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1.
Epigenetics ; 15(8): 781-799, 2020 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-32019393

RESUMO

The Presenilin1 (PSEN1) gene encodes the catalytic peptide of the γ-secretase complex, a key enzyme that cleaves the amyloid-ß protein precursor (AßPP), to generate the amyloid-ß (Aß) peptides, involved in Alzheimer's Disease (AD). Other substrates of the γ-secretase, such as E-cadherin and Notch1, are involved in neurodevelopment and haematopoiesis. Gene-specific DNA methylation influences PSEN1 expression in AD animal models. Here we evaluated canonical and non-canonical cytosine methylation patterns of the PSEN1 5'-flanking during brain development and AD progression, in DNA extracted from the frontal cortex of AD transgenic mice (TgCRND8) and post-mortem human brain. Mapping CpG and non-CpG methylation revealed different methylation profiles in mice and humans. PSEN1 expression only correlated with DNA methylation in adult female mice. However, in post-mortem human brain, lower methylation, both at CpG and non-CpG sites, correlated closely with higher PSEN1 expression during brain development and in disease progression. PSEN1 methylation in blood DNA was significantly lower in AD patients than in controls. The present study is the first to demonstrate a temporal correlation between dynamic changes in PSEN1 CpG and non-CpG methylation patterns and mRNA expression during neurodevelopment and AD neurodegeneration. These observations were made possible by the use of an improved bisulphite methylation assay employing primers that are not biased towards non-CpG methylation. Our findings deepen the understanding of γ-secretase regulation and support the hypothesis that epigenetic changes can promote the pathophysiology of AD. Moreover, they suggest that PSEN1 DNA methylation in peripheral blood may provide a biomarker for AD.

2.
Artigo em Inglês | MEDLINE | ID: mdl-31202182

RESUMO

The multifactorial nature of Late Onset Alzheimer's Disease (LOAD), the AD form of major relevance on epidemiological and social aspects, has driven the original investigation by LC-MS and top-down proteomics approach of the protein repertoire of the brain tissue of TgCRND8 model mice fed with a diet deficient in B vitamins. The analysis of the acid-soluble fraction of brain tissue homogenates identified a list of proteins and peptides, proteoforms and PTMs. In order to disclose possible modulations, their relative quantification in wild type and AD model mice under both B vitamin deficient and control diets was performed. The levels of metallothionein III, guanine nucleotide-binding protein G(I)/G(S)/G(O) subunit gamma-2 and brain acid soluble protein 1 showed statistically significant alterations depending on genotype, diet or both effects, respectively. Particularly, metallothionein III exhibited increased levels in TgCRND8 mice under B vitamin deficient diet with respect to wild type mice under both diets. Brain acid soluble protein 1 showed the opposite, revealing decreased levels in all diet groups of AD model mice with respect to wild type mice in control diet. Lower levels of brain acid soluble protein 1 were also observed in wild type mice under deficiency of B vitamins. These results, besides contributing to increase the knowledge of AD at molecular level, give new suggestions for deeply investigating metallothionein III and brain acid soluble protein 1 in AD.


Assuntos
Doença de Alzheimer/metabolismo , Encéfalo/metabolismo , Hiper-Homocisteinemia/metabolismo , Proteoma/metabolismo , Complexo Vitamínico B/análise , Doença de Alzheimer/genética , Peptídeos beta-Amiloides/metabolismo , Animais , Química Encefálica , Proteínas de Ligação a Calmodulina/genética , Proteínas de Ligação a Calmodulina/metabolismo , Cromatografia Líquida , Proteínas do Citoesqueleto/genética , Proteínas do Citoesqueleto/metabolismo , Modelos Animais de Doenças , Feminino , Humanos , Hiper-Homocisteinemia/etiologia , Hiper-Homocisteinemia/genética , Masculino , Espectrometria de Massas , Camundongos , Camundongos Transgênicos , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismo , Proteoma/química , Proteoma/genética , Complexo Vitamínico B/metabolismo
4.
Antioxidants (Basel) ; 6(4)2017 Sep 30.
Artigo em Inglês | MEDLINE | ID: mdl-28973985

RESUMO

Recent evidence emphasizes the role of dysregulated one-carbon metabolism in Alzheimer's Disease (AD). Exploiting a nutritional B-vitamin deficiency paradigm, we have previously shown that PSEN1 and BACE1 activity is modulated by one-carbon metabolism, leading to increased amyloid production. We have also demonstrated that S-adenosylmethionine (SAM) supplementation contrasted the AD-like features, induced by B-vitamin deficiency. In the present study, we expanded these observations by investigating the effects of SAM and SOD (Superoxide dismutase) association. TgCRND8 AD mice were fed either with a control or B-vitamin deficient diet, with or without oral supplementation of SAM + SOD. We measured oxidative stress by lipid peroxidation assay, PSEN1 and BACE1 expression by Real-Time Polymerase Chain Reaction (PCR), amyloid deposition by ELISA assays and immunohistochemistry. We found that SAM + SOD supplementation prevents the exacerbation of AD-like features induced by B vitamin deficiency, showing synergistic effects compared to either SAM or SOD alone. SAM + SOD supplementation also contrasts the amyloid deposition typically observed in TgCRND8 mice. Although the mechanisms underlying the beneficial effect of exogenous SOD remain to be elucidated, our findings identify that the combination of SAM + SOD could be carefully considered as co-adjuvant of current AD therapies.

5.
Adv Exp Med Biol ; 975 Pt 1: 535-549, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28849480

RESUMO

There is an increasing interest for analytical methods aimed to detect biological sulfur-containing amines, because of their involvement in human diseases and metabolic disorders. This work describes an improved HPLC method for the determination of sulfur containing amino acids and amines from different biological matrices. We optimized a pre-column derivatization procedure using dabsyl chloride, in which dabsylated products can be monitored spectrophotometrically at 460 nm. This method allows the simultaneous analysis of biogenic amines, amino acids and sulfo-amino compounds including carnosine, dopamine, epinephrine, glutathione, cysteine, taurine, lanthionine, and cystathionine in brain specimens, urines, plasma, and cell lysates. Moreover, the method is suitable for the study of physiological and non-physiological derivatives of taurine and glutathione such as hypotaurine, homotaurine, homocysteic acid and S-acetylglutathione. The present method displays good efficiency of derivatization, having the advantage to give rise to stable products compared to other derivatizing agents such as o-phthalaldehyde and dansyl chloride.With this method, we provide a tool to study sulfur cycle from a metabolic point of view in relation to the pattern of biological amino-compounds, allowing researchers to get a complete scenario of organic sulfur and amino metabolism in tissues and cells.


Assuntos
Aminoácidos/análise , Aminas Biogênicas/análise , Cromatografia Líquida de Alta Pressão/métodos , Compostos de Enxofre/análise , Animais , Humanos , Camundongos
6.
Curr Alzheimer Res ; 14(7): 753-759, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-28176663

RESUMO

BACKGROUND: The GSK3ß has been associated to pathological functions in neurodegenerative diseases. This kinase is involved in hyperphosphorylation of microtubule-associated tau protein, leading to aggregation andformation of NFTs. It has clearly been shown that GSK3ß is regulated at posttranslational level: phosphorylation at Tyr216 activates kinase, while phosphorylation at Ser9 is essential to inhibit its activity. OBJECTIVES: At present, there are contradictory findings about the possibility that GSK3ß may be regulated at gene level. Previous data showed overexpression of GSK3ß mRNA in hypomethylating conditions, pointing out to the existence of epigenetic mechanisms responsible for GSK3ß gene regulation. Analysis of human GSK3ß promoter through bisulphite modification, both in neuroblastoma cells and in postmortem frontal cortex from AD patients (AD patients both at Braak stages I-II and at stages V-VI) , allowed us to characterize the methylation pattern of a putative CpG islands in human GSK3ß 5'- flanking region. RESULTS: The analysis evidenced overall hypomethylation of CpG and non-CpG cytosine residues both in cells and in human brain (AD patients and control subjects). We found that GSK3ß mRNA was overexpressed only in patients with initial AD, with no effect on the levels of the protein. On the other hand, we unexpectedly observed the decrease of the inactive GSK3ß in cortex from AD patients at Braak stages I-II, whereas considerable increase was observed in AD patients at stages V-VI compared to the control subjects. CONCLUSIONS: These results point out that GSK3ß hyperactivity, and then NFTs formation, could come into function at an early stage of the disease and then turn off at the last stages.


Assuntos
Doença de Alzheimer/patologia , Metilação de DNA/fisiologia , Lobo Frontal/enzimologia , Glicogênio Sintase Quinase 3 beta/genética , Proteínas 14-3-3/metabolismo , Idoso , Idoso de 80 Anos ou mais , Análise de Variância , Linhagem Celular Tumoral , Feminino , Glicogênio Sintase Quinase 3 beta/metabolismo , Humanos , Masculino , Pessoa de Meia-Idade , Neuroblastoma/patologia , Emaranhados Neurofibrilares/patologia , Fosforilação , Regiões Promotoras Genéticas/genética , RNA Mensageiro/metabolismo , Serina/metabolismo
7.
J Neuropathol Exp Neurol ; 76(1): 27-31, 2017 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-28053004

RESUMO

By means of functional genomics analysis, we recently described the mRNA expression profiles of various genes involved in the neuroinflammatory response in the brains of subjects with late-onset Alzheimer Disease (LOAD). Some of these genes, namely interleukin (IL)-1ß and IL-6, showed distinct expression profiles with peak expression during the first stages of the disease and control-like levels at later stages. IL-1ß and IL-6 genes are modulated by DNA methylation in different chronic and degenerative diseases; it is also well known that LOAD may have an epigenetic basis. Indeed, we and others have previously reported gene-specific DNA methylation alterations in LOAD and in related animal models. Based on these data, we studied the DNA methylation profiles, at single cytosine resolution, of IL-1ß and IL-6 5'-flanking region by bisulphite modification in the cortex of healthy controls and LOAD patients at 2 different disease stages: Braak I-II/A and Braak V-VI/C. Our analysis provides evidence that neuroinflammation in LOAD is associated with (and possibly mediated by) epigenetic modifications.


Assuntos
Doença de Alzheimer/metabolismo , Citocinas/metabolismo , Metilação de DNA/fisiologia , Perfilação da Expressão Gênica/métodos , Mediadores da Inflamação/metabolismo , Idoso , Idoso de 80 Anos ou mais , Doença de Alzheimer/genética , Doença de Alzheimer/patologia , Citocinas/genética , Feminino , Lobo Frontal/metabolismo , Lobo Frontal/patologia , Humanos , Masculino , Pessoa de Meia-Idade , Regiões Promotoras Genéticas/fisiologia
8.
J Alzheimers Dis ; 44(4): 1323-31, 2015.
Artigo em Inglês | MEDLINE | ID: mdl-25672765

RESUMO

Widely confirmed reports were published on association between hyperhomocysteinemia, B vitamin deficiency, oxidative stress, and amyloid-ß in Alzheimer's disease (AD). Homocysteine, cysteine, cysteinylglycine and glutathione are metabolically interrelated thiols that may be potential indicators of health status and disease risk; they all participate in the metabolic pathway of homocysteine. Previous data obtained in one of our laboratories showed that B vitamin deficiency induced exacerbation of AD-like features in TgCRND8 AD mice; these effects were counteracted by S-adenosylmethionine (SAM) supplementation, through the modulation of DNA methylation and antioxidant pathways. Since the cellular response to oxidative stress typically involves alteration in thiols content, a rapid and sensitive HPLC method with fluorescence detection was here used to evaluate the effect of SAM and superoxide-dismutase (SOD) supplementation on thiols level in plasma, in TgCRND8 mice. The quantitative data obtained from HPLC analysis of mice plasma samples showed significant decrease of thiols level when the B vitamin deficient diet was supplemented with SAM + SOD and SOD alone, the latter showing the greatest effect. All these considerations point out the measurement of plasma thiols concentration as a powerful tool of relevance for all clinical purposes involving the evaluation of oxidative stress. The coupling of HPLC with fluorimetric detection, here used, provided a strong method sensitivity allowing thiols determination at very low levels.


Assuntos
Doença de Alzheimer/sangue , Doença de Alzheimer/dietoterapia , Hiper-Homocisteinemia/induzido quimicamente , S-Adenosilmetionina/uso terapêutico , Compostos de Sulfidrila/sangue , Superóxido Dismutase/uso terapêutico , Doença de Alzheimer/genética , Precursor de Proteína beta-Amiloide/genética , Animais , Cromatografia , Cromatografia Líquida de Alta Pressão , Modelos Animais de Doenças , Glutationa/sangue , Homocisteína/sangue , Humanos , Hiper-Homocisteinemia/sangue , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Mutação/genética
9.
Neurobiol Aging ; 33(7): 1482.e1-16, 2012 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-22221883

RESUMO

Methylation reactions linked to homocysteine in the one-carbon metabolism are increasingly elicited in Alzheimer's disease, although the association of hyperhomocysteinemia and of low B vitamin levels with the disease is still debated. We previously demonstrated that hyperhomocysteinemia and DNA hypomethylation induced by B vitamin deficiency are associated with PSEN1 and BACE1 overexpression and amyloid production. The present study is aimed at assessing S-adenosylmethionine effects in mice kept under a condition of B vitamin deficiency. To this end, TgCRND8 mice and wild-type littermates were assigned to control or B vitamin deficient diet, with or without S-adenosylmethionine supplementation. We found that S-adenosylmethionine reduced amyloid production, increased spatial memory in TgCRND8 mice and inhibited the upregulation of B vitamin deficiency-induced PSEN1 and BACE1 expression and Tau phosphorylation in TgCRND8 and wild-type mice. Furthermore, S-adenosylmethionine treatment reduced plaque spreading independently on B vitamin deficiency. These results strengthen our previous observations on the possible role of one-carbon metabolism in Alzheimer's disease, highlighting hyperhomocysteinemia-related mechanisms in dementia onset/progression and encourage further studies aimed at evaluating the use of S-adenosylmethionine as a potential candidate drug for the treatment of the disease.


Assuntos
Doença de Alzheimer/tratamento farmacológico , Doença de Alzheimer/genética , Progressão da Doença , S-Adenosilmetionina/uso terapêutico , Deficiência de Vitaminas do Complexo B/tratamento farmacológico , Deficiência de Vitaminas do Complexo B/genética , Potenciais de Ação/fisiologia , Doença de Alzheimer/patologia , Precursor de Proteína beta-Amiloide/genética , Animais , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Presenilina-1/genética , Deficiência de Vitaminas do Complexo B/patologia
10.
Curr Alzheimer Res ; 9(5): 527-35, 2012 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-22272624

RESUMO

In recent years, in parallel with the growing awareness of the multifactorial nature of Late Onset Alzheimer's Disease, the possibility that epigenetic mechanisms could be involved in the onset and/or progression of the pathology assumed an increasingly intriguing and leading role in Alzheimer's research. Today, many scientific reports indicate the existence of an epigenetic drift during ageing, in particular in Alzheimer's subjects. At the same time, experimental evidences are provided with the aim to demonstrate the causative or consequential role of epigenetic mechanisms. Our research group was involved in the last ten years in studying DNA methylation, the main epigenetic modification, in relationship to altered one-carbon metabolism (namely high homocysteine and low B vitamins levels), in Alzheimer's experimental models. Our previous findings about the demethylation of Presenilin1 gene promoter in nutritionally-induced hyperhomocysteinemia in a transgenic mouse model clearly demonstrated that Presenilin1 is regulated by DNA methylation. One of the open questions raised by our studies was if the observed demethylation was solely due to the induced imbalance of one-carbon metabolism or could be a response to the massive deposition of amyloid plaques in transgenic mice. Here we analyzed old (10 months) mice under standard diet in order to evidence possible changes in Presenilin1 promoter methylation in transgenic (TgCRND8 mice, carrying a double-mutated human APP transgene) vs. wt mice (129Sv) after prolonged exposure to amyloid. We found no differences in Presenilin1 methylation despite a slight increase in gene expression; these results suggest that amyloid production is not responsible for Presenilin1 demethylation in TgCRND8 mice brain.


Assuntos
Envelhecimento/metabolismo , Doença de Alzheimer/metabolismo , Metilação de DNA , Presenilina-1/metabolismo , Regiões Promotoras Genéticas , Envelhecimento/genética , Doença de Alzheimer/complicações , Doença de Alzheimer/genética , Precursor de Proteína beta-Amiloide/genética , Precursor de Proteína beta-Amiloide/metabolismo , Animais , Encéfalo/metabolismo , Modelos Animais de Doenças , Epigênese Genética , Feminino , Regulação da Expressão Gênica/fisiologia , Deriva Genética , Humanos , Hiper-Homocisteinemia/complicações , Hiper-Homocisteinemia/metabolismo , Masculino , Camundongos , Camundongos da Linhagem 129 , Camundongos Transgênicos , Presenilina-1/genética
11.
Neurobiol Aging ; 32(2): 187-99, 2011 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-19329227

RESUMO

We have previously shown that a nutritional model of B vitamin deficiency and homocysteine cycle alteration could lead to increased amyloid ß deposition, due to PSEN1 and BACE over-expression and consequent increase in secretase activity. We hypothesize that nutritional factors causing homocysteine cycle alterations (i.e. hyperhomocysteinemia) could induce sequence-specific DNA hypomethylation and "aberrant" gene activation. Aim of present study was to analyze the methylation pattern of PSEN1 promoter in SK-N-BE neuroblastoma cells and TgCRND8 mice, in a B vitamin (folate, B12 and B6) deficiency paradigm. PSEN1 methylation status has been evaluated through bisulphite modification and genomic sequencing. We demonstrate that B vitamin deficiency induces hypomethylation of specific CpG moieties in the 5'-flanking region; S-adenosylmethionine has been supplemented as methyl donor to reverse this effect. PSEN1 promoter methylation status is correlated with gene expression. These findings pinpoint a direct relationship between B vitamin-dependent alteration of homocysteine cycle and DNA methylation and also indicate that PSEN1 promoter is regulated by methylation of specific CpG moieties.


Assuntos
Metilação de DNA/fisiologia , Regulação da Expressão Gênica/fisiologia , Presenilina-1/genética , S-Adenosilmetionina/efeitos adversos , Deficiência de Vitaminas do Complexo B/etiologia , Deficiência de Vitaminas do Complexo B/metabolismo , Precursor de Proteína beta-Amiloide/genética , Análise de Variância , Animais , Encéfalo/metabolismo , Linhagem Celular Tumoral , Metilação de DNA/efeitos dos fármacos , Modelos Animais de Doenças , Regulação da Expressão Gênica/genética , Humanos , Camundongos , Camundongos Transgênicos , Mutação/genética , Presenilina-1/metabolismo , Regiões Promotoras Genéticas/efeitos dos fármacos , Regiões Promotoras Genéticas/genética , RNA Mensageiro/metabolismo , Análise de Sequência de DNA , Sulfitos/farmacologia , Transfecção/métodos , Deficiência de Vitaminas do Complexo B/genética , Deficiência de Vitaminas do Complexo B/patologia
12.
J Nutr Biochem ; 22(3): 242-51, 2011 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-20573497

RESUMO

Late-onset Alzheimer's disease seems to be a multi-factorial disease with both genetic and non-genetic, environmental, possible causes. Recently, epigenomics is achieving a major role in Alzheimer's research due to its involvement in different molecular pathways leading to neurodegeneration. Among the different epigenetic modifications, DNA methylation is one of the most relevant to the disease. We previously demonstrated that presenilin1 (PSEN1), a gene involved in amyloidogenesis, is modulated by DNA methylation in neuroblastoma cells and Alzheimer's mice in an experimental model of nutritionally altered one-carbon metabolism. This alteration, obtained by nutritional deficiency of B vitamins (folate, B12 and B6) hampered S-adenosylmethionine (SAM)-dependent methylation reactions. The aim of the present paper was to investigate the regulation of DNA methylation machinery in response to hypomethylating (B vitamin deficiency) and hypermethylating (SAM supplementation) alterations of the one-carbon metabolism. We found that DNA methylases (DNMT1, 3a and 3b) and a putative demethylase (MBD2) were differently modulated, in line with the previously observed changes of PSEN1 methylation pattern in the same experimental conditions.


Assuntos
Doença de Alzheimer/metabolismo , Carbono/metabolismo , DNA (Citosina-5-)-Metiltransferases/metabolismo , Metilação de DNA , Proteínas de Ligação a DNA/metabolismo , Modelos Animais de Doenças , Análise de Variância , Animais , Linhagem Celular , Epigenômica , Feminino , Ácido Fólico/metabolismo , Humanos , Masculino , Metilação , Camundongos , Camundongos da Linhagem 129 , Camundongos Transgênicos , S-Adenosilmetionina/análise , S-Adenosilmetionina/metabolismo , Vitamina B 12/metabolismo , Vitamina B 6/metabolismo , Complexo Vitamínico B/metabolismo , Complexo Vitamínico B/uso terapêutico , Deficiência de Vitaminas do Complexo B/metabolismo
13.
J Alzheimers Dis ; 22(4): 1257-68, 2010.
Artigo em Inglês | MEDLINE | ID: mdl-20930296

RESUMO

Late Onset Alzheimer's Disease (LOAD) can be associated to high homocysteine level and alteration of one-carbon metabolism. We previously demonstrated in the TgCRND8 mice strain, over-expressing human amyloid-ß protein precursor, that B vitamin deficiency causes alteration of one-carbon metabolism, together with unbalance of S-adenosylmethionine/S-adenosylhomocysteine levels, and is associated with AD like hallmarks as increased amyloid-ß plaque deposition, hyperhomocysteinemia, and oxidative stress. The same model of nutritional deficit was used here to study the variation of the brain protein expression profile associated to B vitamin deficiency. A group of proteins mainly involved in neuronal plasticity and mitochondrial functions was identified as modulated by one-carbon metabolism. These findings are consistent with increasing data about the pivotal role of mitochondrial abnormalities in AD patho-physiology. The identified proteins might represent new potential biomarkers of LOAD to be further investigated.


Assuntos
Doença de Alzheimer/metabolismo , Encéfalo/metabolismo , Carbono/metabolismo , Proteoma/metabolismo , Doença de Alzheimer/genética , Precursor de Proteína beta-Amiloide/genética , Precursor de Proteína beta-Amiloide/metabolismo , Análise de Variância , Animais , Análise por Conglomerados , Dieta , Modelos Animais de Doenças , Eletroforese em Gel Bidimensional , Feminino , Masculino , Camundongos , Camundongos Transgênicos , Neurônios/metabolismo , Estresse Oxidativo/fisiologia , Deficiência de Vitaminas do Complexo B/genética , Deficiência de Vitaminas do Complexo B/metabolismo
14.
J Alzheimers Dis ; 20(4): 997-1002, 2010.
Artigo em Inglês | MEDLINE | ID: mdl-20413874

RESUMO

Oxidative stress, altered glutathione levels, and hyperhomocysteinemia play critical roles in Alzheimer's disease. We studied the relationships between hyperhomocysteinemia, glutathione, and oxidative stress in TgCRND8 mice maintained in conditions of folate, B12, and B6 deficiency and the effect of S-adenosylmethionine supplementation. We found that hyperhomocysteinemia was correlated with increased reduced/oxidized brain glutathione ratio, with decreased glutathione S-transferase activity and increased lipid peroxidation. S-adenosylmethionine potentiated superoxide dismutase and glutathione S-transferase activity and restored altered brain glutathione and erythrocytes lipid peroxidation. These results underline the importance of S-adenosylmethionine as neuroprotective compound, acting both on methylation and oxidation metabolism.


Assuntos
Glutationa/metabolismo , Estresse Oxidativo/efeitos dos fármacos , S-Adenosilmetionina/farmacologia , Deficiência de Vitaminas do Complexo B/genética , Deficiência de Vitaminas do Complexo B/metabolismo , Animais , Dieta , Suplementos Nutricionais , Eritrócitos/efeitos dos fármacos , Eritrócitos/metabolismo , Glutationa Transferase/metabolismo , Homocisteína/metabolismo , Peroxidação de Lipídeos/efeitos dos fármacos , Metilação , Camundongos , Superóxido Dismutase/metabolismo
15.
J Alzheimers Dis ; 19(3): 895-907, 2010.
Artigo em Inglês | MEDLINE | ID: mdl-20157245

RESUMO

Neurofibrillary tangles (NFTs), composed of intracellular filamentous aggregates of hyperphosphorylated protein tau, are one of the pathological hallmarks of Alzheimer's disease (AD). Tau phosphorylation is regulated by the equilibrium between activities of its protein kinases and phosphatases; unbalance of these activities is proposed to be a reasonable causative factor to the disease process. Glycogen synthase kinase 3beta (GSK3beta) is one of the most important protein kinase in regulating tau phosphorylation; overexpression of active GSK3beta causes ADlike hyperphosphorylation of tau. Protein phosphatase 2A (PP2A) is the major phosphatase that dephosphorylates tau; it was demonstrated that highly conserved carboxyl-terminal sequence of PP2A C-subunit is a focal point for phosphatase regulation. This is the site of a reversible methyl esterification reaction that controls AB_{alpha}C heterotrimers formation. Here we demonstrate that GSK3beta and PP2A genes were upregulated by inhibiting methylation reactions through B vitamin deficiency. In this condition, methylated catalytic subunit PP2Ac was decreased, leading to reduced PP2A activity. By contrast, we observed GSK3beta protein increase and a modulation in phosphorylation sites that regulate GSK3beta activity. Therefore, one-carbon metabolism alteration seems to be a cause of deregulation of the equilibrium between GSK3beta and PP2A, leading to abnormal hyperphosphorylated tau.


Assuntos
Doença de Alzheimer , Quinase 3 da Glicogênio Sintase/genética , Fosforilação/fisiologia , Proteína Fosfatase 2/antagonistas & inibidores , Proteína Fosfatase 2/genética , Deficiência de Vitaminas do Complexo B/fisiopatologia , Proteínas tau/metabolismo , Doença de Alzheimer/genética , Doença de Alzheimer/metabolismo , Doença de Alzheimer/fisiopatologia , Animais , Western Blotting , Linhagem Celular Tumoral , Primers do DNA/genética , Glicogênio Sintase Quinase 3 beta , Humanos , Imuno-Histoquímica , Camundongos , Degeneração Neural/etiologia , Degeneração Neural/patologia , Neuroblastoma/metabolismo , Neuroblastoma/patologia , Emaranhados Neurofibrilares/metabolismo , Emaranhados Neurofibrilares/patologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa
16.
Mol Cell Neurosci ; 37(4): 731-46, 2008 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-18243734

RESUMO

Etiological and molecular studies on the sporadic form of Alzheimer's disease have yet to determine the underlying mechanisms of neurodegeneration. Hyperhomocysteinemia is associated with Alzheimer's disease, and has been hypothesized to promote neurodegeneration, by inhibiting brain methylation activity. The aim of this work was to determine whether a combined folate, B12 and B6 dietary deficiency, would induce amyloid-beta overproduction, and to study the mechanisms linking vitamin deficiency, hyperhomocysteinemia and amyloidogenesis in TgCRND8 and 129Sv mice. We confirmed that B-vitamin deprivation induces hyperhomocysteinemia and imbalance of S-adenosylmethionine and S-adenosylhomocysteine. This effect was associated with PS1 and BACE up-regulation and amyloid-beta deposition. Finally, we detected intraneuronal amyloid-beta and a slight cognitive impairment in a water maze task at a pre-plaque age, supporting the hypothesis of early pathological function of intracellular amyloid. Collectively, these findings are consistent with the hypothesis that abnormal methylation in association with hyperhomocysteinemia may contribute to Alzheimer's disease.


Assuntos
Secretases da Proteína Precursora do Amiloide/biossíntese , Peptídeos beta-Amiloides/metabolismo , Ácido Aspártico Endopeptidases/biossíntese , Hiper-Homocisteinemia/etiologia , Presenilina-1/biossíntese , S-Adenosil-Homocisteína/metabolismo , S-Adenosilmetionina/deficiência , Deficiência de Vitaminas do Complexo B/metabolismo , Secretases da Proteína Precursora do Amiloide/genética , Animais , Ácido Aspártico Endopeptidases/genética , Encéfalo/metabolismo , Encéfalo/patologia , Regulação da Expressão Gênica/fisiologia , Hiper-Homocisteinemia/genética , Hiper-Homocisteinemia/metabolismo , Masculino , Camundongos , Camundongos Transgênicos , Presenilina-1/genética , S-Adenosilmetionina/genética , Deficiência de Vitaminas do Complexo B/complicações , Deficiência de Vitaminas do Complexo B/genética
17.
J Alzheimers Dis ; 11(3): 275-90, 2007 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-17851177

RESUMO

Multiple aspects of homocysteine metabolism were studied to understand the mechanism responsible for hyperhomocysteinemia toxicity in Alzheimer disease. Besides oxidative stress and vascular damage, homocysteine has also a great importance in regulating DNA methylation through S-adenosylmethionine, the main methyl donor in eukaryotes. Alterations of S-adenosylmethionine and methylation were evidenced in Alzheimer disease and in elderly. In order to clarify whether DNA methylation can provide the basis for amyloid-beta overproduction, we used human SK-N-BE neuroblastoma and A172 glioblastoma cell lines. We tested the effects of folate, B12 and B6 deprivation and S-adenosylmethionine addition on methylation metabolism. Our results indicate that homocysteine accumulation induced through vitamin B deprivation could impair the "Methylation Potential" with consequent presenilin 1, BACE and amyloid-beta upregulation. Moreover, we found that homocysteine alterations had an effect on neuroblastoma but not on glioblastoma cells; this suggests a possible differential role of the two cell types in Alzheimer disease.


Assuntos
Secretases da Proteína Precursora do Amiloide/metabolismo , Glioblastoma/metabolismo , Glioblastoma/patologia , Homocisteína/metabolismo , Neuroblastoma/metabolismo , Neuroblastoma/patologia , Idoso , Doença de Alzheimer/genética , Doença de Alzheimer/metabolismo , Doença de Alzheimer/patologia , Secretases da Proteína Precursora do Amiloide/genética , Precursor de Proteína beta-Amiloide/metabolismo , Antioxidantes/administração & dosagem , Antioxidantes/farmacologia , Ácido Aspártico Endopeptidases/genética , Ácido Aspártico Endopeptidases/metabolismo , Western Blotting , Linhagem Celular Tumoral/patologia , Metilação de DNA/efeitos dos fármacos , Primers do DNA/genética , Ácido Fólico/administração & dosagem , Ácido Fólico/farmacologia , Humanos , Estresse Oxidativo/fisiologia , Reação em Cadeia da Polimerase , Presenilina-1/genética , Presenilina-1/metabolismo , S-Adenosilmetionina/administração & dosagem , S-Adenosilmetionina/farmacologia , Regulação para Cima , Deficiência de Vitamina B 12/metabolismo , Deficiência de Vitamina B 6/metabolismo
18.
J Alzheimers Dis ; 9(4): 407-14, 2006 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-16917149

RESUMO

Alzheimer disease (AD) is among the few diseases that may display high homocysteine (HCY) and low B12 and folate in blood. This observation has raised the suspect that amyloid-beta overproduction and accumulation, which may be the cause of the disease, could be due to the loss of epigenetic control in the expression of the genes involved in AbetaPP (amyloid-beta protein precursor) processing. We have shown, in cell culture, that two of the genes responsible for amyloid-beta production are controlled by the methylation of their promoters. The process is strictly related to S-adenosylmethionine (SAM) metabolism. SAM is a natural compound, mainly produced by the liver, which has been found at very low concentrations in AD brains. A further support to this thesis came from the observation that in elderly DNA methylations are consistently lower than in young and mid aged people. We are actually experimenting in transgenic mice the possibility to prevent or to arrest amyloid-beta accumulation, through SAM administration, and therefore its significance and the use of this drug for the treatment of the disease.


Assuntos
Doença de Alzheimer/genética , Doença de Alzheimer/terapia , Metilação de DNA , Epigênese Genética/genética , Inativação Gênica/fisiologia , S-Adenosilmetionina , Envelhecimento , Doença de Alzheimer/metabolismo , Peptídeos beta-Amiloides/metabolismo , Encéfalo/efeitos dos fármacos , Encéfalo/metabolismo , Metilação de DNA/efeitos dos fármacos , Inativação Gênica/efeitos dos fármacos , Humanos , S-Adenosilmetionina/metabolismo , S-Adenosilmetionina/farmacologia , S-Adenosilmetionina/uso terapêutico
19.
J Alzheimers Dis ; 9(4): 415-9, 2006 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-16917150

RESUMO

High homocysteine (Hcy) together with low S-adenosylmethionine (SAM) levels are often observed in Alzheimer disease (AD), and this could be a sign of alteration of SAM/Hcy metabolism. It has already been shown that DNA methylation is involved in amyloid-beta-protein precursor (AbetaPP) processing and amyloid-beta(Abeta) production through the regulation of Presenilin 1 (PS1) expression and that exogenous SAM can silence the gene reducing Abeta. To investigate whether SAM administration globally influenced gene expression in the brain, we analysed 588 genes of the central nervous system in SK-N-BE neuroblastoma cells, with cDNA probes derived from untreated (DM; Differentiation Medium) or SAM treated (DM+SAM) cultures. In these conditions only seven genes were modulated by SAM treatment (and therefore by DNA methylation); three were up-regulated and four down-regulated, showing low levels of modulation.


Assuntos
Doença de Alzheimer/tratamento farmacológico , Doença de Alzheimer/genética , Expressão Gênica/efeitos dos fármacos , Análise de Sequência com Séries de Oligonucleotídeos/métodos , S-Adenosilmetionina/farmacologia , S-Adenosilmetionina/uso terapêutico , Envelhecimento , Precursor de Proteína beta-Amiloide/efeitos dos fármacos , Linhagem Celular Tumoral/patologia , Cromatografia Líquida de Alta Pressão , Metilação de DNA/efeitos dos fármacos , Humanos , Proteínas de Membrana/efeitos dos fármacos , Proteínas de Membrana/metabolismo , Neuroblastoma/metabolismo , Neuroblastoma/patologia , Presenilina-1 , RNA/efeitos dos fármacos , RNA/genética , RNA/metabolismo , S-Adenosilmetionina/administração & dosagem
20.
J Pharm Biomed Anal ; 38(3): 449-56, 2005 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-15925246

RESUMO

Capillary electrophoresis was used for monitoring the stability of S-adenosylmethionine in aqueous solution under different conditions of storage and incubation used for "in vitro" and "in vivo" experiments, by evaluating both the entity of degradation and the possibility of epimerization at the sulfonium group. The determination of S,S-S-adenosylmethionine in presence of its R,S-epimer and degradation products was performed in uncoated capillary of 50 microm ID using 150 mM sodium phosphate buffer at pH 2.5. The analyses were performed in short or long-end injection modes depending if a fast monitoring of the degradation products or the evaluation of the diastereoisomeric ratio were carried out, respectively. In the long-end injection mode the baseline separation of S-adenosylmethionine diastereoisomeric forms and degradation products was obtained in less than 10 min with efficiency values in the range of 172,520-311,439 number of theoretical plates per meter. The results showed that freezing was the optimum storage mode for S-adenosylmethionine aqueous solutions preserving from degradation and diastereoisomeric ratio alterations. Under incubation conditions at 38 degrees C during 14 days period S-adenosylmethionine showed a strong degradation and the formation of three main increasing degradation products. After 7 and 14 days only the 52% and 32% of the initial drug concentration were available and the active S,S-S-adenosylmethionine form was the most affected.


Assuntos
Estabilidade de Medicamentos , Eletroforese Capilar/métodos , S-Adenosilmetionina/análise , Cromatografia Líquida de Alta Pressão/métodos , Armazenamento de Medicamentos/métodos , Eletroforese Capilar/instrumentação , Reprodutibilidade dos Testes , S-Adenosilmetionina/química , Soluções/química , Estereoisomerismo , Tecnologia Farmacêutica/métodos , Temperatura , Fatores de Tempo , Água/química
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