RESUMO
INTRODUCTION: Schizophrenia is considered one of the most disabling and severe human diseases worldwide. The etiology of schizophrenia is thought to be multifactorial and evidence suggests that DNA methylation can play an important role in underlying pivotal neurobiological alterations of this disorder. Some studies have demonstrated the effects of dietary supplementation as an alternative approach to the prevention of schizophrenia, including folic acid. However, no study has ever investigated the role of such supplementation in altering the DNA methylation system in the context of schizophrenia. OBJECTIVES: The present study aims to investigate the effects of maternal folic acid supplementation at different doses on nuclear methyltransferase activity of adult rat offspring subjected to an animal model schizophrenia induced by ketamine. METHODS: Adult female Wistar rats, (60 days old) received folic acid-deficient diet, control diet, or control diet plus folic acid supplementation (at 5, 10, or 50 mg/kg) during pregnancy and lactation. After reaching adulthood (60 days), the male offspring of these dams were subjected to the animal model of schizophrenia induced by 7 days of ketamine intraperitoneal injection (25 mg/kg). After the 7-day protocol, the activity of nuclear methyltransferase was evaluated in the brains of the offspring. RESULTS: Maternal folic acid supplementation at 50 mg/kg increased methyltransferase activity in the frontal cortex, while 10 mg/kg increased methyltransferase activity in the hippocampus. In the striatum of offspring treated with ketamine, maternal deficient diet, control diet, and folic acid supplementation at 5 mg/kg decreased methyltransferase activity compared to the control group. The folic acid supplementation at 10 and 50 mg/kg reversed this ketamine effect. CONCLUSIONS: Maternal FA deficiency could be related to schizophrenia pathophysiology, while FA supplementation could present a protective effect since it demonstrated persistent effects in epigenetic parameters in adult offspring.
Assuntos
Núcleo Celular/enzimologia , Ácido Fólico/uso terapêutico , Metiltransferases/metabolismo , Esquizofrenia/prevenção & controle , Animais , Núcleo Celular/efeitos dos fármacos , Metilação de DNA/efeitos dos fármacos , Dieta , Suplementos Nutricionais , Feminino , Deficiência de Ácido Fólico/complicações , Ketamina , Masculino , Gravidez , Ratos , Ratos Wistar , Esquizofrenia/induzido quimicamente , Esquizofrenia/enzimologia , Psicologia do EsquizofrênicoRESUMO
KEY MESSAGE: We describe the potato CDPK family and place StCDPK7 as a player in potato response to Phytophthora infestans infection, identifying phenylalanine ammonia lyase as its specific phosphorylation target in vitro. Calcium-dependent protein kinases (CDPKs) decode calcium (Ca2+) signals and activate different signaling pathways involved in hormone signaling, plant growth, development, and both abiotic and biotic stress responses. In this study, we describe the potato CDPK/CRK multigene family; bioinformatic analysis allowed us to identify 20 new CDPK isoforms, three CDPK-related kinases (CRKs), and a CDPK-like kinase. Phylogenetic analysis indicated that 26 StCDPKs can be classified into four groups, whose members are predicted to undergo different acylation patterns and exhibited diverse expression levels in different tissues and in response to various stimuli. With the aim of characterizing those members that are particularly involved in plant-pathogen interaction, we focused on StCDPK7. Tissue expression profile revealed that StCDPK7 transcript levels are high in swollen stolons, roots, and mini tubers. Moreover, its expression is induced upon Phytophthora infestans infection in systemic leaves. Transient expression assays showed that StCDPK7 displays a cytosolic/nuclear localization in spite of having a predicted chloroplast transit peptide. The recombinant protein, StCDPK7:6xHis, is an active Ca2+-dependent protein kinase that can phosphorylate phenylalanine ammonia lyase, an enzyme involved in plant defense response. The analysis of the potato CDPK family provides the first step towards the identification of CDPK isoforms involved in biotic stress. StCDPK7 emerges as a relevant player that could be manipulated to deploy disease resistance in potato crops.
Assuntos
Phytophthora infestans/fisiologia , Proteínas de Plantas/genética , Proteínas Quinases/genética , Proteínas Quinases/metabolismo , Solanum tuberosum/genética , Solanum tuberosum/parasitologia , Núcleo Celular/enzimologia , Núcleo Celular/metabolismo , Citosol/enzimologia , Citosol/metabolismo , Resistência à Doença/genética , Fenilalanina Amônia-Liase/genética , Fenilalanina Amônia-Liase/metabolismo , Doenças das Plantas/genética , Doenças das Plantas/microbiologia , Proteínas de Plantas/metabolismo , Solanum tuberosum/enzimologiaRESUMO
The cell cycle-associated neuronal death hypothesis, which has been proposed as a common mechanism for most neurodegenerative diseases, is notably supported by evidencing cell cycle effectors in neurons. However, in naturally occurring nervous system diseases, these markers are not expressed in neuron nuclei but in cytoplasmic compartments. In other respects, the Feline Panleukopenia Virus (FPV) is able to complete its cycle in mature brain neurons in the feline species. As a parvovirus, the FPV is strictly dependent on its host cell reaching the cell cycle S phase to start its multiplication. In this retrospective study on the whole brain of 12 cats with naturally-occurring, FPV-associated cerebellar atrophy, VP2 capsid protein expression was detected by immunostaining not only in some brain neuronal nuclei but also in neuronal cytoplasm in 2 cats, suggesting that viral mRNA translation was still occurring. In these cats, double immunostainings demonstrated the expression of cell cycle S phase markers cyclin A, cdk2 and PCNA in neuronal nuclei. Parvoviruses are able to maintain their host cells in S phase by triggering the DNA damage response. S139 phospho H2A1, a key player in the cell cycle arrest, was detected in some neuronal nuclei, supporting that infected neurons were also blocked into the S phase. PCR studies did not support a co-infection with an adeno or herpes virus. ERK1/2 nuclear accumulation was observed in some neurons suggesting that the ERK signaling pathway might be involved as a mechanism driving these neurons far into the cell cycle.
Assuntos
Biomarcadores/metabolismo , Núcleo Celular/metabolismo , Cérebro/patologia , Vírus da Panleucopenia Felina/fisiologia , Panleucopenia Felina/patologia , Panleucopenia Felina/virologia , Neurônios/patologia , Fase S , Animais , Anticorpos Antivirais/metabolismo , Especificidade de Anticorpos , Pareamento de Bases , Proteínas do Capsídeo/metabolismo , Gatos , Núcleo Celular/enzimologia , DNA Viral/metabolismo , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Vírus da Panleucopenia Felina/genética , Feminino , Genes Virais , Células HEK293 , Humanos , Imuno-Histoquímica , Masculino , Neurônios/virologia , Reação em Cadeia da Polimerase , Reprodutibilidade dos Testes , Tálamo/metabolismoRESUMO
Ubiquitination regulates many processes in plants, including immunity. The E3 ubiquitin ligase PUB17 is a positive regulator of programmed cell death (PCD) triggered by resistance proteins CF4/9 in tomato. Its role in immunity to the potato late blight pathogen, Phytophthora infestans, was investigated here. Silencing StPUB17 in potato by RNAi and NbPUB17 in Nicotiana benthamiana by virus-induced gene silencing (VIGS) each enhanced P. infestans leaf colonization. PAMP-triggered immunity (PTI) transcriptional responses activated by flg22, and CF4/Avr4-mediated PCD were attenuated by silencing PUB17. However, silencing PUB17 did not compromise PCD triggered by P. infestans PAMP INF1, or co-expression of R3a/AVR3a, demonstrating that not all PTI- and PCD-associated responses require PUB17. PUB17 localizes to the plant nucleus and especially in the nucleolus. Transient over-expression of a dominant-negative StPUB17(V314I,V316I) mutant, which retained nucleolar localization, suppressed CF4-mediated cell death and enhanced P. infestans colonization. Exclusion of the StPUB17(V314I,V316I) mutant from the nucleus abolished its dominant-negative activity, demonstrating that StPUB17 functions in the nucleus. PUB17 is a positive regulator of immunity to late blight that acts in the nucleus to promote specific PTI and PCD pathways.
Assuntos
Phytophthora infestans/fisiologia , Doenças das Plantas/imunologia , Imunidade Vegetal , Solanum tuberosum/enzimologia , Ubiquitina-Proteína Ligases/genética , Apoptose , Núcleo Celular/enzimologia , Inativação Gênica , Mutação , Folhas de Planta/enzimologia , Folhas de Planta/genética , Folhas de Planta/imunologia , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Solanum tuberosum/genética , Solanum tuberosum/imunologia , Nicotiana/enzimologia , Nicotiana/genética , Nicotiana/imunologia , Ubiquitina-Proteína Ligases/metabolismoRESUMO
Chard is a plant used as an alternative hypoglycemic agent by diabetic people in Turkey. The aim of this study was to examine the molecular mechanism of hypoglycemic effects of chard extract. Male Sprague-Dawley rats (6-7 months old) were divided into five groups for this investigation: (1) control, (2) hyperglycemic, (3) hyperglycemic+chard, (4) hyperglycemic+insulin, (5) hyperglycemic+chard+insulin. Fourteen days after animals were rendered hyperglycemic by intraperitoneal injection of 60 mg/kg streptozotocin, the chard water extract (2 g/kg/day) or/and insulin (6 U/kg/day) was administered for 45 days. Hypoglycemic effect of chard extract was demonstrated by a significant reduction in the fasting blood glucose and increased glycogen levels in liver of chard extract-treated hyperglycemic rats. Moreover, activity of adenosine deaminase, which is suggested as an important enzyme for modulating the bioactivity of insulin, was decreased by chard treatment. Immunostaining analysis showed increased nuclear translocation of Akt2 and synthesis of GLUT2 in the hepatocytes of chard or/and insulin-treated hyperglycemic rats. The oxidative stress was decreased and antioxidant defense was increased by chard extract or/and insulin treatment to hyperglycemic rats according to the decreased malondialdehyde formation, the activities of catalase, superoxide dismutase, myeloperoxidase and increased glutathione levels. These findings suggest that chard extract might improve glucose response by increasing GLUT2 through Akt2 and antioxidant defense in the liver.
Assuntos
Beta vulgaris/química , Transportador de Glucose Tipo 2/metabolismo , Hiperglicemia/tratamento farmacológico , Hipoglicemiantes/farmacologia , Extratos Vegetais/farmacologia , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transporte Ativo do Núcleo Celular , Animais , Antioxidantes/metabolismo , Catalase/metabolismo , Núcleo Celular/enzimologia , Diabetes Mellitus Experimental/tratamento farmacológico , Avaliação Pré-Clínica de Medicamentos , Hipoglicemiantes/uso terapêutico , Fígado/efeitos dos fármacos , Fígado/enzimologia , Masculino , Estresse Oxidativo , Peroxidase/metabolismo , Extratos Vegetais/uso terapêutico , Ratos Sprague-Dawley , Estreptozocina , Superóxido Dismutase/metabolismoRESUMO
The mitochondrial cytochrome c oxidase (COX) catalyzes the reduction of oxygen to water playing a key role in the respiratory chain and ATP synthesis. The nucleus-encoded COX subunits do not participate in catalysis, but some are known to play a role in the expression, assembly and activity of the enzyme. Since hypoxia continuously affects the shrimp environment, it is important to study COX to understand their ability to deal with low oxygen levels. The goal of this research was to characterize the complementary DNA (cDNA) sequences of three nucleus-encoded subunits -coxIV, coxVa, and coxVb- and to evaluate the shrimp COX response to hypoxia by measuring their gene expression. The cDNA sequence of coxIV consisted of 532bp, which encodes a 17.47kDa protein, while coxVa cDNA consisted of 460bp and coded a protein of 17.11kDa, and the coxVb coding sequence consisted of 364bp encoding a 13.74kDa protein. Shrimp subunits do not have isoforms, and they are not differentially expressed during hypoxia, as observed in mammals. Coordinated changes were detected in the mRNA amounts of nuclear and mitochondrial subnits; these changes, at the transcriptional level, are suggested to be controlled through transcriptional factors Sp1 and NRF2.
Assuntos
DNA Complementar/genética , Complexo IV da Cadeia de Transporte de Elétrons/genética , Mitocôndrias/enzimologia , Subunidades Proteicas/genética , Trifosfato de Adenosina/biossíntese , Sequência de Aminoácidos , Animais , Núcleo Celular/enzimologia , Complexo IV da Cadeia de Transporte de Elétrons/metabolismo , Expressão Gênica , Hipóxia/genética , Hipóxia/metabolismo , Mitocôndrias/genética , Oxigênio/metabolismo , Penaeidae , FilogeniaRESUMO
BACKGROUND: Corticotropin-releasing hormone (CRH) plays an important role in regulating the mammalian stress response. Two of the most extensively studied neuronal populations that express CRH are in the hypothalamus and amygdala. Both regions are involved in the stress response, but the amygdala is also involved in mediating response to fear and anxiety. Given that both hypothalamus and amygdala have overlapping functions, but their CRH-expressing neurons may respond differently to a given perturbation, we sought to identify differentially expressed genes between two neuronal cell types, amygdalar AR-5 and hypothalamic IVB cells. Thus, we performed a microarray analysis. Our hypothesis was that we would identify differentially expressed transcription factors, coregulators and chromatin-modifying enzymes. RESULTS: A total of 31,042 genes were analyzed, 10,572 of which were consistently expressed in both cell lines at a 95% confidence level. Of the 10,572 genes, 2,320 genes in AR-5 were expressed at ≥ 2-fold relative to IVBs, 1,104 genes were expressed at ≥2-fold in IVB relative to AR-5 and 7,148 genes were expressed at similar levels between the two cell lines. The greatest difference was in six mitochondrial DNA-encoded genes, which were highly abundant in AR-5 relative to IVB cells. The relative abundance of these genes ranged from 413 to 885-fold according to the microarray results. Differential expression of these genes was verified by RTqPCR. The differentially expressed mitochondrial genes were cytochrome b (MT-CYB), cytochrome c oxidase subunit 1 and 2 (MT-CO1 and MT-CO2) and NADH-ubiquinone oxidoreductase chain 1, 2, and 3 (MT-ND1, MT-ND2, MT-ND3). CONCLUSION: As expected, the array revealed differential expression of transcription factors and coregulators; however the greatest difference between the two cell lines was in genes encoded by the mitochondrial genome. These genes were abundant in AR-5 relative to IVBs. At present, the reason for the marked difference is unclear. The cells may differ in mtDNA copy number, number of mitochondria, or regulation of the mitochondrial genome. The specific functions served by having such different levels of mitochondrial expression have not been determined. It is possible that the greater expression of the mitochondrial genes in the amygdalar cells reflects higher energy requirements than in the hypothalamic cell line.
Assuntos
Tonsila do Cerebelo/citologia , Núcleo Celular/genética , Hipotálamo/citologia , Mitocôndrias/genética , Transcriptoma , Animais , Linhagem Celular , Núcleo Celular/enzimologia , Núcleo Celular/metabolismo , Cromatina/genética , Cromatina/metabolismo , Hormônio Liberador da Corticotropina/genética , Humanos , Mitocôndrias/enzimologia , Mitocôndrias/metabolismo , Análise de Sequência com Séries de Oligonucleotídeos , Ratos , Fatores de Transcrição/metabolismoRESUMO
Phytopathogenic oomycetes, such as Phytophthora infestans, secrete an arsenal of effector proteins that modulate plant innate immunity to enable infection. We describe CRN8, a host-translocated effector of P. infestans that has kinase activity in planta. CRN8 is a modular protein of the CRN effector family. The C-terminus of CRN8 localizes to the host nucleus and triggers cell death when the protein is expressed in planta. Cell death induction by CRN8 is dependent on its localization to the plant nucleus, which requires a functional nuclear localization signal (NLS). The C-terminal sequence of CRN8 has similarity to a serine/threonine RD kinase domain. We demonstrated that CRN8 is a functional RD kinase and that its auto-phosphorylation is dependent on an intact catalytic site. Co-immunoprecipitation experiments revealed that CRN8 forms a dimer or multimer. Heterologous expression of CRN8 in planta resulted in enhanced virulence by P. infestans. In contrast, in planta expression of the dominant-negative CRN8(R469A;D470A) resulted in reduced P. infestans infection, further implicating CRN8 in virulence. Overall, our results indicate that similar to animal parasites, plant pathogens also translocate biochemically active kinase effectors inside host cells.
Assuntos
Núcleo Celular/enzimologia , Phytophthora infestans/patogenicidade , Doenças das Plantas/microbiologia , Multimerização Proteica , Proteínas Serina-Treonina Quinases/metabolismo , Solanum tuberosum/microbiologia , Núcleo Celular/genética , Phytophthora infestans/enzimologia , Phytophthora infestans/genética , Doenças das Plantas/genética , Doenças das Plantas/imunologia , Imunidade Vegetal , Proteínas Serina-Treonina Quinases/genética , Solanum tuberosum/imunologia , Nicotiana/metabolismo , Nicotiana/microbiologiaRESUMO
Apoptosis, the main type of programmed cell death, plays an essential role in a variety of biological events. Whereas "classical" apoptosis is dependent on caspase activation, caspase-independent death is increasingly recognized as an alternative pathway. To develop new anticancer agents, oleifolioside A was isolated from Dendropanax morbifera Leveille and the biochemical mechanisms of oleifolioside A-induced apoptosis in HeLa cells were investigated. Exposure to oleifolioside A resulted in caspase activation and typical features of apoptosis, although cell death was not prevented by caspase inhibition. Oleifolioside A treatment induced up-regulation of Bad, loss of mitochondrial membrane potential, nuclear relocation of mitochondrial factors, apoptosis-inducing factor (AIF), endonuclease G (EndoG), and apoptosis induction. This is the first report of anticancer activity of oleifolioside A, and nuclear translocation of AIF and EndoG in oleifolioside A-treated HeLa cells might represent an alternative death signaling pathway in the absence of caspase activity.
Assuntos
Fator de Indução de Apoptose/metabolismo , Apoptose/efeitos dos fármacos , Araliaceae/química , Núcleo Celular/metabolismo , Endodesoxirribonucleases/metabolismo , Extratos Vegetais/farmacologia , Saponinas/farmacologia , Neoplasias do Colo do Útero/metabolismo , Caspases/metabolismo , Núcleo Celular/enzimologia , Feminino , Células HeLa , Humanos , Transporte Proteico , Neoplasias do Colo do Útero/tratamento farmacológico , Neoplasias do Colo do Útero/enzimologia , Neoplasias do Colo do Útero/fisiopatologiaRESUMO
Activation-induced deaminase (AID) is an enzyme required for class switch recombination (CSR) and somatic hypermutation (SHM), processes that ensure antibody maturation and expression of different immunoglobulin isotypes. AID function is tightly regulated by tissue- and stage-specific expression, nuclear localization, and protein stability. Transcription factor YY1 is crucial for early B cell development, but its function at late B cell stages is unknown. Here, we show that YY1 conditional knockout in activated splenic B cells interferes with CSR. Knockout of YY1 did not affect B cell proliferation, transcription of the AID and IgM genes, or levels of various switch region germ line transcripts. However, we show that YY1 physically interacts with AID and controls the accumulation of nuclear AID, at least in part, by increasing nuclear AID stability. We show for the first time that YY1 plays a novel role in CSR and controls nuclear AID protein levels.
Assuntos
Linfócitos B/metabolismo , Citidina Desaminase/metabolismo , Switching de Imunoglobulina/genética , Fator de Transcrição YY1/metabolismo , Animais , Linfócitos B/imunologia , Linhagem Celular , Núcleo Celular/enzimologia , Citidina Desaminase/genética , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Ligação Proteica , Baço/imunologia , Fator de Transcrição YY1/genéticaRESUMO
The leaves of Eriobotrya japonica Lindl. have been widely used as a traditional medicine for the treatment of many diseases including gastroenteric disorders, diabetes mellitus, chronic bronchitis and asthma. In the present study, the anti-metastatic action of the EtOAc fraction of the leaves of E. japonica (LEJ) was investigated. LEJ showed potent inhibitory effects on MMP-2 and MMP-9 activities and expressions via down-regulation of NF-κB translocation to the nucleus in B16F10 cells. In addition, the cell migration and invasion were down-regulated by LEJ. LEJ also significantly suppressed lung metastasis in vivo. Moreover, we isolated the compounds ursolic acid and 2α-hydroxyursolic acid from LEJ and both compounds also significantly suppressed MMP-2 and MMP-9 activities, indicating that they are the active components of LEJ. The present results demonstrate that LEJ may be used as valuable antimetastatic agent for the treatment of cancer metastasis.
Assuntos
Antineoplásicos Fitogênicos/farmacologia , Eriobotrya/química , Extratos Vegetais/farmacologia , Animais , Antineoplásicos Fitogênicos/isolamento & purificação , Antineoplásicos Fitogênicos/uso terapêutico , Movimento Celular/efeitos dos fármacos , Núcleo Celular/efeitos dos fármacos , Núcleo Celular/enzimologia , Proliferação de Células/efeitos dos fármacos , Humanos , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/secundário , Masculino , Metaloproteinase 9 da Matriz , Inibidores de Metaloproteinases de Matriz , Melanoma Experimental/tratamento farmacológico , Melanoma Experimental/secundário , Camundongos , Camundongos Endogâmicos C57BL , Estrutura Molecular , NF-kappa B/metabolismo , Invasividade Neoplásica , Extratos Vegetais/isolamento & purificação , Extratos Vegetais/uso terapêutico , Folhas de Planta/química , Transporte Proteico , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
High fluence low-power laser irradiation (HF-LPLI) is a newly discovered stimulus through generating reactive oxygen species (ROS) to trigger cell apoptosis. Activation of glycogen synthase kinase 3ß (GSK3ß) is proved to be involved in intrinsic apoptotic pathways under various stimuli. However, whether the proapoptotic factor GSK3ß participates in HF-LPLI-induced apoptosis has not been elucidated. Therefore, in the present study, we investigated the involvement of GSK3ß in apoptosis under HF-LPLI treatment (120 J/cm2, 633 nm). We found that GSK3ß activation could promote HF-LPLI-induced apoptosis, which could be prevented by lithium chloride (a selective inhibitor of GSK3ß) exposure or by GSK3ß-KD (a dominant-negative GSK3ß) overexpression. We also found that the activation of GSK3ß by HF-LPLI was due to the inactivation of protein kinase B (Akt), a widely reported and important upstream negative regulator of GSK3ß, indicating the existence and inactivation of Akt/GSK3ß signaling pathway. Moreover, the inactivation of Akt/GSK3ß pathway depended on the fluence of HF-LPLI treatment. Furthermore, vitamin c, a ROS scavenger, completely prevented the inactivation of Akt/GSK3ß pathway, indicating ROS generation was crucial for the inactivation. In addition, GSK3ß promoted Bax activation by down-regulating Mcl-1 upon HF-LPLI treatment. Taken together, we have identified a new and important proapoptotic signaling pathway that is consisted of Akt/GSK3ß inactivation for HF-LPLI stimulation. Our research will extend the knowledge into the biological mechanisms induced by LPLI.
Assuntos
Apoptose/efeitos da radiação , Quinase 3 da Glicogênio Sintase/metabolismo , Luz , Terapia com Luz de Baixa Intensidade , Proteínas Proto-Oncogênicas c-akt/metabolismo , Transdução de Sinais , Animais , Linhagem Celular , Núcleo Celular/enzimologia , Núcleo Celular/efeitos da radiação , Relação Dose-Resposta à Radiação , Regulação para Baixo/efeitos da radiação , Ativação Enzimática/efeitos da radiação , Glicogênio Sintase Quinase 3 beta , Humanos , Modelos Biológicos , Proteína de Sequência 1 de Leucemia de Células Mieloides , Transporte Proteico/efeitos da radiação , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Proteína X Associada a bcl-2/metabolismoRESUMO
A rare sugar, D-psicose has progressively been evaluated as a unique metabolic regulator of glucose and lipid metabolism, and thus represents a promising compound for the treatment of type 2 diabetes mellitus (T2DM). The present study was undertaken to examine the underlying effector organs of D-psicose in lowering blood glucose and abdominal fat by exploiting a T2DM rat model, Otsuka Long-Evans Tokushima Fatty (OLETF) rats. Rats were fed 5% D-psicose or 5% D-glucose supplemented in drinking water, and only water in the control for 13 weeks and the protective effects were compared. A non-diabetic Long-Evans Tokushima Otsuka (LETO), fed with water served as a counter control of OLETF. After 13 weeks feeding, D-psicose treatment significantly reduced the increase in body weight and abdominal fat mass. Oral glucose tolerance test (OGTT) showed the reduced blood glucose and insulin levels suggesting the improvement of insulin resistance in OLETF rats. Oil-red-O staining elucidated that D-psicose significantly reduced lipid accumulation in the liver. Immunohistochemical analysis showed D-psicose induced glucokinase translocation from nucleus to cytoplasm of the liver which enhances glucokinase activity and subsequent synthesis of glycogen in the liver. D-psicose also protected the pathological change of the ß-cells of pancreatic islets. These data demonstrate that D-psicose controls blood glucose levels by reducing lipotoxicity in liver and by preserving pancreatic ß-cell function.
Assuntos
Glicemia/efeitos dos fármacos , Diabetes Mellitus Tipo 2/tratamento farmacológico , Frutose/uso terapêutico , Resistência à Insulina , Gordura Abdominal/efeitos dos fármacos , Gordura Abdominal/patologia , Animais , Peso Corporal/efeitos dos fármacos , Núcleo Celular/enzimologia , Citoplasma/enzimologia , Citoproteção , Ingestão de Alimentos/efeitos dos fármacos , Fígado Gorduroso/tratamento farmacológico , Fígado Gorduroso/patologia , Glucoquinase/metabolismo , Teste de Tolerância a Glucose , Homeostase/efeitos dos fármacos , Células Secretoras de Insulina/efeitos dos fármacos , Células Secretoras de Insulina/patologia , Metabolismo dos Lipídeos/efeitos dos fármacos , Masculino , Pâncreas/efeitos dos fármacos , Pâncreas/patologia , Transporte Proteico/efeitos dos fármacos , Ratos , Ratos Endogâmicos OLETFRESUMO
All tRNA(His) possess an essential extra G(-1) guanosine residue at their 5' end. In eukaryotes after standard processing by RNase P, G(-1) is added by a tRNA(His) guanylyl transferase. In prokaryotes, G(-1) is genome-encoded and retained during maturation. In plant mitochondria, although trnH genes possess a G(-1) we find here that both maturation pathways can be used. Indeed, tRNA(His) with or without a G(-1) are found in a plant mitochondrial tRNA fraction. Furthermore, a recombinant Arabidopsis mitochondrial RNase P can cleave tRNA(His) precursors at both positions G(+1) and G(-1). The G(-1) is essential for recognition by plant mitochondrial histidyl-tRNA synthetase. Whether, as shown in prokaryotes and eukaryotes, the presence of uncharged tRNA(His) without G(-1) has a function or not in plant mitochondrial gene regulation is an open question. We find that when a mutated version of a plant mitochondrial trnH gene containing no encoded extra G is introduced and expressed into isolated potato mitochondria, mature tRNA(His) with a G(-1) are recovered. This shows that a previously unreported tRNA(His) guanylyltransferase activity is present in plant mitochondria.
Assuntos
Mitocôndrias/genética , Processamento Pós-Transcricional do RNA , RNA de Plantas/metabolismo , RNA de Transferência de Histidina/metabolismo , RNA/metabolismo , Arabidopsis/enzimologia , Núcleo Celular/enzimologia , Mitocôndrias/enzimologia , Nucleotidiltransferases/análise , Nucleotidiltransferases/metabolismo , RNA/biossíntese , RNA/classificação , Precursores de RNA/metabolismo , RNA Mitocondrial , RNA de Plantas/biossíntese , RNA de Plantas/classificação , RNA de Transferência de Histidina/biossíntese , RNA de Transferência de Histidina/classificação , Ribonuclease P/metabolismo , Solanum tuberosum/enzimologia , Solanum tuberosum/genéticaRESUMO
Thimet oligopeptidase (EC 3.4.24.15; also called EP24.15 and TOP; referred to here as TOP) is a neuropeptidase involved in the regulation of several physiological functions including reproduction. Among its substrates is gonadotrophin-releasing hormone (GnRH), an important hypothalamic hormone that regulates the synthesis and release of oestradiol and facilitates female sexual behaviour. Using immunohistochemistry, we found that TOP is expressed in the nucleus of cells throughout the female mouse brain, and in high levels in steroid-sensitive regions of the hypothalamus, which is consistent with previous findings in male rats. Furthermore, dual-label immunofluorescence revealed that TOP and oestrogen receptor alpha (ERalpha) coexpress in several reproductively-relevant brain regions, including the medial preoptic area (mPOA), arcuate nucleus (ARC), ventrolateral portion of the ventromedial hypothalamic nucleus (VMNvl) and the midbrain central grey (MCG). Previous studies in rats have shown that oestradiol decreases hypothalamic TOP levels or activity, possibly potentiating the effects of GnRH. In the present study, analysis by immunohistochemistry revealed that oestradiol decreased TOP immunoreactivity in the VMNvl, whereas no differences were detected in the mPOA, ARC or median eminence. Overall, the present findings indicate that TOP is coexpressed with ERalpha, and oestradiol regulates TOP expression in a brain region-specific manner in female mice, providing neuroanatomical evidence that TOP may function in reproductive physiology and/or behaviour.
Assuntos
Núcleo Celular/enzimologia , Estradiol/metabolismo , Receptor alfa de Estrogênio/metabolismo , Hipotálamo/citologia , Metaloendopeptidases/metabolismo , Animais , Receptor alfa de Estrogênio/genética , Feminino , Hipotálamo/metabolismo , Masculino , Metaloendopeptidases/genética , Camundongos , Camundongos Endogâmicos C57BL , Ovariectomia , RatosRESUMO
AIM OF THE STUDY: To determine the inhibitory effect of tetramethylpyrazine (TMP) on lipopolysaccharide (LPS)-induced over-production of nitric oxide (NO) and inducible nitric oxide synthase (iNOS) in N9 microglial cells. MATERIALS AND METHODS: N9 cells were pretreated with vehicle or TMP and then exposed to LPS for the time indicated. Cell viability was determined by methylthiazoyltetrazolium (MTT) assay. Nitrite assay was performed by Griess reaction. Expression of iNOS mRNA was examined by RT-PCR. Protein levels of iNOS, p38 mitogen-activated protein kinase (MAPK), ERK1/2, JNK, phosphatidylinositol 3-kinase (PI3K) and Akt were determined by western blot analysis. Formation of reactive oxygen species (ROS) was evaluated by fluorescence image system. RESULTS: TMP inhibited LPS-induced over-production of NO and iNOS in N9 cells. TMP also inhibited the NF-kappaB translocation from cytoplasm into nucleus of N9 cells. In addition, TMP showed blocking effect on the phosphorylation of p38 MAPK, ERK1/2, JNK and Akt, but not PI3K. Further, TMP suppressed the formation of intracellular ROS in LPS-induced N9 cells. CONCLUSIONS: TMP inhibited production of NO and iNOS in LPS-induced N9 cells through blocking MAPK and PI3K/Akt activation and suppressing ROS production.
Assuntos
Sequestradores de Radicais Livres/farmacologia , Microglia/efeitos dos fármacos , Proteínas Quinases Ativadas por Mitógeno/antagonistas & inibidores , Óxido Nítrico Sintase Tipo II/antagonistas & inibidores , Óxido Nítrico/biossíntese , Proteína Oncogênica v-akt/antagonistas & inibidores , Inibidores de Fosfoinositídeo-3 Quinase , Pirazinas/farmacologia , Espécies Reativas de Oxigênio/antagonistas & inibidores , Animais , Western Blotting , Linhagem Celular , Núcleo Celular/efeitos dos fármacos , Núcleo Celular/enzimologia , Núcleo Celular/metabolismo , Sobrevivência Celular/efeitos dos fármacos , Citosol/efeitos dos fármacos , Citosol/enzimologia , Citosol/metabolismo , Relação Dose-Resposta a Droga , Sequestradores de Radicais Livres/isolamento & purificação , Lipopolissacarídeos/toxicidade , Camundongos , Microglia/enzimologia , Microglia/metabolismo , Óxido Nítrico/antagonistas & inibidores , Pirazinas/isolamento & purificação , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Transdução de Sinais/efeitos dos fármacosRESUMO
Oxidative stress plays a pivotal role in chronic heart failure. SIRT1, an NAD(+)-dependent histone/protein deacetylase, promotes cell survival under oxidative stress when it is expressed in the nucleus. However, adult cardiomyocytes predominantly express SIRT1 in the cytoplasm, and its function has not been elucidated. The purpose of this study was to investigate the functional role of SIRT1 in the heart and the potential use of SIRT1 in therapy for heart failure. We investigated the subcellular localization of SIRT1 in cardiomyocytes and its impact on cell survival. SIRT1 accumulated in the nucleus of cardiomyocytes in the failing hearts of TO-2 hamsters, postmyocardial infarction rats, and a dilated cardiomyopathy patient but not in control healthy hearts. Nuclear but not cytoplasmic SIRT1-induced manganese superoxide dismutase (Mn-SOD), which was further enhanced by resveratrol, and increased the resistance of C2C12 myoblasts to oxidative stress. Resveratrol's enhancement of Mn-SOD levels depended on the level of nuclear SIRT1, and it suppressed the cell death induced by antimycin A or angiotensin II. The cell-protective effects of nuclear SIRT1 or resveratrol were canceled by the Mn-SOD small interfering RNA or SIRT1 small interfering RNA. The oral administration of resveratrol to TO-2 hamsters increased Mn-SOD levels in cardiomyocytes, suppressed fibrosis, preserved cardiac function, and significantly improved survival. Thus, Mn-SOD induced by resveratrol via nuclear SIRT1 reduced oxidative stress and participated in cardiomyocyte protection. SIRT1 activators such as resveratrol could be novel therapeutic tools for the treatment of chronic heart failure.
Assuntos
Cardiomiopatia Dilatada/metabolismo , Insuficiência Cardíaca/metabolismo , Fibras Musculares Esqueléticas/enzimologia , Miócitos Cardíacos/enzimologia , Sirtuína 1/metabolismo , Superóxido Dismutase/metabolismo , Adulto , Animais , Cardiomiopatia Dilatada/patologia , Núcleo Celular/enzimologia , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/fisiologia , Células Cultivadas , Doença Crônica , Cricetinae , Modelos Animais de Doenças , Ativação Enzimática/efeitos dos fármacos , Ativação Enzimática/fisiologia , Insuficiência Cardíaca/patologia , Humanos , Masculino , Mesocricetus , Camundongos , Fibras Musculares Esqueléticas/citologia , Infarto do Miocárdio/metabolismo , Infarto do Miocárdio/patologia , Miócitos Cardíacos/citologia , Estresse Oxidativo/efeitos dos fármacos , Estresse Oxidativo/fisiologia , Fenóis/farmacologia , Extratos Vegetais/farmacologia , Ratos , Resveratrol , Sirtuína 1/genética , Estilbenos/farmacologiaRESUMO
The essential trace element selenium and polyunsaturated fatty acids (PUFA) have been used for the prevention of cancer. Both nutrients enhance the apoptosis of malignant cells and provide health benefits. However, an increased dietary intake of PUFA augments the susceptibility of lipid peroxidation and oxidative damage in many cells. So far, relatively few data are available about the interaction of selenium and PUFA in testis and thus a possible effect of both dietary components on the prevention of testicular cancer or on the apoptosis of testicular germ cells. Male germ cells in the rat contain most of the testicular phospholipid hydroperoxide glutathione peroxidase (PHGPx), mainly as the mitochondrial isoform of this selenoprotein (m-PHGPx). An experiment was therefore carried out to determine the action of fish oil, a nutrient rich in PUFA, on the testicular expression of PHGPx. Because the PHGPx formation remains nearly unchanged in the animals fed the PUFA-enriched diet, we conclude that no apoptosis of testicular germ cells is induced by an increased intake of this nutrient. The intake of fish oil in the selenium-deficient animal led to a markedly altered formation of several selenium-containing proteins, including sperm nuclei glutathione peroxidase (snGPx), also designated as the nuclear form of PHGPx (n-PHGPx), and a 10-kDa selenium-containing protein.
Assuntos
Apoptose/efeitos dos fármacos , Gorduras na Dieta/farmacologia , Ácidos Graxos Insaturados/farmacologia , Espermatozoides/efeitos dos fármacos , Animais , Núcleo Celular/enzimologia , Gorduras na Dieta/administração & dosagem , Eletroforese em Gel de Poliacrilamida , Ácidos Graxos Insaturados/administração & dosagem , Óleos de Peixe/administração & dosagem , Óleos de Peixe/química , Óleos de Peixe/farmacologia , Glutationa Peroxidase/metabolismo , Masculino , Fosfolipídeo Hidroperóxido Glutationa Peroxidase , Ratos , Compostos de Selênio/administração & dosagem , Compostos de Selênio/farmacologia , Selenoproteínas/metabolismo , Espermatozoides/citologia , Espermatozoides/metabolismo , Testículo/citologia , Testículo/efeitos dos fármacos , Testículo/metabolismoRESUMO
The bioassay-guided fractionation of a MeOH extract of the heartwood of Caesalpinia sappan L. provided two neuroprotective compounds, sappanchalcone (2) and 4-O-methylepisappanol (3), together with a methoxychalcone, isoliquiritigenin 2'-methyl ether (1), and three aromatic compounds, 4-O-methylsappanol (4), caesalpine J (5), pluchoic acid (6). At concentrations of 20-40 microM, compound 2 showed significant cytoprotective effects against glutamate-induced oxidative stress through the induction of heme oxygenase (HO)-1 in HT22-immortalized hippocampal cells. Compound 3 also showed moderate neuroprotective effect at 40 microM, but compounds 1, 4-6 did not show any protective effects against glutamate-induced cytotoxicity in HT22 cells.
Assuntos
Caesalpinia/química , Chalcona/análogos & derivados , Ácido Glutâmico/efeitos adversos , Fármacos Neuroprotetores/farmacologia , Estresse Oxidativo/efeitos dos fármacos , Extratos Vegetais/farmacologia , Animais , Western Blotting , Linhagem Celular , Núcleo Celular/efeitos dos fármacos , Núcleo Celular/enzimologia , Núcleo Celular/metabolismo , Sobrevivência Celular/efeitos dos fármacos , Chalcona/isolamento & purificação , Chalcona/farmacologia , Citosol/efeitos dos fármacos , Citosol/enzimologia , Citosol/metabolismo , Relação Dose-Resposta a Droga , Heme Oxigenase-1/biossíntese , Hipocampo/citologia , Camundongos , Estrutura Molecular , Fármacos Neuroprotetores/isolamento & purificação , Extratos Vegetais/isolamento & purificação , Espécies Reativas de Oxigênio/metabolismo , Relação Estrutura-Atividade , MadeiraRESUMO
When a plant cell is challenged by a well-defined stimulus, complex signal transduction pathways are activated to promote the modulation of specific sets of genes and eventually to develop adaptive responses. In this context, protein phosphorylation plays a fundamental role through the activation of multiple protein kinase families. Although the involvement of protein kinases at the plasma membrane and cytosolic levels are now well-documented, their nuclear counterparts are still poorly investigated. In the field of plant defence reactions, no known study has yet reported the activation of a nuclear protein kinase and/or its nuclear activity in plant cells, although some protein kinases, e.g. MAPK (mitogen-activated protein kinase), are known to be translocated into the nucleus. In the present study, we investigated the ability of cryptogein, a proteinaceous elicitor of tobacco defence reactions, to induce different nuclear protein kinase activities. We found that at least four nuclear protein kinases are activated in response to cryptogein treatment in a time-dependent manner, some of them exhibiting Ca(2+)-dependent activity. The present study focused on one 47 kDa protein kinase with a Ca(2+)-independent activity, closely related to the MAPK family. After purification and microsequencing, this protein kinase was formally identified as SIPK (salicyclic acid-induced protein kinase), a biotic and abiotic stress-activated MAPK of tobacco. We also showed that cytosolic activation of SIPK is not sufficient to promote a nuclear SIPK activity, the latter being correlated with cell death. In that way, the present study provides evidence of a functional nuclear MAPK activity involved in response to an elicitor treatment.