RESUMO
Docosahexaenoic acid (DHA, C22:6 ω3) may be involved in various neuroprotective mechanisms that could prevent Alzheimer's disease (AD). Its influence has still been little explored regarding the dysfunction of the endolysosomal pathway, known as an early key event in the physiopathological continuum triggering AD. This dysfunction could result from the accumulation of degradation products of the precursor protein of AD, in particular the C99 fragment, capable of interacting with endosomal proteins and thus contributing to altering this pathway from the early stages of AD. This study aims to evaluate whether neuroprotection mediated by DHA can also preserve the endolysosomal function. AD-typical endolysosomal abnormalities were recorded in differentiated human SH-SY5Y neuroblastoma cells expressing the Swedish form of human amyloid precursor protein. This altered phenotype included endosome enlargement, the reduced secretion of exosomes, and a higher level of apoptosis, which confirmed the relevance of the cellular model chosen for studying the associated deleterious mechanisms. Second, neuroprotection mediated by DHA was associated with a reduced interaction of C99 with the Rab5 GTPase, lower endosome size, restored exosome production, and reduced neuronal apoptosis. Our data reveal that DHA may influence protein localization and interactions in the neuronal membrane environment, thereby correcting the dysfunction of endocytosis and vesicular trafficking associated with AD.
Assuntos
Doença de Alzheimer , Ácidos Docosa-Hexaenoicos , Endossomos , Lisossomos , Neurônios , Proteínas rab5 de Ligação ao GTP , Humanos , Ácidos Docosa-Hexaenoicos/farmacologia , Ácidos Docosa-Hexaenoicos/metabolismo , Doença de Alzheimer/metabolismo , Doença de Alzheimer/patologia , Proteínas rab5 de Ligação ao GTP/metabolismo , Endossomos/metabolismo , Neurônios/metabolismo , Neurônios/patologia , Neurônios/efeitos dos fármacos , Lisossomos/metabolismo , Linhagem Celular Tumoral , Precursor de Proteína beta-Amiloide/metabolismo , Apoptose , Fármacos Neuroprotetores/farmacologia , Sobrevivência Celular/efeitos dos fármacosRESUMO
Perturbations of cholesterol metabolism have been linked to neurodegenerative diseases. Glia-neuron crosstalk is essential to achieve a tight regulation of brain cholesterol trafficking. Adequate cholesterol supply from glia via apolipoprotein E-containing lipoproteins ensures neuronal development and function. The lipolysis-stimulated lipoprotein receptor (LSR), plays an important role in brain cholesterol homeostasis. Aged heterozygote Lsr+/- mice show altered brain cholesterol distribution and increased susceptibility to amyloid stress. Since LSR expression is higher in astroglia as compared to neurons, we sought to determine if astroglial LSR deficiency could lead to cognitive defects similar to those of Alzheimer's disease (AD). Cre recombinase was activated in adult Glast-CreERT/lsrfl/fl mice by tamoxifen to induce astroglial Lsr deletion. Behavioral phenotyping of young and old astroglial Lsr KO animals revealed hyperactivity during the nocturnal period, deficits in olfactory function affecting social memory and causing possible apathy, as well as visual memory and short-term working memory problems, and deficits similar to those reported in neurodegenerative diseases, such as AD. Furthermore, GFAP staining revealed astroglial activation in the olfactory bulb. Therefore, astroglial LSR is important for working, spatial, and social memory related to sensory input, and represents a novel pathway for the study of brain aging and neurodegeneration.
Assuntos
Astrócitos/metabolismo , Transtornos da Memória/metabolismo , Memória de Curto Prazo , Receptores de Lipoproteínas/metabolismo , Olfato , Animais , Colesterol/metabolismo , Transtornos da Memória/genética , Camundongos , Receptores de Lipoproteínas/genéticaRESUMO
Perturbations of lipid homeostasis manifest as dyslipidemias and obesity, which are significant risk factors for atherosclerosis and diabetes. Lipoprotein receptors in the liver are key players in the regulation of lipid homeostasis, among which the hepatic lipolysis stimulated lipoprotein receptor, LSR, was recently shown to play an important role in the removal of lipoproteins from the circulation during the postprandial phase. Since heterozygous LSR+/- mice demonstrate moderate dyslipidemia and develop higher body weight gain in response to high-fat diet compared with littermate LSR+/+ controls, we questioned if LSR heterozygosity could affect genes related to hepatic lipid metabolism. A target-specific qPCR array for 84 genes related to lipid metabolism was performed on mRNA isolated from livers of 6 mo old female LSR+/- mice and LSR+/+ littermates following a 6 wk period on a standard (STD) or high-fat diet (60% kcal, HFD). Of the 84 genes studied, 32 were significantly downregulated in STD-LSR+/- mice compared with STD-LSR+/+, a majority of which were PPARα target genes involved in lipid metabolism and transport, and insulin and adipokine-signaling pathways. Of these 32 genes, 80% were also modified in HFD-LSR+/+, suggesting that STD-LSR+/- mice demonstrated a predisposition towards a "high-fat"-like profile, which could reflect dysregulation of liver lipid homeostasis. Since similar profiles of genes were affected by either LSR heterozygosity or by high-fat diet, this would suggest that LSR is a key receptor in regulating hepatic lipid homeostasis, and whose downregulation combined with a Western-type diet may increase predisposition to diet-induced obesity.
Assuntos
Dieta Hiperlipídica/efeitos adversos , Homeostase/genética , Metabolismo dos Lipídeos/genética , Fígado/metabolismo , Receptores de Lipoproteínas/genética , Transcriptoma/genética , Animais , Regulação para Baixo/genética , Feminino , Heterozigoto , Insulina/genética , Lipídeos/genética , Camundongos , Obesidade/genética , Aumento de Peso/genéticaRESUMO
Deciphering the mechanism of Alzheimer's disease is a key element for designing an efficient therapeutic strategy. Molecular dynamics (MD) calculations, atomic force microscopy, and infrared spectroscopy were combined to investigate ß-amyloid (Aß1-42) peptide interactions with supported lipid bilayers (SLBs). The MD simulations showed that nascent Aß1-42 monomers remain anchored within a model phospholipid bilayer's hydrophobic core, which suggests their stability in their native environment. We tested this prediction experimentally by studying the behavior of Aß1-42 monomers and oligomers when interacting with SLBs. When Aß1-42 monomers and oligomers were self-assembled with a lipid bilayer and deposited as an SLB, they remain within the bilayers. Their presence in the bilayers induces destabilization of the model membranes. No specific interactions between Aß1-42 and the SLBs were detected when SLBs free of Aß1-42 were exposed to Aß1-42. This study suggests that Aß can remain in the membrane after cleavage by γ-secretase and cause severe damage to the membrane.
Assuntos
Doença de Alzheimer , Humanos , Peptídeos beta-Amiloides/química , Fragmentos de Peptídeos/química , Bicamadas Lipídicas/químicaRESUMO
The development of novel therapeutic strategies for Alzheimer's disease (AD) represents one of the biggest unmet medical needs today. Application of neurotrophic factors able to modulate neuronal survival and synaptic connectivity is a promising therapeutic approach for AD. We aimed to determine whether the loco-regional delivery of ciliary neurotrophic factor (CNTF) could prevent amyloid-beta (Abeta) oligomer-induced synaptic damages and associated cognitive impairments that typify AD. To ensure long-term administration of CNTF in the brain, we used recombinant cells secreting CNTF encapsulated in alginate polymers. The implantation of these bioreactors in the brain of Abeta oligomer-infused mice led to a continuous secretion of recombinant CNTF and was associated with the robust improvement of cognitive performances. Most importantly, CNTF led to full recovery of cognitive functions associated with the stabilization of synaptic protein levels in the Tg2576 AD mouse model. In vitro as well as in vivo, CNTF activated a Janus kinase/signal transducer and activator of transcription-mediated survival pathway that prevented synaptic and neuronal degeneration. These preclinical studies suggest that CNTF and/or CNTF receptor-associated pathways may have AD-modifying activity through protection against progressive Abeta-related memory deficits. Our data also encourage additional exploration of ex vivo gene transfer for the prevention and/or treatment of AD.
Assuntos
Doença de Alzheimer/complicações , Fator Neurotrófico Ciliar/biossíntese , Fator Neurotrófico Ciliar/uso terapêutico , Transtornos da Memória/etiologia , Transtornos da Memória/terapia , Sinapses/efeitos dos fármacos , Doença de Alzheimer/genética , Peptídeos beta-Amiloides/administração & dosagem , Peptídeos beta-Amiloides/metabolismo , Peptídeos beta-Amiloides/farmacologia , Precursor de Proteína beta-Amiloide/genética , Animais , Apoptose/genética , Encéfalo/patologia , Contagem de Células/métodos , Terapia Baseada em Transplante de Células e Tecidos/métodos , Células Cultivadas , Fator Neurotrófico Ciliar/administração & dosagem , Modelos Animais de Doenças , Inibidores Enzimáticos/farmacologia , Ensaio de Imunoadsorção Enzimática/métodos , Regulação da Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica/genética , Proteínas de Fluorescência Verde/genética , Humanos , Masculino , Aprendizagem em Labirinto/fisiologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Mutação/genética , Neurônios/efeitos dos fármacos , Neurônios/metabolismo , Fragmentos de Peptídeos/administração & dosagem , Fragmentos de Peptídeos/farmacologia , Transdução de Sinais/efeitos dos fármacos , Sinapses/metabolismo , Sinaptossomos/metabolismo , Sinaptossomos/patologia , Sinaptossomos/ultraestrutura , Fatores de Tempo , Transfecção/métodosRESUMO
Alzheimer's disease (AD) is a major public health concern due to longer life expectancy in the Western countries. Amyloid-beta (Abeta) oligomers are considered the proximate effectors in the early stages of AD. AD-related cognitive impairment, synaptic loss and neurodegeneration result from interactions of Abeta oligomers with the synaptic membrane and subsequent activation of pro-apoptotic signalling pathways. Therefore, membrane structure and lipid status appear determinant in Abeta-induced toxicity. Numerous epidemiological studies have highlighted the beneficial influence of docosahexaenoic acid (DHA, C22:6 n-3) on the preservation of synaptic function and memory capacities in aged individuals or upon Abeta exposure, whereas its deficiency is presented as a risk factor for AD. An elevated number of studies have been reporting the beneficial effects of dietary DHA supplementation on cognition and synaptic integrity in various AD models. In this review, we describe the important potential of DHA to preserve neuronal and brain functions and classified its numerous molecular and cellular effects from impact on membrane lipid content and organisation to activation of signalling pathways sustaining synaptic function and neuronal survival. DHA appears as one of the most valuable diet ingredients whose neuroprotective properties could be crucial for designing nutrition-based strategies able to prevent AD as well as other lipid- and age-related diseases whose prevalence is progressing in elderly populations.
Assuntos
Doença de Alzheimer/patologia , Modelos Animais de Doenças , Ácidos Docosa-Hexaenoicos/farmacologia , Camundongos , Sinapses/efeitos dos fármacos , Doença de Alzheimer/metabolismo , Doença de Alzheimer/fisiopatologia , Peptídeos beta-Amiloides/metabolismo , Animais , Transtornos Cognitivos/metabolismo , Transtornos Cognitivos/fisiopatologia , Transtornos Cognitivos/prevenção & controle , Citoproteção/efeitos dos fármacos , Ácidos Docosa-Hexaenoicos/metabolismo , Humanos , Modelos Biológicos , Neurônios/efeitos dos fármacos , Fármacos Neuroprotetores/farmacologiaRESUMO
As a hepatic receptor for triglyceride-rich lipoproteins, the lipolysis-stimulated lipoprotein receptor (LSR) may be involved in the dynamics of lipid distribution between the liver and peripheral tissues. Here, we explore the potential role of leptin in regulating LSR. At physiological concentrations (1-10 ng/ml), leptin increased LSR protein and mRNA levels in Hepa1-6 cells through an ERK1/2-dependent and α-amanitin-sensitive pathway. In vivo, leptin treatment of C57BL6/Rj mice (1 µg 2×/d, 8 d) led to a significant increase in hepatic LSR mRNA and protein, decreased liver triglycerides and increased VLDL secretion as compared to controls. LSR(+/-) mice with elevated postprandial lipemia placed on a high-fat (60% kcal) diet exhibited accelerated weight gain and increased fat mass as compared to controls. While plasma leptin levels were increased 3-fold, hepatic leptin receptor protein levels and phosphorylation of ERK1/2 were significantly reduced. Therefore, leptin is an important regulator of LSR protein levels providing the means for the control of hepatic uptake of lipids during the postprandial phase. However, this may no longer be functional in LSR(+/-) mice placed under a chronic dietary fat load, suggesting that this animal model could be useful for the study of molecular mechanisms involved in peripheral leptin resistance.
Assuntos
Leptina/farmacologia , Metabolismo dos Lipídeos/efeitos dos fármacos , Lipólise/efeitos dos fármacos , Fígado/efeitos dos fármacos , Período Pós-Prandial , Receptores de Lipoproteínas/metabolismo , Regulação para Cima/efeitos dos fármacos , Animais , Western Blotting , Peso Corporal/efeitos dos fármacos , Linhagem Celular , Imunofluorescência , Leptina/sangue , Fígado/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Reação em Cadeia da PolimeraseRESUMO
The regulation of cholesterol, an essential brain lipid, ensures proper neuronal development and function, as demonstrated by links between perturbations of cholesterol metabolism and neurodegenerative diseases, including Alzheimer's disease. The central nervous system (CNS) acquires cholesterol via de novo synthesis, where glial cells provide cholesterol to neurons. Both lipoproteins and lipoprotein receptors are key elements in this intercellular transport, where the latter recognize, bind and endocytose cholesterol containing glia-produced lipoproteins. CNS lipoprotein receptors are like those in the periphery, among which include the ApoB, E binding lipolysis stimulated lipoprotein receptor (LSR). LSR is a multimeric protein complex that has multiple isoforms including α and α', which are seen as a doublet at 68 kDa, and ß at 56 kDa. While complete inactivation of murine lsr gene is embryonic lethal, studies on lsr +/- mice revealed altered brain cholesterol distribution and cognitive functions. In the present study, LSR profiling in different CNS regions revealed regiospecific expression of LSR at both RNA and protein levels. At the RNA level, the hippocampus, hypothalamus, cerebellum, and olfactory bulb, all showed high levels of total lsr compared to whole brain tissues, whereas at the protein level, only the hypothalamus, olfactory bulb, and retina showed the highest levels of total LSR. Interestingly, major regional changes in LSR expression were observed in aged mice which suggests changes in cholesterol homeostasis in specific structures in the aging brain. Immunocytostaining of primary cultures of mature murine neurons and glial cells isolated from different CNS regions showed that LSR is expressed in both neurons and glial cells. However, lsr RNA expression in the cerebellum was predominantly higher in glial cells, which was confirmed by the immunocytostaining profile of cerebellar neurons and glia. Based on this observation, we would propose that LSR in glial cells may play a key role in glia-neuron cross talk, particularly in the feedback control of cholesterol synthesis to avoid cholesterol overload in neurons and to maintain proper functioning of the brain throughout life.
Assuntos
Envelhecimento/metabolismo , Encéfalo/metabolismo , Receptores de Lipoproteínas/metabolismo , Envelhecimento/genética , Animais , Encéfalo/anatomia & histologia , Colesterol/metabolismo , Homeostase , Humanos , Lipólise , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Neuroglia/metabolismo , Neurônios/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Receptores de Lipoproteínas/deficiência , Receptores de Lipoproteínas/genética , Distribuição Tecidual , TranscriptomaRESUMO
The ε4 allele of the apolipoprotein E (APOE) gene common polymorphism is the strongest genetic risk factor for Alzheimer's disease (AD). Human APOE gene is located on chromosome 19q13.1, a region linked to AD that also includes the LSR gene, which encodes the lipolysis-stimulated lipoprotein receptor (LSR). As an APOE receptor, LSR is involved in the regulation of lipid homeostasis in both periphery and brain. This study aimed to determine the potential interactions between 2 LSR genetic variants, rs34259399 and rs916147, and the APOE common polymorphism in 142 AD subjects (mean age: 73.16 ± 8.50 years) and 63 controls (mean age: 70.41 ± 8.49 years). A significant epistatic interaction was observed between APOE and both LSR variants, rs34259399 (beta = -0.95; p = 2 × 10-5) and rs916147 (beta = -0.83; p = 6.8 × 10-3). Interestingly, the interaction of LSR polymorphisms with APOE non-ε4 alleles increased AD risk. This indicates the existence of complex molecular interactions between these 2 neighboring genes involved in the pathogenesis of AD, which merits further investigation.
Assuntos
Doença de Alzheimer/genética , Apolipoproteínas E/metabolismo , Epistasia Genética , Receptores de Lipoproteínas/genética , Receptores de Lipoproteínas/metabolismo , Idoso , Feminino , Frequência do Gene , Genótipo , Humanos , Masculino , Polimorfismo de Nucleotídeo Único , Fatores de TranscriçãoRESUMO
Alzheimer's disease (AD) is a major public health concern in all countries. Although the precise cause of AD is still unknown, a growing body of evidence supports the notion that soluble amyloid beta-peptide (Abeta) may be the proximate cause of synaptic injuries and neuronal death early in the disease. AD patients display lower levels of docosahexaenoic acid (DHA, C22:6 ; n-3) in plasma and brain tissues as compared to age-matched controls. Furthermore, epidemiological studies suggest that high DHA intake might have protective properties against neurodegenerative diseases. These observations are supported by in vivo studies showing that DHA-rich diets limits the synaptic loss and cognitive defects induced by Abeta peptide. Although the molecular basis of these neuroprotective effects remains unknown, several mechanisms have been proposed such as (i) regulation of the expression of potentially protective genes, (ii) activation of anti-inflammatory pathways, (iii) modulation of functional properties of the synaptic membranes along with changes in their physicochemical and structural features.
Assuntos
Doença de Alzheimer/dietoterapia , Doença de Alzheimer/prevenção & controle , Peptídeos beta-Amiloides/antagonistas & inibidores , Encéfalo/efeitos dos fármacos , Encéfalo/metabolismo , Ácidos Docosa-Hexaenoicos/uso terapêutico , Encefalite/dietoterapia , Fármacos Neuroprotetores/uso terapêutico , Doença de Alzheimer/metabolismo , Peptídeos beta-Amiloides/metabolismo , Animais , Encéfalo/fisiopatologia , Ácidos Docosa-Hexaenoicos/metabolismo , Encefalite/metabolismo , Encefalite/fisiopatologia , Alimentos Formulados/normas , Humanos , Metabolismo dos Lipídeos/efeitos dos fármacos , Metabolismo dos Lipídeos/fisiologia , Fármacos Neuroprotetores/metabolismo , Sinapses/efeitos dos fármacos , Sinapses/metabolismoRESUMO
Although a major risk factor for Alzheimer's disease (AD), the "aging" parameter is not systematically considered in preclinical validation of anti-AD drugs. To explore how aging affects neuronal reactivity to anti-AD agents, the ciliary neurotrophic factor (CNTF)-associated pathway was chosen as a model. Comparison of the neuroprotective properties of CNTF in 6- and 18-month old mice revealed that CNTF resistance in the older animals is associated with the exclusion of the CNTF-receptor subunits from rafts and their subsequent dispersion to non-raft cortical membrane domains. This age-dependent membrane remodeling prevented both the formation of active CNTF-receptor complexes and the activation of prosurvival STAT3 and ERK1/2 pathways, demonstrating that age-altered membranes impaired the reactivity of potential therapeutic targets. CNTF-receptor distribution and CNTF signaling responses were improved in older mice receiving dietary docosahexaenoic acid, with CNTF-receptor functionality being similar to those of younger mice, pointing toward dietary intervention as a promising adjuvant strategy to maintain functional neuronal membranes, thus allowing the associated receptors to respond appropriately to anti-AD agents.
Assuntos
Envelhecimento/genética , Envelhecimento/fisiologia , Encéfalo/citologia , Membrana Celular/fisiologia , Neurônios/citologia , Nootrópicos/uso terapêutico , Animais , Fator Neurotrófico Ciliar/fisiologia , Gorduras Insaturadas na Dieta , Ácidos Docosa-Hexaenoicos , Sistema de Sinalização das MAP Quinases/fisiologia , Masculino , Microdomínios da Membrana , Camundongos Endogâmicos C57BL , Receptor do Fator Neutrófico Ciliar/fisiologia , Fator de Transcrição STAT3/metabolismo , Transdução de SinaisRESUMO
Recent data have revealed that soluble oligomeric forms of amyloid peptide (Abeta) may be the proximate effectors of the neuronal injury and death occurring in Alzheimer's disease (AD). However, the molecular mechanisms associated with the neuronal cell death induced by the nonfibrillar Abeta remain to be elucidated. In this study, we investigated the role of the cytosolic Ca2+-dependent phospholipase A2 (cPLA2), and its associated metabolic pathway, i.e., the arachidonic acid (AA) cascade, in the apoptotic cell death induced by soluble oligomers of Abeta. The treatment of rat cortical neurons with low concentrations of soluble Abeta(1-40) or Abeta(1-42) peptide resulted in an early calcium-dependent release of AA associated with a transient relocalization of cPLA2. Both cPLA2 antisense oligonucleotides and a selective inhibitor of cPLA2 activity abolished the release of AA from neurons and also protected cells against apoptosis induced by Abeta. Furthermore, inhibitors of the PKC, p38, and MEK/ERK pathways that are involved in cPLA2 phosphorylation and activation reduced Abeta-induced cell death. Finally, we demonstrate that inhibitors of cyclooxygenase-2 reduced the Abeta-induced cell death by 55%. Our studies suggest a novel neuronal response of soluble oligomers of Abeta, which occurs through a cPLA2 signaling cascade and an AA-dependent death pathway. This may prove to be crucial in AD processes and could provide important targets for drug development.
Assuntos
Peptídeos beta-Amiloides/metabolismo , Apoptose/fisiologia , Citosol/enzimologia , Neurônios/enzimologia , Neurônios/fisiologia , Fragmentos de Peptídeos/metabolismo , Fosfolipases A/metabolismo , Animais , Ácido Araquidônico/metabolismo , Células Cultivadas , Córtex Cerebral/citologia , Ciclo-Oxigenase 2 , Humanos , Proteínas de Membrana , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Fosfolipases A2 , Prostaglandina-Endoperóxido Sintases/metabolismo , Ratos , SolubilidadeRESUMO
Lactational exposure to low levels of the sum of the six indicator polychlorinated biphenyls (Σ6 NDL-PCBs, 10ng/kg/day) is known to lead to persistent anxious behavior in young and adult offspring mice at postnatal days 40 and 160, respectively. At more advanced life stages, we evaluated the effects on the mouse brain of neuronal stress induced by the synaptotoxic amyloid-beta (Aß) peptide. Perinatal exposure of lactating mice to Σ6 NDL-PCBs did not affect short-term memory performances of their offspring male mice aged 14 months as compared to control PCB-naive mice. However, following intracerebroventricular injection of soluble Aß oligomers, significant impairments in long-term memory were detected in the mice that had been lactationally treated with Σ6 NDL-PCBs. In addition, immunoblot analyses of the synaptosomal fraction of hippocampal tissues from treated mice revealed a lower expression of the synaptic proteins synaptophysin and PSD-95. Though preliminary, our findings suggest for the first time that early exposure to low levels of NDL-PCBs induce late neuronal vulnerability to amyloid stress. Additional experiments are needed to confirm whether early environmental influences are involved in the etiology of brain aging and cognitive decline.
Assuntos
Envelhecimento , Poluentes Ambientais/toxicidade , Lactação/efeitos dos fármacos , Bifenilos Policlorados/toxicidade , Efeitos Tardios da Exposição Pré-Natal/induzido quimicamente , Efeitos Tardios da Exposição Pré-Natal/fisiopatologia , Peptídeos beta-Amiloides/administração & dosagem , Animais , Transtornos Cognitivos/etiologia , Suscetibilidade a Doenças , Proteína 4 Homóloga a Disks-Large , Feminino , Guanilato Quinases/metabolismo , Injeções Intraventriculares , Masculino , Aprendizagem em Labirinto/efeitos dos fármacos , Proteínas de Membrana/metabolismo , Memória de Curto Prazo/efeitos dos fármacos , Camundongos , GravidezRESUMO
The developing central nervous system is particularly vulnerable to environmental contaminants such as non-dioxin-like polychlorinated biphenyls (NDL-PCBs). This study investigated the potential oxidative effects in mice pups exposed via lactation to the sum of the six indicator NDL-PCBs (∑6 NDL-PCBs) at 0, 1, 10 and 100 ng/kg per 14 days, constituting levels below the guidance values fixed by French food safety agencies for humans at 10 ng/kg body weight per day. For this purpose, the oxidative status was assessed by flow cytometry via dichloro-dihydro-fluorescein diacetate in the cerebellum of juvenile male offspring mice during brain growth spurt [postnatal day (PND) 14]. No significant differences were found in the levels of reactive oxygen species in the cerebellar neurons or glial cells (astrocytes, oligodendrocytes and microglia) of lactationally exposed male mice at PND 14 (p>0.05). Concordantly, oxidative-stress related gene expression was measured by qPCR for catalase, copper zinc superoxide dismutase 1, glyoxalase 1, glutathione peroxidase 1, and glutathione reductase 1, in the cerebellum at PND 14 appeared unaffected, as also verified at the protein level by immunoblots. Moreover, transcriptomic data from our previous work have not shown differences in the mRNA expressions of genes belonging to GO terms involved in oxidative stress in neurons of male mice exposed to ∑6 NDL-PCBs compared to controls; except for glyoxalase 1 which was downregulated in neurons isolated from exposed group compared to controls. Our findings suggest that lactational exposure to NDL-PCBs at environmental relevant concentrations may not cause significant oxidative effect on juvenile cerebellum.
Assuntos
Cerebelo/efeitos dos fármacos , Lactação , Estresse Oxidativo/efeitos dos fármacos , Bifenilos Policlorados/toxicidade , Animais , Antioxidantes/metabolismo , Encéfalo/crescimento & desenvolvimento , Cerebelo/citologia , Cerebelo/crescimento & desenvolvimento , Feminino , Inocuidade dos Alimentos , Masculino , Camundongos , Neuroglia/metabolismo , Neurônios/metabolismo , Bifenilos Policlorados/farmacocinética , Espécies Reativas de Oxigênio/metabolismo , TranscriptomaRESUMO
Oligomeric amyloid-ß (Aß) peptide contributes to impaired synaptic connections and neurodegenerative processes, and as such, represents a primary therapeutic target for Alzheimer's disease (AD)-modifying approaches. However, the lack of efficacy of drugs that inhibit production of Aß demonstrates the need for a better characterization of its toxic effects, both on synaptic and neuronal function. Here, we used conditioned medium obtained from recombinant HEK-AßPP cells expressing the human amyloid-ß protein precursor (Aß-CM), to investigate Aß-induced neurotoxic and synaptotoxic effects. Characterization of Aß-CM revealed that it contained picomolar amounts of cell-secreted Aß in its soluble form. Incubation of primary cortical neurons with Aß-CM led to significant decreases in synaptic protein levels as compared to controls. This effect was no longer observed in neurons incubated with conditioned medium obtained from HEK-AßPP cells grown in presence of the γ-secretase inhibitor, Semagacestat or LY450139 (LY-CM). However, neurotoxic and pro-apoptotic effects of Aß-CM were only partially prevented using LY-CM, which could be explained by other deleterious compounds related to chronic oxidative stress that were released by HEK-AßPP cells. Indeed, full neuroprotection was observed in cells exposed to LY-CM by additional treatment with the antioxidant resveratrol, or with the pluripotent n-3 polyunsaturated fatty acid docosahexaenoic acid. Inhibition of Aß production appeared necessary but insufficient to prevent neurodegenerative effects associated with AD due to other neurotoxic compounds that could exert additional deleterious effects on neuronal function and survival. Therefore, association of various types of protective agents needs to be considered when developing strategies for AD treatment.
Assuntos
Peptídeos beta-Amiloides/farmacologia , Neurônios/efeitos dos fármacos , Fármacos Neuroprotetores/farmacologia , Alanina/análogos & derivados , Alanina/farmacologia , Precursor de Proteína beta-Amiloide/genética , Precursor de Proteína beta-Amiloide/metabolismo , Análise de Variância , Animais , Azepinas/farmacologia , Caspase 3/metabolismo , Células Cultivadas , Córtex Cerebral/citologia , Meios de Cultivo Condicionados/farmacologia , Embrião de Mamíferos , Proteína Glial Fibrilar Ácida/metabolismo , Humanos , Estresse Oxidativo/efeitos dos fármacos , Estresse Oxidativo/genética , Fosfopiruvato Hidratase/metabolismo , Ratos , Transdução de Sinais/efeitos dos fármacos , TransfecçãoRESUMO
Lipids are the fundamental structural components of biological membranes. For a long time considered as simple barriers segregating aqueous compartments, membranes are now viewed as dynamic interfaces providing a molecular environment favorable to the activity of membrane-associated proteins. Interestingly, variations in membrane lipid composition, whether quantitative or qualitative, play a crucial role in regulation of membrane protein functionalities. Indeed, a variety of alterations in brain lipid composition have been associated with the processes of normal and pathological aging. Although not establishing a direct cause-and-effect relationship between these complex modifications in cerebral membranes and the process of cognitive decline, evidence shows that alterations in membrane lipid composition affect important physicochemical properties notably impacting the lateral organization of membranes, and thus microdomains. It has been suggested that preservation of microdomain functionality may represent an effective strategy for preventing or decelerating neuronal dysfunction and cerebral vulnerability, processes that are both aggravated by aging. The working hypothesis developed in this review proposes that preservation of membrane organization, for example, through nutritional supplementation of docosahexaenoic acid, could prevent disturbances in and preserve effective cerebral function.
Assuntos
Envelhecimento , Encéfalo/metabolismo , Lipídeos de Membrana/metabolismo , Microdomínios da Membrana/metabolismo , Proteínas de Membrana/metabolismo , Doença de Alzheimer/metabolismo , Doença de Alzheimer/prevenção & controle , Encéfalo/efeitos dos fármacos , Ácidos Docosa-Hexaenoicos/administração & dosagem , Ácidos Docosa-Hexaenoicos/metabolismo , Humanos , Microdomínios da Membrana/química , Microdomínios da Membrana/efeitos dos fármacos , Modelos Biológicos , Fármacos Neuroprotetores/administração & dosagem , Fármacos Neuroprotetores/metabolismoRESUMO
Neuronal cell death in Alzheimer's disease (AD) is partly induced by the interaction of the amyloid-beta peptide (Abeta) with the plasma membrane of target cells. Accordingly, recent studies have suggested that cholesterol, an important component of membranes that controls their physical properties and functions, plays a critical role in neurodegenerative diseases. We report here that the enrichment of the neuronal plasma membrane with cholesterol protects cortical neurons from apoptosis induced by soluble oligomers of the Abeta(1-40) peptide. Conversely, cholesterol depletion using cyclodextrin renders cells more vulnerable to the cytotoxic effects of the Abeta-soluble oligomers. Increasing the cholesterol content of small unilamellar liposomes also decreases Abeta-dependent liposome fusion. We clearly demonstrate that cholesterol protection is specific to the soluble conformation of Abeta, because we observed no protective effects on cortical neurons treated by amyloid fibrils of the Abeta(1-40) peptide. This may provide a new opportunity for the development of an effective AD therapy as well as elucidate the impact of the cholesterol level during AD development.
Assuntos
Peptídeos beta-Amiloides/antagonistas & inibidores , Apoptose/efeitos dos fármacos , Colesterol/farmacologia , Lipídeos de Membrana/farmacologia , Neurônios/efeitos dos fármacos , Fragmentos de Peptídeos/antagonistas & inibidores , Doença de Alzheimer/metabolismo , Doença de Alzheimer/patologia , Peptídeos beta-Amiloides/farmacologia , Peptídeos beta-Amiloides/toxicidade , Animais , Biopolímeros , Caspase 8 , Caspases/metabolismo , Células Cultivadas/efeitos dos fármacos , Células Cultivadas/ultraestrutura , Córtex Cerebral/citologia , Córtex Cerebral/embriologia , Citoesqueleto/efeitos dos fármacos , Citoesqueleto/ultraestrutura , Relação Dose-Resposta a Droga , Ativação Enzimática/efeitos dos fármacos , Bicamadas Lipídicas , Lipossomos , Fluidez de Membrana , Fusão de Membrana/efeitos dos fármacos , Neurônios/ultraestrutura , Estresse Oxidativo , Fragmentos de Peptídeos/farmacologia , Fragmentos de Peptídeos/toxicidade , Conformação Proteica , Ratos , Ratos Wistar , SolubilidadeRESUMO
Alzheimer's disease (AD) is a neurodegenerative disease that has been linked to changes in cholesterol metabolism. Neuronal cholesterol content significantly influences the pro-apoptotic effect of amyloid-ß peptide42 (Aß42), which plays a key role in AD development. We previously reported that aged mice with reduced expression of the lipolysis stimulated lipoprotein receptor (LSR+/-), demonstrate membrane cholesterol accumulation and decreased intracellular lipid droplets in several brain regions, suggesting a potential role of LSR in brain cholesterol distribution. We questioned if these changes rendered the LSR+/- mouse more susceptible to Aß42-induced cognitive and biochemical changes. Results revealed that intracerebroventricular injection of oligomeric Aß42 in male 15-month old LSR+/+ and LSR+/- mice led to impairment in learning and long-term memory and decreased cortical cholesterol content of both groups; these effects were significantly amplified in the Aß42-injected LSR+/- group. Total latency of the Morris test was significantly and negatively correlated with cortical cholesterol content of the LSR+/- mice, but not of controls. Significantly lower cortical PSD95 and SNAP-25 levels were detected in Aß42-injected LSR+/- mice as compared to Aß42-injected LSR+/+ mice. In addition, 24S-hydroxy cholesterol metabolite levels were significantly higher in the cortex of LSR+/- mice. Taken together, these results suggest that changes in cortex cholesterol regulation as a result of the LSR+/- genotype were linked to increased susceptibility to amyloid stress, and we would therefore propose the aged LSR+/- mouse as a new model for understanding the link between modified cholesterol regulation as a risk factor for AD.
Assuntos
Peptídeos beta-Amiloides/farmacologia , Córtex Cerebral/efeitos dos fármacos , Córtex Cerebral/metabolismo , Colesterol/metabolismo , Fragmentos de Peptídeos/farmacologia , Receptores de Lipoproteínas/deficiência , Análise de Variância , Animais , Proteína 4 Homóloga a Disks-Large , Regulação da Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica/genética , Guanilato Quinases/metabolismo , Hipocampo/efeitos dos fármacos , Hipocampo/metabolismo , Hidroxicolesteróis/metabolismo , Aprendizagem em Labirinto/efeitos dos fármacos , Aprendizagem em Labirinto/fisiologia , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , RNA Mensageiro , Receptores de Lipoproteínas/genética , Análise de Regressão , Proteína 25 Associada a Sinaptossoma/metabolismoRESUMO
Docosahexaenoic acid (DHA) is increasingly considered for its health benefits. However, its use as functional food ingredient is still limited by its instability. In this work, we developed an efficient and solvent-free bioprocess for the synthesis of a phenolic ester of DHA. A fed-batch process catalyzed by Candida antarctica lipase B was optimised, leading to the production of 440 g/L vanillyl ester (DHA-VE). Structural characterisation of the purified product indicated acylation of the primary OH group of vanillyl alcohol. DHA-VE exhibited a high radical scavenging activity in acellular systems. In vivo experiments showed increased DHA levels in erythrocytes and brain tissues of mice fed DHA-VE-supplemented diet. Moreover, in vitro neuroprotective properties of DHA-VE were demonstrated in rat primary neurons exposed to amyloid-ß oligomers. In conclusion, DHA-VE synergized the main beneficial effects of two common natural biomolecules and therefore appears a promising functional ingredient for food applications.
Assuntos
Álcoois Benzílicos/química , Ácidos Docosa-Hexaenoicos/química , Ésteres/metabolismo , Oxirredutases/química , Animais , Encéfalo/efeitos dos fármacos , Encéfalo/metabolismo , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Dieta , Ácidos Docosa-Hexaenoicos/metabolismo , Enzimas Imobilizadas/metabolismo , Eritrócitos/efeitos dos fármacos , Eritrócitos/metabolismo , Proteínas Fúngicas/metabolismo , Lipase/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Neurônios/citologia , Neurônios/efeitos dos fármacos , Neurônios/metabolismo , Fármacos Neuroprotetores/química , Fármacos Neuroprotetores/metabolismo , Fármacos Neuroprotetores/farmacologia , Oxirredutases/biossíntese , Oxirredutases/farmacologia , Ratos , Ratos WistarRESUMO
Oxidative stress has been demonstrated in Alzheimer's disease (AD) brain and may affect glutamate transport (GT), thereby leading to excitotoxic neuronal death. Since oxidative stress markers have been shown also in peripheral tissues, we investigated possible GT alterations in fibroblast cultures obtained from 18 patients with AD and 15 control patients and analyzed the effects of the lipoperoxidation product 4-hydroxynonenal (4-HNE) and antioxidants. Basal GT was decreased by 60% in fibroblasts from patients with AD versus control patients. Exposure to HNE did not affect GT in control patients, but it reduced GT by 50% in patients with AD, without any concomitant change in cell viability; conversely, HNE exposure induced a larger increase in ROS intracellular levels in AD than in control fibroblasts. Glutathione and N-acetylcysteine completely blocked 4-HNE effects and also increased basal uptake in AD cells. Moreover, inhibition of glutathione synthesis in control fibroblasts by pretreatment with buthionine sulfoximine resulted in GT reduction (40%) and an increase in ROS levels after exposure to 4-HNE. Nevertheless, since there are no differences between GSH basal level in controls and patients with AD, the alteration of other antioxidant systems cannot be excluded. Our study supports the hypothesis of a systemic impairment of GT in AD, possibly linked to oxidative stress and to reduced antioxidant defenses, which may be partially reversed by antioxidant treatment. Therefore, we suggest fibroblast cultures as a tool for exploring pathogenetic mechanisms and possible therapeutic strategies in patients with AD.