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Olfactory dysfunction and atrophy of olfactory brain regions are observed early in mild cognitive impairment and Alzheimer disease. Despite substantial evidence showing neuroprotective effects in MCI/AD with treatment of docosahexaenoic acid (DHA), an omega-3 fatty acid, few studies have assessed DHA and its effects on the olfactory system deficits. We therefore performed structural (MRI), functional (olfactory behavior, novel object recognition), and molecular (markers of apoptosis and inflammation) assessments of APOE4 and wild-type mice ± DHA treatment at 3, 6, and 12 months of age. Our results demonstrate that APOE4 mice treated with the control diet show recognition memory deficits, abnormal olfactory habituation, and discrimination abilities and an increase in IBA-1 immunoreactivity in the olfactory bulb. These phenotypes were not present in APOE4 mice treated with a DHA diet. Alterations in some brain regions' weights and/or volumes were observed in the APOPE4 mice and may be due to caspase activation and/or neuroinflammatory events. These results suggest that the consumption of a diet rich in DHA may provide some benefit to E4 carriers but may not alleviate all symptoms.
Assuntos
Doença de Alzheimer , Disfunção Cognitiva , Camundongos , Animais , Camundongos Transgênicos , Ácidos Docosa-Hexaenoicos/farmacologia , Ácidos Docosa-Hexaenoicos/uso terapêutico , Apolipoproteína E4/genética , Apolipoproteína E4/metabolismo , Encéfalo/metabolismo , Disfunção Cognitiva/complicações , Disfunção Cognitiva/tratamento farmacológico , Doença de Alzheimer/genética , CogniçãoRESUMO
Olfactory dysfunction is a common symptom in neurodegenerative disorders and is regarded as a potential early predictor of impending cognitive decline. This study was undertaken in order to determine if olfactory dysfunction observed in the elderly is due to a general loss of smell or the inability to detect specific odours, and if misidentification of odours correlates with cognitive scores. Seniors for the Olfactory Response and Cognition in Aging (ORCA) sub-study were recruited from the Quebec Nutrition and Successful Aging (NuAge) cohort. The University of Pennsylvania smell identification test (UPSIT) was performed to measure olfactory function and the telephone Mini Mental State Examination (t-MMSE) and the French version of the Telephone Interview for Cognitive Status Modified (F-TICS-m) for cognitive status. The results demonstrate that seniors exhibit specific olfactory loss and had severe difficulty in particular in identifying lemon, pizza, fruit punch, cheddar cheese and lime. Furthermore, there was a significant difference in the ability to detect certain odours between the sexes. Results also showed that misidentification of certain scents was associated with cognitive scores, and when the sexes were assessed separately sex-specific misidentification of cognitive-associated odours was observed. The relationship between the cognitive scores and scent misidentification suggests that impending cognitive decline may be highlighted by the inability to smell specific odours. Our study provides additional support for the testing of olfactory function in the elderly and suggests that loss of smell for particular scents may become a useful diagnostic tool.
Assuntos
Disfunção Cognitiva , Transtornos do Olfato , Masculino , Feminino , Humanos , Idoso , Olfato , Transtornos do Olfato/diagnóstico , Anosmia , Envelhecimento , Disfunção Cognitiva/diagnósticoRESUMO
BACKGROUND: In many neurological disorders, including Alzheimer disease, early olfactory dysfunction is observed. OBJECTIVE: In order to determine if deficits in olfactory memory are present in the elderly and if olfactory dysfunction correlates with cognitive impairment in the aging population, olfactory testing has been done on seniors from the NuAge cohort accepting to participate in the Olfactory Response Cognition and Aging (ORCA) secondary sub-study. The t-Mini Mental Statement Examination and the Telephone Interview for Cognitive Status tests were done to assess cognition levels. RESULTS: Overall, 94% of the ORCA cohort displayed olfactory dysfunction. Deficits in olfactory memory were also present. A correlation was observed between olfactory function and cognitive test scores. Moreover, in women who smoked, there was an association between olfactory memory and cognitive scores. CONCLUSION: Our results suggest that olfactory dysfunction may predict impending cognitive decline and highlights the need for olfactory training in seniors to improve olfaction and overall well-being.
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Olfactory dysfunction is observed in several neurological disorders including Mild Cognitive Impairment (MCI) and Alzheimer disease (AD). These deficits occur early and correlate with global cognitive performance, depression and degeneration of olfactory regions in the brain. Despite extensive human studies, there has been little characterization of the olfactory system in models of AD. In order to determine if olfactory structural and/or molecular phenotypes are observed in a model expressing a genetic risk factor for AD, we assessed the olfactory bulb (OB) in APOE4 transgenic mice. A significant decrease in OB weight was observed at 12 months of age in APOE4 mice concurrent with inflammation and decreased NeuN expression. In order to determine if a diet rich in omega-3s may alleviate the olfactory system phenotypes observed, we assessed WT and APOE4 mice on a docosahexaenoic acid (DHA) diet. APOE4 mice on a DHA diet did not present with atrophy of the OB, and the alterations in NeuN and IBA-1 expression were alleviated. Furthermore, alterations in caspase mRNA and protein expression in the APOE4 OB were not observed with a DHA diet. Similar to the human AD condition, OB atrophy is an early phenotype in the APOE4 mice and concurrent with inflammation. These data support a link between the structural olfactory brain region atrophy and the olfactory dysfunction observed in AD and suggest that inflammation and cell death pathways may contribute to the olfactory deficits observed. Furthermore, the results suggest that diets enriched in DHA may provide benefit to APOE4 allele carriers.
Assuntos
Doença de Alzheimer , Apolipoproteína E4 , Ácidos Docosa-Hexaenoicos/fisiologia , Transtornos do Olfato/dietoterapia , Bulbo Olfatório , Doença de Alzheimer/complicações , Doença de Alzheimer/genética , Animais , Apolipoproteína E4/genética , Atrofia , Dieta , Modelos Animais de Doenças , Camundongos , Camundongos Transgênicos , Transtornos do Olfato/etiologia , Transtornos do Olfato/genética , Bulbo Olfatório/crescimento & desenvolvimento , Bulbo Olfatório/metabolismo , Bulbo Olfatório/patologiaRESUMO
OBJECTIVE: Alzheimer disease (AD) is a chronic neurodegenerative disorder that affects millions of individuals worldwide. Symptoms include memory dysfunction and deficits in attention, planning, language, and overall cognitive function. Olfactory dysfunction is a common symptom of AD and evidence supports that it is an early marker. Furthermore, olfactory bulb and entorhinal cortex atrophy are well described in AD. However, in AD, no studies have assessed the olfactory cortex as a whole and if sex effects are observed. METHODS: Magnetic Resonance Imaging was used to scan 39 participants with an average age of 72 years and included men and women. AAL Single-Subject Atlas (implemented in PNEURO tool - PMOD 3.8) was used to determine the volume of the olfactory cortex and the hippocampus. Olfactory cortex volume was lower in both men and women AD cases compared with controls. This decrease was more apparent in the left olfactory cortex and was influenced by age. As expected, hippocampal volume was also significantly reduced in AD. However, this was only observed in the male cohort. A significant correlation was observed between levels of education and hippocampal volume in controls that were not detected in the AD participants. Asymmetry was observed in the olfactory cortex volume when comparing left and right volumes in both the control and AD participants, which was not observed in the hippocampus. RESULTS: These data highlight the importance of the role of olfactory cortical atrophy in the pathogenesis of AD and the interplay between the olfactory deficits and degeneration of olfactory regions in the brain.
Assuntos
Doença de Alzheimer/patologia , Atrofia/patologia , Processamento de Imagem Assistida por Computador , Córtex Olfatório/patologia , Idoso , Encéfalo/patologia , Disfunção Cognitiva/fisiopatologia , Estudos de Coortes , Feminino , Hipocampo/patologia , Humanos , Imageamento por Ressonância Magnética , Masculino , Fatores SexuaisRESUMO
Excitotoxicity, due to overstimulation of N-methyl D-aspartate receptors (NMDARs), has a pivotal role in many neurological disorders. However, NMDAR antagonists often cause side effects, and identifying more druggable therapeutic targets for NMDAR excitotoxicity is an important goal. Activation of caspases is a downstream effect of excitotoxicity that may be critically involved in NMDAR-mediated cell death. Caspase-6 (casp6) in particular has been shown to play a key role in the pathogenesis of stroke, Huntington disease, and Alzheimer disease. Using N-methyl D-aspartate (NMDA)-induced excitotoxic injuries of primary rat neurons, we demonstrate that there is an early increase in caspase profiles after an excitotoxic event at the level of mRNA, protein, and activity. Casp6 is elevated and activated first, followed by caspase-8 and caspase-3. Similarly, known casp6 substrates huntingtin, as well as novel casp6 substrates serine/threonine kinase 3 and death domain-associated protein, are cleaved in similar temporal patterns post NMDA. On the basis of these data, we propose that casp6 may be an initiator caspase in apoptotic cascades leading to neuronal death after an excitotoxic event and suggest casp6 as a potential therapeutic target for neurological disorders where NMDAR-mediated excitotoxicity has been shown to play a role.
Assuntos
Caspase 6/metabolismo , Receptores de N-Metil-D-Aspartato/metabolismo , Animais , Córtex Cerebral/enzimologia , Córtex Cerebral/metabolismo , Ativação Enzimática , Neurônios/enzimologia , Neurônios/metabolismo , Neurotoxinas/farmacologia , Cultura Primária de Células , Ratos Sprague-DawleyRESUMO
Caspases and their substrates are key mediators of apoptosis and strongly implicated in various physiological processes. As the serine/threonine kinase family is involved in apoptosis and serine/threonine kinase 3 (STK3) is a recently identified caspase-6 substrate, we assessed the expression and cleavage of STK3 in murine peripheral organs and brain regions during the aging process. We also assessed caspase-3, -6, -7, and -8 expression and activity in order to delineate potential mechanism(s) underlying the generation of the STK3 fragments observed and their relation to the apoptotic pathway. We demonstrate for the first time the cleavage of STK3 by caspase-7 and show that STK3 protein levels globally increase throughout the organism with age. In contrast, caspase-3, -6, -7, and -8 expression and activity vary significantly among the different organs analyzed suggesting differential effects of aging on the apoptotic mechanism and/or nonapoptotic functions of caspases throughout the organism. These results further our understanding of the role of caspases and their substrates in the normal aging process and highlight a potential role for STK3 in neurodegeneration.
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Envelhecimento/genética , Envelhecimento/metabolismo , Apoptose/genética , Caspases/genética , Caspases/metabolismo , Expressão Gênica/genética , Especificidade de Órgãos/genética , Proteínas Serina-Treonina Quinases/metabolismo , Proteólise , Animais , Encéfalo/metabolismo , Caspases/fisiologia , Masculino , Camundongos Endogâmicos C57BL , Doenças Neurodegenerativas/genética , Serina-Treonina Quinase 3RESUMO
Death-associated protein 6 (DAXX) is a ubiquitous protein implicated in various cellular processes such as apoptosis, tumorigenesis, development and transcription. The role of DAXX is however ambiguous and many contradictory results regarding its function in apoptosis upon various cellular stresses are described in the literature. In order to have a better understanding of the role of DAXX throughout the entire organism under physiological stress conditions, we have characterized the mRNA levels, protein expression and the proteolytic processing of DAXX in the normal aging process in peripheral organs and brain regions in C57BL/6 male mice. Overall, Daxx mRNA expression decreases with aging in the liver, kidney, heart, cortex and cerebellum. In contrast, an increase is observed in the striatum. The protein expression of DAXX and of its proteolytic fragments increases with aging in the kidney, heart and cortex. In liver and spleen, no changes are observed while in the striatum and cerebellum, certain forms increase and others decrease with age, suggesting that the functions of DAXX may be cell type dependent. This study provides important details regarding the expression and post-translational modifications of DAXX in aging in the entire organism and provides reference data for the deregulation observed in age-associated diseases.
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Envelhecimento/metabolismo , Encéfalo/metabolismo , Proteínas de Transporte/metabolismo , Regulação da Expressão Gênica no Desenvolvimento/fisiologia , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Proteínas Nucleares/metabolismo , Vísceras/metabolismo , Animais , Proteínas Correpressoras , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Chaperonas Moleculares , Especificidade de Órgãos/fisiologiaRESUMO
Caspase-6 (CASP6) has emerged as an important player in Huntington disease (HD), Alzheimer disease (AD) and cerebral ischemia, where it is activated early in the disease process. CASP6 also plays a key role in axonal degeneration, further underscoring the importance of this protease in neurodegenerative pathways. As a protein's function is modulated by its protein-protein interactions, we performed a high-throughput yeast-2-hybrid (Y2H) screen against â¼17,000 human proteins to gain further insight into the function of CASP6. We identified a high-confidence list of 87 potential CASP6 interactors. From this list, 61% are predicted to contain a CASP6 recognition site. Of nine candidate substrates assessed, six are cleaved by CASP6. Proteins that did not contain a predicted CASP6 recognition site were assessed using a LUMIER assay approach, and 51% were further validated as interactors by this method. Of note, 54% of the high-confidence interactors identified show alterations in human HD brain at the mRNA level, and there is a significant enrichment for previously validated huntingtin (HTT) interactors. One protein of interest, STK3, a pro-apoptotic kinase, was validated biochemically to be a CASP6 substrate. Furthermore, our results demonstrate that in striatal cells expressing mutant huntingtin (mHTT), an increase in full length and fragment levels of STK3 are observed. We further show that caspase-3 is not essential for the endogenous cleavage of STK3. Characterization of the interaction network provides important new information regarding key pathways of interactors of CASP6 and highlights potential novel therapeutic targets for HD, AD and cerebral ischemia.
Assuntos
Caspase 6/metabolismo , Doença de Huntington/metabolismo , Doença de Huntington/patologia , Mapas de Interação de Proteínas , Proteínas Serina-Treonina Quinases/metabolismo , Sítios de Ligação , Linhagem Celular , Regulação da Expressão Gênica , Humanos , Proteína Huntingtina/genética , Modelos Biológicos , Processamento de Proteína Pós-Traducional , Serina-Treonina Quinase 3 , Técnicas do Sistema de Duplo-HíbridoRESUMO
There is evidence of substantial benefit of cardiac rehabilitation (CR) for patients with low exercise capacity at admission. Nevertheless, some patients are not able to perform an initial exercise stress test (EST). We aimed to describe this group using data of 1094 consecutive patients after a cardiac event (71±7 years, 78% men) enrolled in nine centres for inpatient CR. We analysed sociodemographic and clinical variables (e.g. cardiovascular risk factors, comorbidities, complications at admission), amount of therapy (e.g. exercise training, nursing care) and the results of the initial and the final 6-min walking test (6MWT) with respect to the application of an EST. Fifteen per cent of patients did not undergo an EST (non-EST group). In multivariable analysis, the probability of obtaining an EST was higher for men [odds ratio (OR) 1.89, P=0.01], a 6MWT (per 10 m, OR 1.07, P<0.01) and lower for patients with diabetes mellitus (OR 0.48, P<0.01), NYHA-class III/IV (OR 0.27, P<0.01), osteoarthritis (OR 0.39, P<0.01) and a longer hospital stay (per 5 days, OR 0.87, P=0.02). The non-EST group received fewer therapy units of exercise training, but more units of nursing care and physiotherapy than the EST group. However, there were no significant differences between both groups in the increase of the 6MWT during CR (123 vs. 108 m, P=0.122). The present study confirms the feasibility of an EST at the start of CR as an indicator of disease severity. Nevertheless, patients without EST benefit from CR even if exercising less. Thus, there is a justified need for individualized, comprehensive and interdisciplinary CR.
Assuntos
Síndrome Coronariana Aguda/reabilitação , Assistência Integral à Saúde , Avaliação da Deficiência , Teste de Esforço , Doenças das Valvas Cardíacas/reabilitação , Infarto do Miocárdio/reabilitação , Síndrome Coronariana Aguda/classificação , Síndrome Coronariana Aguda/diagnóstico , Idoso , Idoso de 80 Anos ou mais , Ponte de Artéria Coronária/reabilitação , Tolerância ao Exercício , Feminino , Doenças das Valvas Cardíacas/classificação , Doenças das Valvas Cardíacas/diagnóstico , Implante de Prótese de Valva Cardíaca/reabilitação , Humanos , Masculino , Pessoa de Meia-Idade , Infarto do Miocárdio/classificação , Infarto do Miocárdio/diagnóstico , Intervenção Coronária Percutânea/reabilitação , CaminhadaRESUMO
BACKGROUND: Arrhythmias associated with QT prolongation on the ECG often lead to sudden unexpected death in epilepsy. The mechanism causing a prolongation of the QT interval during epilepsy remains unknown. Based on observations showing an upregulation of neuronal sodium channels in the brain during epilepsy, we tested the hypothesis that a similar phenomenon occurs in the heart and contributes to QT prolongation by altering cardiac sodium current properties (INa). METHODS AND RESULTS: We used the patch clamp technique to assess the effects of epilepsy on the cardiac action potential and INa in rat ventricular myocytes. Consistent with QT prolongation, epileptic rats had longer ventricular action potential durations attributable to a sustained component of INa (INaL). The increase in INaL was because of a larger contribution of neuronal Na channels characterized by their high sensitivity to tetrodotoxin. As in the brain, epilepsy was associated with an enhanced expression of the neuronal isoform NaV1.1 in cardiomyocyte. Epilepsy was also associated with a lower INa activation threshold resulting in increased cell excitability. CONCLUSIONS: This is the first study correlating increased expression of neuronal sodium channels within the heart to epilepsy-related cardiac arrhythmias. This represents a new paradigm in our understanding of cardiac complications related to epilepsy.
Assuntos
Potenciais de Ação/fisiologia , Arritmias Cardíacas/genética , DNA/genética , Morte Súbita/etiologia , Epilepsia/metabolismo , Regulação da Expressão Gênica , Miócitos Cardíacos/metabolismo , Canal de Sódio Disparado por Voltagem NAV1.5/genética , Animais , Arritmias Cardíacas/complicações , Arritmias Cardíacas/metabolismo , Western Blotting , Epilepsia/complicações , Epilepsia/mortalidade , Masculino , Canal de Sódio Disparado por Voltagem NAV1.5/biossíntese , Técnicas de Patch-Clamp , Ratos , Reação em Cadeia da Polimerase em Tempo RealRESUMO
Assemblies of huntingtin (HTT) fragments with expanded polyglutamine (polyQ) tracts are a pathological hallmark of Huntington's disease (HD). The molecular mechanisms by which these structures are formed and cause neuronal dysfunction and toxicity are poorly understood. Here, we utilized available gene expression data sets of selected brain regions of HD patients and controls for systematic interaction network filtering in order to predict disease-relevant, brain region-specific HTT interaction partners. Starting from a large protein-protein interaction (PPI) data set, a step-by-step computational filtering strategy facilitated the generation of a focused PPI network that directly or indirectly connects 13 proteins potentially dysregulated in HD with the disease protein HTT. This network enabled the discovery of the neuron-specific protein CRMP1 that targets aggregation-prone, N-terminal HTT fragments and suppresses their spontaneous self-assembly into proteotoxic structures in various models of HD. Experimental validation indicates that our network filtering procedure provides a simple but powerful strategy to identify disease-relevant proteins that influence misfolding and aggregation of polyQ disease proteins.
Assuntos
Algoritmos , Proteínas do Tecido Nervoso/metabolismo , Agregação Patológica de Proteínas/metabolismo , Dobramento de Proteína , Sequência de Aminoácidos , Animais , Encéfalo/metabolismo , Encéfalo/patologia , Linhagem Celular Tumoral , Drosophila/genética , Drosophila/metabolismo , Proteína Huntingtina , Dados de Sequência Molecular , Proteínas do Tecido Nervoso/química , Proteínas do Tecido Nervoso/genética , Células PC12 , Ligação Proteica , RatosRESUMO
In order to further understand age-related physiological changes and to have in depth reference values for C57BL/6 mice, we undertook a study to assess the effects of aging on peripheral organ weights, and brain region weights in wild type C57BL/6 male mice. Peripheral organs, body and brain region weights were collected from young (3-4 months), mid (12 months), old (23-28 months) and very old (>30 months) mice. Significant increases are observed with aging in body, liver, heart, kidney and spleen organ weights. A decrease in organ weight is observed with aging in liver and kidney only in the very old mice. In contrast, testes weight decreases with age. Within the brain, hippocampi, striata and olfactory bulbs weight decreases with age. These data further our knowledge of the anatomical and biological changes that occur with aging and provide reference values for physiological-based pharmacokinetic studies in C57BL/6 mice.
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Envelhecimento , Peso Corporal/fisiologia , Encéfalo/anatomia & histologia , Animais , Coração/anatomia & histologia , Rim/anatomia & histologia , Fígado/anatomia & histologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Tamanho do Órgão , Valores de Referência , Baço/anatomia & histologia , Testículo/anatomia & histologiaRESUMO
Huntington Disease (HD) is a progressive neurodegenerative disease caused by an elongated CAG repeat in the huntingtin (HTT) gene that encodes a polyglutamine tract in the HTT protein. Proteolysis of the mutant HTT protein (mHTT) has been detected in human and murine HD brains and is implicated in the pathogenesis of HD. Of particular importance is the site at amino acid (aa) 586 that contains a caspase-6 (Casp6) recognition motif. Activation of Casp6 occurs presymptomatically in human HD patients and the inhibition of mHTT proteolysis at aa586 in the YAC128 mouse model results in the full rescue of HD-like phenotypes. Surprisingly, Casp6 ablation in two different HD mouse models did not completely prevent the generation of this fragment, and therapeutic benefits were limited, questioning the role of Casp6 in the disease. We have evaluated the impact of the loss of Casp6 in the YAC128 mouse model of HD. Levels of the mHTT-586 fragment are reduced but not absent in the absence of Casp6 and we identify caspase 8 as an alternate enzyme that can generate this fragment. In vivo, the ablation of Casp6 results in a partial rescue of body weight gain, normalized IGF-1 levels, a reversal of the depression-like phenotype and decreased HTT levels. In the YAC128/Casp6-/- striatum there is a concomitant reduction in p62 levels, a marker of autophagic activity, suggesting increased autophagic clearance. These results implicate the HTT-586 fragment as a key contributor to certain features of HD, irrespective of the enzyme involved in its generation.
Assuntos
Caspase 6/metabolismo , Doença de Huntington/enzimologia , Proteínas da Membrana Plasmática de Transporte de Serotonina/metabolismo , Animais , Peso Corporal , Fator Neurotrófico Derivado do Encéfalo/metabolismo , Caspase 6/genética , Corpo Estriado/metabolismo , Depressão/metabolismo , Modelos Animais de Doenças , Fator de Crescimento Insulin-Like I/metabolismo , Camundongos , Camundongos Transgênicos , Atividade MotoraRESUMO
HIP14 is the most highly conserved of 23 human palmitoyl acyltransferases (PATs) that catalyze the post-translational addition of palmitate to proteins, including huntingtin (HTT). HIP14 is dysfunctional in the presence of mutant HTT (mHTT), the causative gene for Huntington disease (HD), and we hypothesize that reduced palmitoylation of HTT and other HIP14 substrates contributes to the pathogenesis of the disease. Here we describe the yeast two-hybrid (Y2H) interactors of HIP14 in the first comprehensive study of interactors of a mammalian PAT. Unexpectedly, we discovered a highly significant overlap between HIP14 interactors and 370 published interactors of HTT, 4-fold greater than for control proteins (P = 8 × 10(-5)). Nearly half of the 36 shared interactors are already implicated in HD, supporting a direct link between HIP14 and the disease. The HIP14 Y2H interaction set is significantly enriched for palmitoylated proteins that are candidate substrates. We confirmed that three of them, GPM6A, and the Sprouty domain-containing proteins SPRED1 and SPRED3, are indeed palmitoylated by HIP14; the first enzyme known to palmitoylate these proteins. These novel substrates functions might be affected by reduced palmitoylation in HD. We also show that the vesicular cargo adapter optineurin, an established HTT-binding protein, co-immunoprecipitates with HIP14 but is not palmitoylated. mHTT leads to mislocalization of optineurin and aberrant cargo trafficking. Therefore, it is possible that optineurin regulates trafficking of HIP14 to its substrates. Taken together, our data raise the possibility that defective palmitoylation by HIP14 might be an important mechanism that contributes to the pathogenesis of HD.
Assuntos
Aciltransferases/genética , Proteínas Adaptadoras de Transdução de Sinal/genética , Doença de Huntington/genética , Proteínas do Tecido Nervoso/genética , Processamento de Proteína Pós-Traducional , Aciltransferases/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Animais , Células COS , Proteínas de Ciclo Celular , Chlorocebus aethiops , Redes Reguladoras de Genes , Células HEK293 , Humanos , Proteína Huntingtina , Doença de Huntington/metabolismo , Doença de Huntington/patologia , Peptídeos e Proteínas de Sinalização Intracelular/genética , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Lipoilação , Glicoproteínas de Membrana/genética , Glicoproteínas de Membrana/metabolismo , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Proteínas de Membrana Transportadoras , Anotação de Sequência Molecular , Proteínas do Tecido Nervoso/metabolismo , Ligação Proteica , Mapeamento de Interação de Proteínas , Proteínas Repressoras/genética , Proteínas Repressoras/metabolismo , Transdução de Sinais , Fator de Transcrição TFIIIA/genética , Fator de Transcrição TFIIIA/metabolismo , Técnicas do Sistema de Duplo-HíbridoRESUMO
Huntington's disease is caused by an expanded polyglutamine repeat in the huntingtin protein (HTT), but the pathophysiological sequence of events that trigger synaptic failure and neuronal loss are not fully understood. Alterations in N-methyl-D-aspartate (NMDA)-type glutamate receptors (NMDARs) have been implicated. Yet, it remains unclear how the HTT mutation affects NMDAR function, and direct evidence for a causative role is missing. Here we show that mutant HTT redirects an intracellular store of juvenile NMDARs containing GluN3A subunits to the surface of striatal neurons by sequestering and disrupting the subcellular localization of the endocytic adaptor PACSIN1, which is specific for GluN3A. Overexpressing GluN3A in wild-type mouse striatum mimicked the synapse loss observed in Huntington's disease mouse models, whereas genetic deletion of GluN3A prevented synapse degeneration, ameliorated motor and cognitive decline and reduced striatal atrophy and neuronal loss in the YAC128 Huntington's disease mouse model. Furthermore, GluN3A deletion corrected the abnormally enhanced NMDAR currents, which have been linked to cell death in Huntington's disease and other neurodegenerative conditions. Our findings reveal an early pathogenic role of GluN3A dysregulation in Huntington's disease and suggest that therapies targeting GluN3A or pathogenic HTT-PACSIN1 interactions might prevent or delay disease progression.
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Comportamento Animal , Doença de Huntington/metabolismo , Doença de Huntington/patologia , Receptores de N-Metil-D-Aspartato/metabolismo , Sinapses/metabolismo , Proteínas Adaptadoras de Transdução de Sinal , Animais , Proteínas de Transporte/metabolismo , Morte Celular/efeitos dos fármacos , Proteínas do Citoesqueleto , Modelos Animais de Doenças , Deleção de Genes , Células HEK293 , Humanos , Doença de Huntington/fisiopatologia , Imunoprecipitação , Peptídeos e Proteínas de Sinalização Intracelular , Camundongos , Atividade Motora/efeitos dos fármacos , Proteínas Mutantes/química , Proteínas Mutantes/metabolismo , Proteínas Mutantes/toxicidade , Neostriado/metabolismo , Neostriado/patologia , Proteínas do Tecido Nervoso/química , Proteínas do Tecido Nervoso/metabolismo , Neurônios/efeitos dos fármacos , Neurônios/metabolismo , Neurônios/patologia , Neuropeptídeos/metabolismo , Fosfoproteínas/metabolismo , Ligação Proteica/efeitos dos fármacos , Estrutura Quaternária de Proteína , Teste de Desempenho do Rota-Rod , Sinapses/efeitos dos fármacos , Sinapses/ultraestruturaRESUMO
Huntington disease (HD), a neurodegenerative disorder caused by an expanded CAG repeat in the HTT gene, remains without a treatment to modify the course of the illness. Lithium, a drug widely used for the treatment of bipolar disorder, has been shown to exert neuroprotective effects in a number of models of neurological disease but may have various toxic effects at conventional therapeutic doses. We examined whether NP03, a novel low-dose lithium microemulsion, would improve the disease phenotypes in the YAC128 mouse model of HD. We demonstrate that NP03 improves motor function, ameliorates the neuropathological deficits in striatal volume, neuronal counts, and DARPP-32 expression, and partially rescues testicular atrophy in YAC128 mice. These positive effects were accompanied by improvements in multiple biochemical endpoints associated with the pathogenesis of HD, including normalization of caspase-6 activation and amelioration of deficits in BDNF levels, and with no lithium-related toxicity. Our findings demonstrate that NP03 ameliorates the motor and neuropathological phenotypes in the YAC128 mouse model of HD, and represents a potential therapeutic approach for HD.
Assuntos
Encéfalo/efeitos dos fármacos , Doença de Huntington/tratamento farmacológico , Lítio/administração & dosagem , Fármacos Neuroprotetores/administração & dosagem , Animais , Encéfalo/patologia , Modelos Animais de Doenças , Feminino , Humanos , Doença de Huntington/patologia , Immunoblotting , Lítio/efeitos adversos , Masculino , Camundongos , Atividade Motora/efeitos dos fármacos , Fármacos Neuroprotetores/efeitos adversos , Ratos , Ratos Wistar , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
Huntington's disease (HD) is caused by a polyglutamine expansion in the Huntingtin (Htt) protein. Proteolytic cleavage of Htt into toxic N-terminal fragments is believed to be a key aspect of pathogenesis. The best characterized putative cleavage event is at amino acid 586, hypothesized to be mediated by caspase 6. A corollary of the caspase 6 cleavage hypothesis is that the caspase 6 fragment should be a toxic fragment. To test this hypothesis, and further characterize the role of this fragment, we have generated transgenic mice expressing the N-terminal 586 aa of Htt with a polyglutamine repeat length of 82 (N586-82Q), under the control of the prion promoter. N586-82Q mice show a clear progressive rotarod deficit by 4 months of age, and are hyperactive starting at 5 months, later changing to hypoactivity before early mortality. MRI studies reveal widespread brain atrophy, and histologic studies demonstrate an abundance of Htt aggregates, mostly cytoplasmic, which are predominantly composed of the N586-82Q polypeptide. Smaller soluble N-terminal fragments appear to accumulate over time, peaking at 4 months, and are predominantly found in the nuclear fraction. This model appears to have a phenotype more severe than current full-length Htt models, but less severe than HD mouse models expressing shorter Htt fragments. These studies suggest that the caspase 6 fragment may be a transient intermediate, that fragment size is a factor contributing to the rate of disease progression, and that short soluble nuclear fragments may be most relevant to pathogenesis.
Assuntos
Caspase 6/fisiologia , Doença de Huntington/metabolismo , Degeneração Neural/metabolismo , Proteínas do Tecido Nervoso/genética , Proteínas Nucleares/genética , Fragmentos de Peptídeos/genética , Animais , Atrofia , Modelos Animais de Doenças , Humanos , Proteína Huntingtina , Doença de Huntington/patologia , Doença de Huntington/fisiopatologia , Camundongos , Camundongos Endogâmicos , Camundongos Transgênicos , Degeneração Neural/patologia , Degeneração Neural/fisiopatologia , Proteínas do Tecido Nervoso/metabolismo , Proteínas do Tecido Nervoso/toxicidade , Proteínas Nucleares/metabolismo , Proteínas Nucleares/toxicidade , Fragmentos de Peptídeos/biossíntese , Fragmentos de Peptídeos/toxicidade , Expansão das Repetições de Trinucleotídeos/fisiologiaRESUMO
Apoptosis, or programmed cell death, is a cellular pathway involved in normal cell turnover, developmental tissue remodeling, embryonic development, cellular homeostasis maintenance and chemical-induced cell death. Caspases are a family of intracellular proteases that play a key role in apoptosis. Aberrant activation of caspases has been implicated in human diseases. In particular, numerous findings implicate Caspase-6 (Casp6) in neurodegenerative diseases, including Alzheimer disease (AD) and Huntington disease (HD), highlighting the need for a deeper understanding of Casp6 biology and its role in brain development. The use of targeted caspase-deficient mice has been instrumental for studying the involvement of caspases in apoptosis. The goal of this study was to perform an in-depth neuroanatomical and behavioral characterization of constitutive Casp6-deficient (Casp6-/-) mice in order to understand the physiological function of Casp6 in brain development, structure and function. We demonstrate that Casp6-/- neurons are protected against excitotoxicity, nerve growth factor deprivation and myelin-induced axonal degeneration. Furthermore, Casp6-deficient mice show an age-dependent increase in cortical and striatal volume. In addition, these mice show a hypoactive phenotype and display learning deficits. The age-dependent behavioral and region-specific neuroanatomical changes observed in the Casp6-/- mice suggest that Casp6 deficiency has a more pronounced effect in brain regions that are involved in neurodegenerative diseases, such as the striatum in HD and the cortex in AD.
Assuntos
Caspase 6/fisiologia , Degeneração Neural/enzimologia , Envelhecimento/patologia , Envelhecimento/fisiologia , Doença de Alzheimer/enzimologia , Doença de Alzheimer/patologia , Animais , Apoptose/fisiologia , Sequência de Bases , Comportamento Animal/fisiologia , Encéfalo/enzimologia , Encéfalo/patologia , Caspase 6/deficiência , Caspase 6/genética , Humanos , Doença de Huntington/enzimologia , Doença de Huntington/patologia , Camundongos , Camundongos Knockout , Atividade Motora/fisiologia , Degeneração Neural/genética , Degeneração Neural/patologia , Neurônios/enzimologia , Neurônios/patologia , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Receptores de N-Metil-D-Aspartato/fisiologiaRESUMO
BACKGROUND: The amelioration of behavioral and neuropathological deficits in mice expressing caspase-6-resistant (C6R) mutant huntingtin (mhtt), despite the presence of an expanded polyglutamine tract, highlights proteolysis of htt at the 586aa caspase-6 (casp6) site may be an important mechanism in the pathogenesis of Huntington disease (HD). One possible explanation of these effects is that C6R mhtt could act as a dominant negative on mhtt. OBJECTIVE AND METHODS: To determine if the neuroprotective effect observed in the C6R mice is due to dominant negative effects, we crossed the C6R mice to the YAC128 HD mouse model to generate mice expressing both caspase-cleavable and C6R mhtt (YAC/C6R) concurrently and assessed previously defined behavioral and neuropathological endpoints. RESULTS: Our results demonstrate that YAC/C6R animals exhibit similar motor abnormalities and learning deficits as the YAC128 mice. Neuropathological analysis reveals a significant decrease in brain weight and striatal volume in the YAC/C6R mice comparable to the YAC128 mice. In contrast, and similar to previous findings, C6R mice demonstrate preserved brain weight and striatal volume. As expected, body weight is significantly increased in the YAC/C6R mice due to the increased levels of htt. CONCLUSIONS: The results of this study suggest that the lack of an HD phenotype in the C6R mice is most likely due to the absence of cleavage of htt and not due to suppression of expression of mhtt.