RESUMO
BACKGROUND: Immunological mechanisms are suspected in sensory neuropathy (SN) occurring with systemic autoimmune diseases and in some idiopathic cases, but so far there are no antibodies (Abs) identifying these neuropathies. METHODS: In the search for such specific antibodies, serum samples were collected from 106 patients with SN of these 72 fulfilled the diagnosis criteria of sensory neuronopathy (SNN) and 211 control subjects including patients with sensorimotor neuropathies, other neurological diseases (ONDs), systemic autoimmune diseases and healthy blood donors. RESULTS: In the first step, a protein array with 8000 human proteins allowed identification of the intracellular domain of the fibroblast growth factor receptor 3 (FGFR3) as a target of Abs in 7/16 SNN and 0/30 controls. In the second step, an ELISA method was used to test the 317 patients and controls for anti-FGFR3 Abs. Abs were detected in 16/106 patients with SN and 1/211 controls (p<0.001). Among the 106 patients with SN, anti-FGFR3 Abs were found in 11/38 patients with autoimmune context, 5/46 with idiopathic neuropathy and 0/22 with neuropathy of other aetiology (p=0.006). The only control patient with anti-FGFR3 Abs had lupus and no recorded neuropathy. Sensitivity, specificity, and positive and negative predictive values of anti-FGFR3 Abs for a diagnosis of idiopathic or dysimmune SN were 19%, 99.6%, 94.1% and 77.3%, respectively. A cell-based assay confirmed serum reactivity against the intracellular domain of FGFR3. The neuropathy in patients with anti-FGF3 Abs was non-length dependent in 87% of patients and fulfilled the criteria of probable SNN in 82%. Trigeminal nerve involvement and pain were frequent features. CONCLUSIONS: A anti-FGFR3 Abs identify a subgroup of patients with SN in whom an underlying autoimmune disorder affecting sensory neurons in the dorsal root and trigeminal nerve ganglia is suspected.
Assuntos
Anticorpos/análise , Doenças do Sistema Nervoso Periférico/imunologia , Receptor Tipo 3 de Fator de Crescimento de Fibroblastos/imunologia , Adulto , Idoso , Especificidade de Anticorpos , Reações Cruzadas , Ensaio de Imunoadsorção Enzimática , Feminino , Células HEK293 , Humanos , Masculino , Pessoa de Meia-Idade , Proteínas do Tecido Nervoso/análise , Proteínas do Tecido Nervoso/imunologia , Doenças do Sistema Nervoso Periférico/diagnóstico , Valor Preditivo dos Testes , Estudos Retrospectivos , Células Receptoras Sensoriais/imunologiaRESUMO
Amyotrophic lateral sclerosis (ALS) is a progressive, lethal, degenerative disorder of motor neurons. The causes of most cases of ALS are as yet undefined. In a previous study, it was shown that N-methyl-D-aspartate (NMDA) and H(2)O(2) stimuli reduce neuronal survival in cortical neurons in culture (Boutahar et al., 2008). To identify variations in gene expression in response to these neurotoxins in transgenic vs. control cortical neurons cultures, both microarray and RT-PCR analysis were performed. High-density oligonucleotide microarrays showed changes in the expression of about 600 genes involved in protein degradation, neurotrophic factors pathway, cell cycle, inflammation, cytoskeleton, cell adhesion, transcription, or signalling. The most up-regulated genes following H(2)O(2) treatment were involved in cytoskeletal organization and axonal transport, such as ARAP2, KIF17, and DKK2, or in trophic factors pathways, such as insulin-like growth factor-binding protein 4 (IGFBP4), FGF17, and serpin2. The most down-regulated genes were involved in ion transport, such as TRPV1. After NMDA treatment, the most up-regulated genes were involved in protein degradation, such as ubiquitin-conjugating enzyme E2I and cathepsin H, and the most down-regulated genes were involved in ion transport, such as SCN7A. We conclude that these neurotoxins act through different transcriptional inductions, and these changes may reflect an adaptative cellular response to the cellular stress induced by the neurotoxins involved in ALS in the presence of mutant human SOD1.
Assuntos
Esclerose Lateral Amiotrófica/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Neurônios/metabolismo , Superóxido Dismutase/metabolismo , Análise de Variância , Animais , Sobrevivência Celular , Células Cultivadas , Córtex Cerebral/citologia , Córtex Cerebral/metabolismo , Perfilação da Expressão Gênica , Humanos , Peróxido de Hidrogênio/farmacologia , Camundongos , Camundongos Mutantes Neurológicos , Camundongos Transgênicos , N-Metilaspartato/farmacologia , Neurotoxinas/farmacologia , Análise de Sequência com Séries de Oligonucleotídeos , Oxidantes/farmacologia , Estresse Oxidativo , Estatísticas não Paramétricas , Superóxido Dismutase/genética , Superóxido Dismutase-1RESUMO
Neurotrophins protect neurons against glutamate and oxidative stress, but the underlying mechanism remains unclear. We investigated the neuroprotective role of the neurotrophin brain-derived neurotrophic factor (BDNF) in neuronal cultures subjected to NMDA or H(2)O(2) toxicity and analyzed the molecular mechanisms involved, particularly those related to regulation of cell cycle or endoplasmic reticulum (ER) stress. Preincubation with BDNF of cortical neuron cultures prevented NMDA- or H(2)O(2)-induced neuronal death as well as MAPK-ERK1/2 activation. Inhibition of phosphatidylinositol 3-kinase (PI3-K) abolished the protective effect of BDNF. NMDA and H(2)O(2) induced activation of cell cycle reentry regulators such as retinoblastoma (Rb) protein and E2F1 transcription factor. However, BDNF abolished the activation of both factors. NMDA-induced expression of chaperone encoding gene BIP was slightly inhibited by BDNF, but it did not affect expression of ER stress protein CHOP. Our results suggest that BDNF neuroprotection may be mediated through inhibition of Ras-MAPK pathway and cell cycle reentry during oxidative or excitotoxic stress responses. However, BDNF did not modify expression of ER stress signal induced by NMDA.
Assuntos
Fator Neurotrófico Derivado do Encéfalo/metabolismo , Ciclo Celular/fisiologia , Retículo Endoplasmático/metabolismo , Neurônios/fisiologia , Estresse Oxidativo/fisiologia , Estresse Fisiológico/fisiologia , Animais , Morte Celular/fisiologia , Córtex Cerebral/fisiologia , Peróxido de Hidrogênio/metabolismo , Sistema de Sinalização das MAP Quinases/fisiologia , Camundongos , N-Metilaspartato/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Fatores de TempoRESUMO
Oxidative stress and excitotoxicity are both involved in the pathogenesis of neuronal degenerative diseases like ALS. In order to compare their action, some key proteins involved in their respective signaling pathways, particularly ERK and p53, were analyzed in primary cultures of cortical neurons subjected to NMDA or H(2)O(2) treatment. Early ERK activation was detected after NMDA treatment and was maintained during 24 h, but not after H(2)O(2) treatment. Early p53 expression was also found after NMDA treatment but diminished later. On the other hand, it progressively increased from 6 h to 24 h after H(2)O(2) treatment. Blocking ERK1/2 activation with the upstream inhibitor U0126 inhibited NMDA-mediated p53 expression, suggesting that ERK1/2 signals drive the cells to apoptosis under these conditions. In order to identify the initial membrane target of these neurotoxins, PAK1 was analyzed. Early increase of PAK1 expression was measured after NMDA treatment and was still present after 24 h. Conversely increased PAK1 expression was only detected 24 h after H(2)O(2) treatment. In order to define the components through which NMDA or H(2)O(2) induce the final elements of these pathways, p21 and c-jun, we have performed a detailed functional analysis of c-jun and p21 promoters following plasmid transfection. Both p21 and c-jun were activated after NMDA treatment, but this activation was abolished after H(2)O(2) treatment. We conclude that NMDA induces an early effect that involves activation of p53, ERK, PAK1, p21 and c-jun. On the other hand, H(2)O(2) induces long-term p53 expression, late expression of PAK1 without activation of p21 promoter. The timing differences of the action of these neurotoxins may explain why the presence of both compounds is needed to induce neuronal death.