that was depleted of albumin and immunoglobulin G, four different charge variants of DJ-1 could be detected. Taken together, the capillary isoelectric focusing immunoassay for DJ-1 represents a promising tool that may ultimately serve in clinical biomarker studies.
Asunto(s)
Química Encefálica , Péptidos y Proteínas de Señalización Intracelular/líquido cefalorraquídeo , Focalización Isoeléctrica/métodos , Proteínas Oncogénicas/análisis , Proteínas Oncogénicas/líquido cefalorraquídeo , Enfermedad de Parkinson/líquido cefalorraquídeo , Animales , Western Blotting , Encéfalo/metabolismo , Dieta Alta en Grasa , Humanos , Inmunoensayo/métodos , Masculino , Ratones , Ratones Endogámicos C57BL , Proteínas Oncogénicas/metabolismo , Peroxirredoxinas , Proteína Desglicasa DJ-1 , Isoformas de Proteínas/análisis , Isoformas de Proteínas/líquido cefalorraquídeoRESUMEN
INTRODUCTION: Here, we present a stem-cell based study on the de-novo generation of beta-III-tubulin-positive neurons after treatment with the classic antipsychotic drug haloperidol or after treatment with the second-generation antipsychotic (SGA) ziprasidone. METHODS: Adult neural stem cells (ANSC) dissociated from the adult mouse hippocampus were expanded in cell culture with basic fibroblast growth factor (bFGF) and epidermal growth factor (EGF). ANSC differentiated upon withdrawal of EGF and bFGF. RESULTS AND DISCUSSION: Ziprasidone generated significantly more beta-III-tubulin-positive neurons than haloperidol during the differentiation of adult neural stem cells isolated from murine hippocampus (ANSC). We assume that this net increase in neurogenesis by ziprasidone relies on this drug's 5-HT1A receptor affinity, which is not present in the haloperidol molecule, since the inactivation by WAY100621 impeded this process. These data could possibly suggest a clinical relevance for studying antipsychotic drugs in the stem cell paradigm employed in this study.
Asunto(s)
Antipsicóticos/farmacología , Haloperidol/farmacología , Hipocampo/citología , Células-Madre Neurales/efectos de los fármacos , Neurogénesis/efectos de los fármacos , Piperazinas/farmacología , Tiazoles/farmacología , Animales , Diferenciación Celular/efectos de los fármacos , Movimiento Celular/efectos de los fármacos , Células Cultivadas , Factor de Crecimiento Epidérmico/farmacología , Factor 2 de Crecimiento de Fibroblastos/farmacología , Hipocampo/efectos de los fármacos , Inmunohistoquímica , Ratones , Ratones Endogámicos C57BL , Receptor de Serotonina 5-HT1A/efectos de los fármacos , Antagonistas de la Serotonina/farmacología , Tubulina (Proteína)/biosíntesisRESUMEN
In light of the dramatically increasing prevalence of Alzheimer's disease (AD) to be expected in the future, the development of novel therapeutics, improved differential and early diagnostics, and means for the identification of individuals at risk are urgently needed. At present, instruments for a reliable differential diagnosis in clinical dementia, mild cognitive impairment, or prodromal stages have direct practical implications for differentiating secondary dementias from neurodegenerative conditions and for treatment decisions. It may also be reasonable to enforce the incorporation of biomarkers into clinical studies as surrogate outcome parameters and as an attempt to optimize recruitment criteria. Recently, revised research criteria increasingly rely on the interpretation of biomarker patterns, including neuroimaging and CSF-based neurochemical dementia diagnosis (NDD) in supporting the clinical diagnosis. Here, we review the performance of current core CSF biomarkers (Aß(42) peptide, total tau protein and phosphorylated tau species) and try to define objectives for prospective markers, also considering blood-based tests, which would increase the acceptance and wide application of NDD. Moreover, we evaluate the role and the limitations of genotyping in the predictive diagnosis of AD.
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Enfermedad de Alzheimer/diagnóstico , Biomarcadores/líquido cefalorraquídeo , Demencia/diagnóstico , Diagnóstico Diferencial , Enfermedad de Alzheimer/líquido cefalorraquídeo , Enfermedad de Alzheimer/complicaciones , Enfermedad de Alzheimer/genética , Péptidos beta-Amiloides/líquido cefalorraquídeo , Apolipoproteínas E/genética , Demencia/líquido cefalorraquídeo , Demencia/etiología , Demencia/genética , Predisposición Genética a la Enfermedad , Humanos , Enfermedades Neurodegenerativas/complicaciones , Neuroimagen , Fragmentos de Péptidos/líquido cefalorraquídeo , Valor Predictivo de las Pruebas , Proteínas tau/líquido cefalorraquídeoRESUMEN
Even if more extensively investigated in affective disorders, the serotonergic system is likely to be also implicated in modulating the pathogenesis of schizophrenia, where it closely interacts with the dopaminergic and glutamatergic system. To substantiate this notion, we studied the intensity and dynamics of cellular Ca(2+) responses to serotonin (5-hydoxytryptamine, 5-HT) in peripheral lymphocytes taken from currently non-psychotic schizophrenic patients. To this aim, peripheral lymphocytes were freshly obtained from healthy controls and a naturalistic collective of patients with schizophrenia in remission. Intracellular Ca(2+) responses were recorded in real-time by ratiometric fluorometry after 5-HT or phythaemagglutinin (PHA) stimulation, which served as an internal reference for Ca(2+) responsivity to non-specific stimulation. The intracellular Ca(2+) peak early after applying the 5-HT trigger was significantly elevated in schizophrenic patients. No significant differences of Ca(2+) peak levels were seen in response to stimulation with the mitogenic agent PHA, although responses to 5-HT and PHA were positively correlated in individual patients or controls. In conclusion, the serotonergic response patterns in peripheral lymphocytes from schizophrenic patients seem to be elevated, if employing sensitive tools like determination of intracellular Ca(2+) responses. Our observations suggest that the participation of serotonergic neurotransmitter system in the pathogenesis of schizophrenia may deserve more interest, even if it should only act as a modulator on the main pathology in the dopaminergic and glutamatergic systems. We hope that this pilot study will prompt further studies with larger patient collectives to revisit this question.
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Calcio/metabolismo , Receptores de Serotonina/metabolismo , Esquizofrenia/metabolismo , Linfocitos T/metabolismo , Adulto , Estudios de Casos y Controles , Femenino , Humanos , Masculino , Persona de Mediana Edad , Proyectos Piloto , Serotonina/farmacología , Agonistas de Receptores de Serotonina/farmacología , Transducción de SeñalRESUMEN
Antagonists of the N-methyl-D-aspartate (NMDA)-type glutamate receptor induce psychosis in healthy individuals and exacerbate schizophrenia symptoms in patients. In this study we have produced an animal model of NMDA receptor hypofunction by chronically treating rats with low doses of the NMDA receptor antagonist MK-801. Subsequently, we performed an expression study and identified 20 genes showing altered expression in the brain of these rats compared with untreated animals. We then explored whether the human orthologs of these genes are associated with schizophrenia in the largest schizophrenia genome-wide association study published to date, and found evidence for association for 4 out of the 20 genes: SF3B1, FOXP1, DLG2 and VGLL4. Interestingly, three of these genes, FOXP1, SF3B1 and DLG2, have previously been implicated in neurodevelopmental disorders.
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Maleato de Dizocilpina/farmacología , Factores de Transcripción Forkhead/genética , Guanilato-Quinasas/genética , Hipocampo , Fosfoproteínas/genética , Proteínas Represoras/genética , Ribonucleoproteína Nuclear Pequeña U2/genética , Esquizofrenia , Factores de Transcripción/genética , Proteínas Supresoras de Tumor/genética , Adulto , Anciano , Animales , Estudios de Casos y Controles , Femenino , Estudio de Asociación del Genoma Completo , Hipocampo/metabolismo , Hipocampo/patología , Humanos , Masculino , Persona de Mediana Edad , Psicotrópicos/farmacología , Factores de Empalme de ARN , Ratas , Receptores de N-Metil-D-Aspartato/antagonistas & inhibidores , Esquizofrenia/genética , Esquizofrenia/patologíaRESUMEN
BACKGROUND AND PURPOSE: The vitamin K-dependent protein Z (PZ) has been shown to possess anticoagulant as well as procoagulant properties. Plasma levels of PZ show a broad interindividual variation, but it is unknown to which extent this variation is under genetic control. Recent clinical studies revealed contradictory results on the association of PZ plasma levels and the risk of ischemic stroke. METHODS: We performed a case-control study including 200 patients with cerebral ischemia aged < or =50 years and 199 control subjects from the same South German region. We investigated a possible association of 2 common single nucleotide mutations in the PZ gene with the risk of cerebral ischemia. Furthermore, enzyme-linked immunosorbent assay measurements were done in control subjects without vascular disease to detect a potential association of different genotypes with PZ plasma (antigen) levels. RESULTS: In patients, the frequency of the A allele of the intron F polymorphism G79A was significantly lower than in controls (15.7% versus 24.4%; odds ratio, 0.58; 95% CI, 0.39 to 0.86; P=0.007; adjusted for age, sex, and conventional risk factors). The G allele of the promoter polymorphism A-13G tended to be less common in patients (4.2% versus 7.0%; adjusted odds ratio, 0.56; 95% CI, 0.28 to 1.13; P=0.105). In 42 control subjects, the A allele of the intron F polymorphism was associated with lower PZ antigen levels (P=0.0032; Spearman correlation coefficient rs=-0.48). CONCLUSIONS: The A allele of an intron F polymorphism of the PZ gene appears to be a novel protective genetic marker for the risk of cerebral ischemia in young adults. In the context of juvenile stroke, high PZ plasma levels may represent a prothrombotic condition.
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Proteínas Sanguíneas/análisis , Proteínas Sanguíneas/genética , Isquemia Encefálica/genética , Polimorfismo de Nucleótido Simple , Adulto , Isquemia Encefálica/epidemiología , Estudios de Casos y Controles , Factor V/genética , Femenino , Frecuencia de los Genes , Variación Genética , Alemania/epidemiología , Humanos , Intrones/genética , Masculino , Proteínas de la Membrana , Prevalencia , Regiones Promotoras Genéticas , Protrombina/genética , Medición de Riesgo , Proteínas Supresoras de TumorRESUMEN
Treatment of human hepatoma cells (HepG2) with NO-donors for 24 h inhibited hypoxia-induced erythropoietin (EPO) gene activation. NO was found to increase the production of reactive oxygen species (ROS), the putative signaling molecules between a cellular O2-sensor and hypoxia inducible factor 1 (HIF-1). HIF-1 is the prime regulator of O2-dependent genes such as EPO. NO-treatment for more than 20 h reduced HIF-1-driven reporter gene activity. In contrast, immediately after the addition of NO, ROS levels in HepG2 cells decreased below control values for as long as 4 h. Corresponding to these lowered ROS-levels, HIF-1 reporter gene activity and EPO gene expression transiently increased but were reduced when ROS levels rose thereafter. Our findings of a bimodal effect of NO on ROS production shed new light on the involvement of ROS in the mechanism of O2-sensing and may explain earlier conflicting data about the effect of NO on O2-dependent gene expression.
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Carcinoma Hepatocelular/metabolismo , Óxido Nítrico/metabolismo , Oxígeno/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Factores de Transcripción , Acridinas/metabolismo , Anaerobiosis , Carcinoma Hepatocelular/enzimología , Carcinoma Hepatocelular/patología , Proteínas de Unión al ADN/genética , Eritropoyetina/análisis , Eritropoyetina/genética , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Genes Reporteros , Humanos , Peróxido de Hidrógeno/metabolismo , Factor 1 Inducible por Hipoxia , Subunidad alfa del Factor 1 Inducible por Hipoxia , NADH NADPH Oxidorreductasas/metabolismo , NADPH Oxidasas , Donantes de Óxido Nítrico/farmacología , Óxidos de Nitrógeno , Proteínas Nucleares/genética , Compuestos Onio/farmacología , Penicilamina/análogos & derivados , Penicilamina/farmacología , ARN Mensajero/genética , ARN Mensajero/metabolismo , Transducción de Señal , Espermina/análogos & derivados , Espermina/farmacología , Activación Transcripcional , Transfección , Células Tumorales CultivadasRESUMEN
For decades treatment of schizophrenia was restricted to drugs, which mainly target positive symptoms by interfering with the dopaminergic neurotransmission. Since a large body of experimental and clinical data implicate that schizophrenia may primarily be a consequence of an imbalance in the glutamatergic system, specifically the networks containing GABAergic interneurons (γ-amino butyric acid), new drugs modulating glutamatergic neurotransmission are being developed. Targeting this dysfunction may follow different strategies, including application of direct or indirect NMDA (N-methyl-D-aspartate) receptor agonists or drugs modulating the function of metabotropic glutamate receptors. Meanwhile, the first substances have proven to be effective in animal models of schizophrenia and now enter the stage of clinical trials. The most promising data have been obtained in studies employing agonists of the metabotropic glutamate receptor. A choice of these substances is presented in this review.
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Interneuronas/fisiología , Esquizofrenia/fisiopatología , Aminoácidos/fisiología , Animales , Antipsicóticos/farmacología , Antipsicóticos/uso terapéutico , Humanos , Receptores Dopaminérgicos/fisiología , Receptores de Glutamato/fisiología , Esquizofrenia/tratamiento farmacológicoAsunto(s)
Regulación de la Expresión Génica , Oxígeno/metabolismo , Especies Reactivas de Oxígeno/metabolismo , Animales , Línea Celular , Grupo Citocromo b/metabolismo , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Eritropoyetina/biosíntesis , Factor 1 Inducible por Hipoxia , Subunidad alfa del Factor 1 Inducible por Hipoxia , Ratones , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Transducción de Señal , Factores de Transcripción/genética , Factores de Transcripción/metabolismoRESUMEN
There is evidence for a strong genetic component in the etiology of schizophrenia, as demonstrated by family, twin and adoption studies suggesting a heritability of about 80%. There are several approaches in the search for genetic risk factors such as linkage or association studies. Additionally, much effort was done in refining the phenotype including neuropsychology, neurophysiology, imaging or the generation of animal models. Genes becoming associated with schizophrenia have to be tested for functionality including e.g. metabolomics, transcriptomics, proteomics, generation of transgenic mice, analysis of protein-protein interactions, allele-specific RNA expression analysis, analysis of neuronal and stem cell cultures, as well as post mortem studies and behavioral studies in rodents. This amount of data requires complex data analysis. A system's perspective can help in the analysis of the structural and functional complexity of the brain. New tools will be needed for a more complex and systemic view. The systems biology approach could be a pivotal tool in understanding of complex behavior and diseases in future.
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Predisposición Genética a la Enfermedad , Esquizofrenia , Biología de Sistemas , Animales , Humanos , Esquizofrenia/genética , Esquizofrenia/patología , Esquizofrenia/fisiopatologíaRESUMEN
The supplementation of creatine (Cr) has a marked neuroprotective effect in mouse models of neurodegenerative diseases. This has been assigned to the known bioenergetic, anti-apoptotic, anti-excitotoxic, and anti-oxidant properties of Cr. As aging and neurodegeneration share pathophysiological pathways, we investigated the effect of oral Cr supplementation on aging in 162 aged C57Bl/6J mice. Outcome variables included "healthy" life span, neurobehavioral phenotyping, as well as morphology, biochemistry, and expression profiling from brain. The median healthy life span of Cr-fed mice was 9% higher than in control mice, and they performed significantly better in neurobehavioral tests. In brains of Cr-treated mice, there was a trend towards a reduction of reactive oxygen species and significantly lower accumulation of the "aging pigment" lipofuscin. Expression profiling showed an upregulation of genes implicated in neuronal growth, neuroprotection, and learning. These data show that Cr improves health and longevity in mice. Cr may be a promising food supplement to promote healthy human aging.
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Conducta Animal/fisiología , Cognición/fisiología , Creatina/administración & dosificación , Suplementos Dietéticos , Estado de Salud , Tasa de Supervivencia , Animales , Conducta Animal/efectos de los fármacos , Cognición/efectos de los fármacos , Ratones , Ratones Endogámicos C57BL , Ratones Endogámicos , SobrevidaRESUMEN
In a primary study on proinflammatory genetic profiles in stroke, the authors found the E469K polymorphism of the intercellular adhesion molecule 1 (ICAM-1) highly represented in the subgroup with spontaneous cervical artery dissection (sCAD). They further investigated the same genetic variant in a second group of 65 patients with sCAD. An association between sCAD and EE genotype was confirmed (odds ratio 3.16; p < 0.01), indicating that a proinflammatory predisposition is a risk factor for sCAD.