RESUMO
Since its detection in the brain, the cannabinoid receptor type 2 (CB2) has been considered a promising therapeutic target for various neurological and psychiatric disorders. However, precise brain mapping of its expression is still lacking. Using magnetic cell sorting, calibrated RT-qPCR and single-nucleus RNAseq, we show that CB2 is expressed at a low level in all brain regions studied, mainly by few microglial cells, and by neurons in an even lower proportion. Upon lipopolysaccharide stimulation, modeling neuroinflammation in non-sterile conditions, we demonstrate that the inflammatory response is associated with a transient reduction in CB2 mRNA levels in brain tissue, particularly in microglial cells. This result, confirmed in the BV2 microglial cell line, contrasts with the positive correlation observed between CB2 mRNA levels and the inflammatory response upon stimulation by interferon-gamma, modeling neuroinflammation in sterile condition. Discrete brain CB2 expression might thus be up- or down-regulated depending on the inflammatory context.
Assuntos
Encéfalo , Lipopolissacarídeos , Camundongos Endogâmicos C57BL , Microglia , Receptor CB2 de Canabinoide , Animais , Microglia/metabolismo , Receptor CB2 de Canabinoide/metabolismo , Receptor CB2 de Canabinoide/genética , Receptor CB2 de Canabinoide/biossíntese , Camundongos , Encéfalo/metabolismo , Lipopolissacarídeos/farmacologia , Masculino , Regulação da Expressão Gênica/efeitos dos fármacos , Inflamação/metabolismo , Inflamação/patologia , Doenças Neuroinflamatórias/metabolismoRESUMO
BACKGROUND: Attention-deficit hyperactivity disorder (ADHD) is a frequent comorbidity in children with epilepsy, which management mostly relies on the usual treatments of ADHD, especially methylphenidate. Supplementation with polyunsaturated n-3 Fatty Acid (PUFA) has been proposed as an alternative therapeutic approach in ADHD without epilepsy but has never been evaluated in epilepsy-associated ADHD. METHODS: A multicenter double blind randomized placebo-controlled trial evaluating supplementation with PUFA, in eicosapentaenoic- and docosahexaenoic-acid form, conjugated to a phospholipid vector (PS-Omega3) in children aged >6 and <16-years old, and suffering from any type of epilepsy and ADHD (inattentive or combined type) according to DSM-V. After a 4-week baseline period, patients were allocated (1:1) either to placebo group or to PS-Omega 3 group and entered a 12 week-double-blind treatment period which was followed by a 12 week-open-label treatment period. The primary outcome was the reduction of the ADHD-rating scale IV attention-deficit subscore after 12 weeks of treatment. RESULTS: The study was stopped early because of lack of eligible participants and the expected sample size was not reached. Seventy-four patients were randomized, 44 in PS-Omega3, and 30 in the placebo group. The reduction after 12 weeks of treatment in the inattention subscore of the ADHD-IV scale was -1.57 in the PS-Omega3 group, and -2.90 in the placebo group (p = 0.33, α = 5%). Results were similar after 24 weeks of treatment and for all other ADHD-related secondary outcomes, with no difference between placebo and PS-Omega3. CONCLUSION: Our study remaining underpowered, no formal conclusion about the effect of Ps-Omega3 could be drawn. However, our data strongly suggested that the PS-Omega 3 formulation used in the current study did not improve ADHD symptoms in children with epilepsy. PLAIN LANGUAGE SUMMARY: Supplementation with polyunsaturated n-3 Fatty Acid (PUFA) has been proposed in ADHD but has never been evaluated in patients with both epilepsy and ADHD. To address this issue, we conducted a multicenter double blind randomized placebo-controlled trial evaluating supplementation with PUFA in children with epilepsy and ADHD. The study was stopped early because of lack of eligible participants, hampering formal conclusion. However, the evolution of the ADHD symptoms at 12 and 24 weeks did not differ between placebo and PUFA supplementation, strongly suggesting that PUFA did not improve ADHD symptoms in children with epilepsy.
Assuntos
Transtorno do Deficit de Atenção com Hiperatividade , Epilepsia , Ácidos Graxos Ômega-3 , Criança , Humanos , Adolescente , Transtorno do Deficit de Atenção com Hiperatividade/complicações , Transtorno do Deficit de Atenção com Hiperatividade/tratamento farmacológico , Fosfatidilserinas/uso terapêutico , Resultado do Tratamento , Ácidos Graxos Ômega-3/uso terapêutico , Ácidos Graxos Insaturados/uso terapêutico , Epilepsia/tratamento farmacológico , Suplementos NutricionaisRESUMO
Despite low levels of cannabinoid receptor type 2 (CB2R) expression in the central nervous system in human and rodents, a growing body of evidence shows CB2R involvement in many processes at the behavioral level, through both immune and neuronal modulations. Recent in vitro and in vivo evidence have highlighted the complex role of CB2R under physiological and inflammatory conditions. Under neuroinflammatory states, its activation seems to protect the brain and its functions, making it a promising target in a wide range of neurological disorders. Here, we provide a complete and updated overview of CB2R function in the central nervous system of rodents, spanning from modulation of immune function in microglia but also in other cell types, to behavior and neuronal activity, in both physiological and neuroinflammatory contexts.
Assuntos
Sistema Nervoso Central , Receptor CB2 de Canabinoide , Humanos , Receptor CB2 de Canabinoide/metabolismo , Sistema Nervoso Central/metabolismo , Encéfalo/metabolismo , Neurônios/metabolismo , Microglia/metabolismo , Receptor CB1 de Canabinoide/metabolismoRESUMO
The function of cannabinoid receptor type 2 (CB2R), mainly expressed by leukocytes, has long been limited to its peripheral immunomodulatory role. However, the use of CB2R-specific ligands and the availability of CB2R-Knock Out mice revealed that it could play a functional role in the CNS not only under physiological but also under pathological conditions. A direct effect on the nervous system emerged when CB2R mRNA was detected in neural tissues. However, accurate mapping of CB2R protein expression in the nervous system is still lacking, partly because of the lack of specificity of antibodies available. This review examines the regions and cells of the nervous system where CB2R protein is most likely present by cross-referencing mRNA and protein data published to date. Of the many antibodies developed to target CB2R, only a few have partially passed specificity tests and detected CB2R in the CNS. Efforts must be continued to support the development of more specific and better validated antibodies in each of the species in which CB2R protein is sought or needs to be quantified.
Assuntos
Canabinoides , Camundongos , Animais , Camundongos Knockout , Sistema Nervoso , RNA Mensageiro , Receptor CB2 de Canabinoide/genéticaRESUMO
Free-living amoebae are known to act as replication niches for the pathogenic bacterium Legionella pneumophila in freshwater environments. However, we previously reported that some strains of the Willaertia magna species are more resistant to L. pneumophila infection and differ in their ability to support its growth. From this observation, we hypothesize that L. pneumophila growth in environment could be partly dependent on the composition of amoebic populations and on the possible interactions between different amoebic species. We tested this hypothesis by studying the growth of L. pneumophila and of a permissive free-living amoeba, Vermamoeba vermiformis (formerly named Hartmannella vermiformis), in co-culture with or without other free-living amoebae (Acanthamoeba castellanii and W. magna). We demonstrate the occurrence of inter-amoebic phagocytosis with A. castellanii and W. magna being able to ingest V. vermiformis infected or not infected with L. pneumophila. We also found that L. pneumophila growth is strongly impacted by the permissiveness of each interactive amoeba demonstrating that L. pneumophila proliferation and spread are controlled, at least in part, by inter-amoebic interactions.
Assuntos
Amébidos/microbiologia , Legionella pneumophila/crescimento & desenvolvimento , Fagocitose , Amébidos/classificação , Amébidos/crescimento & desenvolvimento , Técnicas de Cocultura , Interações entre Hospedeiro e Microrganismos , Doença dos Legionários/transmissão , Microbiologia da ÁguaRESUMO
Legionella pneumophila is a human pathogen responsible for a severe form of pneumonia named Legionnaire disease. Its natural habitat is aquatic environments, being in a free state or intracellular parasites of free-living amoebae, such as Acanthamoeba castellanii. This pathogen is able to replicate within some amoebae. Willaertia magna C2c Maky, a non-pathogenic amoeba, was previously demonstrated to resist to L. pneumophila and even to be able to eliminate the L. pneumophila strains Philadelphia, Lens, and Paris. Here, we studied the induction of seven virulence genes of three L. pneumophila strains (Paris, Philadelphia, and Lens) within W. magna C2c Maky in comparison within A. castellanii and with the gene expression level of L. pneumophila strains alone used as controls. We defined a gene expression-based virulence index to compare easily and without bias the transcript levels in different conditions and demonstrated that W. magna C2c Maky did not increase the virulence of L. pneumophila strains in contrast to A. castellanii. These results confirmed the non-permissiveness of W. magna C2c Maky toward L. pneumophila strains.
RESUMO
Repeated deep dives are highly pro-oxidative events for air-breathing aquatic foragers such as penguins. At fledging, the transition from a strictly terrestrial to a marine lifestyle may therefore trigger a complex set of anti-oxidant responses to prevent chronic oxidative stress in immature penguins but these processes are still undefined. By combining in vivo and in vitro approaches with transcriptome analysis, we investigated the adaptive responses of sea-acclimatized (SA) immature king penguins (Aptenodytes patagonicus) compared with pre-fledging never-immersed (NI) birds. In vivo, experimental immersion into cold water stimulated a higher thermogenic response in SA penguins than in NI birds, but both groups exhibited hypothermia, a condition favouring oxidative stress. In vitro, the pectoralis muscles of SA birds displayed increased oxidative capacity and mitochondrial protein abundance but unchanged reactive oxygen species (ROS) generation per g tissue because ROS production per mitochondria was reduced. The genes encoding oxidant-generating proteins were down-regulated in SA birds while mRNA abundance and activity of the main antioxidant enzymes were up-regulated. Genes encoding proteins involved in repair mechanisms of oxidized DNA or proteins and in degradation processes were also up-regulated in SA birds. Sea life also increased the degree of fatty acid unsaturation in muscle mitochondrial membranes resulting in higher intrinsic susceptibility to ROS. Oxidative damages to protein or DNA were reduced in SA birds. Repeated experimental immersions of NI penguins in cold-water partially mimicked the effects of acclimatization to marine life, modified the expression of fewer genes related to oxidative stress but in a similar way as in SA birds and increased oxidative damages to DNA. It is concluded that the multifaceted plasticity observed after marine life may be crucial to maintain redox homeostasis in active tissues subjected to high pro-oxidative pressure in diving birds. Initial immersions in cold-water may initiate an hormetic response triggering essential changes in the adaptive antioxidant response to marine life.
Assuntos
Antioxidantes/metabolismo , Hormese , Spheniscidae/fisiologia , Termotolerância , Animais , Metabolismo Basal , Metabolismo Energético , Peróxido de Hidrogênio/metabolismo , Mitocôndrias Musculares/metabolismo , OxirreduçãoRESUMO
The impacts of high-fat diets (HFDs) on the onset of metabolic endotoxemia and low-grade inflammation are well established in rodent models. However, the dose-effect of dietary lipid intakes on these parameters is not known. We hypothesized that increasing dietary lipid amounts could be linked to parallel increases of endotoxemia, low-grade inflammation, and metabolic and intestinal alterations. Six-week-old male C57BL/6J mice were fed a low-fat diet (LFD, 2.6 wt% of lipids), a moderate HFD (mHFD, 22 wt% of lipids), or a very HFD (vHFD, 45 wt% of lipids) formulated mainly using chow ingredients and milk fat. After 12 weeks, white adipose tissues, liver, intestine, distal colon contents, and plasma were collected. Only vHFD mice significantly increased body weight and fat mass vs LFD mice. This was associated with increases of plasma concentrations of triglycerides, leptin and adiponectin, and liver lipids. No such differences were observed between LFD and mHFD mice. However, mHFD developed metabolic endotoxemia and inflammation, unlike vHFD mice. In turn, vHFD mice showed more goblet cells in all intestine segments vs both other groups and a decrease of Bacteroides-Prevotella in their microbiota vs LFD mice. Finally, mHFD mice colon exhibited a decrease in lactobacilli and in the levels of occludin phosphorylation. Altogether, using complex HFD, no associations were observed between dietary lipid amounts and the magnitude of endotoxemia, inflammation, and physiological alterations developed. These results reveal the impact of the diet composition on intestinal goblet cells and mucus coat, bringing new insights about further consequences on HFD-induced metabolic disorders.
Assuntos
Gorduras na Dieta/administração & dosagem , Endotoxemia/fisiopatologia , Células Caliciformes/metabolismo , Inflamação/fisiopatologia , Adiponectina/sangue , Tecido Adiposo Branco/metabolismo , Animais , Colo/metabolismo , Dieta com Restrição de Gorduras , Dieta Hiperlipídica , Interleucina-6/sangue , Mucosa Intestinal/metabolismo , Intestinos/citologia , Leptina/sangue , Lipopolissacarídeos , Fígado/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Ocludina/metabolismo , Triglicerídeos/sangue , Aumento de Peso , Proteína da Zônula de Oclusão-1/metabolismoRESUMO
Dairy products derived from the milk of cows fed in pastures are characterised by higher amounts of conjugated linoleic acid and α-linolenic acid (ALA), and several studies have shown their ability to reduce cardiovascular risk. However, their specific metabolic effects compared with standard dairy in a high-fat diet (HFD) context remain largely unknown; this is what we determined in the present study with a focus on the metabolic and intestinal parameters. The experimental animals were fed for 12 weeks a HFD containing 20 % fat in the form of a pasture dairy cream (PDC) or a standard dairy cream (SDC). Samples of plasma, liver, white adipose tissue, duodenum, jejunum and colon were analysed. The PDC mice, despite a higher food intake, exhibited lower fat mass, plasma and hepatic TAG concentrations, and inflammation in the adipose tissue than the SDC mice. Furthermore, they exhibited a higher expression of hepatic PPARα mRNA and adipose tissue uncoupling protein 2 mRNA, suggesting an enhanced oxidative activity of the tissues. These results might be explained, in part, by the higher amounts of ALA in the PDC diet and in the liver and adipose tissue of the PDC mice. Moreover, the PDC diet was found to increase the proportions of two strategic cell populations involved in the protective function of the intestinal epithelium, namely Paneth and goblet cells in the small intestine and colon, compared with the SDC diet. In conclusion, a PDC HFD leads to improved metabolic outcomes and to a stronger gut barrier compared with a SDC HFD. This may be due, at least in part, to the protective mechanisms induced by specific lipids.
Assuntos
Bovinos/fisiologia , Dieta/veterinária , Gorduras na Dieta/uso terapêutico , Alimento Funcional , Leite , Obesidade/fisiopatologia , Paniculite/prevenção & controle , Tecido Adiposo Branco/imunologia , Tecido Adiposo Branco/metabolismo , Tecido Adiposo Branco/patologia , Criação de Animais Domésticos , Animais , Laticínios/efeitos adversos , Laticínios/análise , Dieta Hiperlipídica/efeitos adversos , Gorduras na Dieta/efeitos adversos , Gorduras na Dieta/análise , Gorduras na Dieta/metabolismo , Feminino , Alimento Funcional/análise , Hipertrigliceridemia/etiologia , Hipertrigliceridemia/prevenção & controle , Mucosa Intestinal/imunologia , Mucosa Intestinal/metabolismo , Mucosa Intestinal/patologia , Mucosa Intestinal/fisiopatologia , Lactação , Fígado/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Leite/efeitos adversos , Leite/química , Leite/metabolismo , Obesidade/imunologia , Obesidade/metabolismo , Obesidade/patologia , Paniculite/etiologia , Componentes Aéreos da Planta/química , Componentes Aéreos da Planta/crescimento & desenvolvimento , Poaceae/química , Poaceae/crescimento & desenvolvimento , Distribuição AleatóriaRESUMO
Thyroid hormones (TH) are major contributor to oxidative stress in mammals because they (1) stimulate reactive oxygen species generation (ROS), (2) impair antioxidant defenses, and (3) increase the susceptibility to free radicals of most tissues. Unlike mammals, THs seem to diminish mitochondrial ROS while they have limited effect on the antioxidant machinery in birds. However, how THs modify the susceptibility to ROS has never been explored in an avian model, and very little is known about their effect on oxidative balance in birds. Therefore, the objective of our study was to examine the effect of chronic pharmacological hypo- and hyperthyroidism on (i) the susceptibility of mitochondrial membranes to ROS; and (ii) the level of oxidative stress assessed by measuring oxidative damage to lipids, nucleic acids and proteins in the gastrocnemius muscle of ducklings. We show that hypothyroidism had no effect on the susceptibility of mitochondrial membranes to free radicals. Hypothyroid ducklings had lower oxidized lipids (-31%) and DNA (-25%) but a similar level of protein carbonylation relative to controls. Conversely, mitochondrial membranes of hyperthyroid ducklings exhibited higher unsaturation (+12%) and peroxidation (+31%) indexes than in controls indicating a greater susceptibility to free radicals. However, hyperthyroid ducklings exhibited more oxidative damages on proteins (+67%) only, whereas lipid damages remained unchanged, and there was a slight reduction (-15%) in damages to DNA compared to euthyroid controls. Our results indicate that birds and mammals present fundamental differences in their oxidative stress response to thyroid status.
Assuntos
DNA/metabolismo , Patos/metabolismo , Radicais Livres/metabolismo , Hipertireoidismo/metabolismo , Hipotireoidismo/metabolismo , Músculo Esquelético/metabolismo , Estresse Oxidativo/fisiologia , Animais , Antioxidantes , Dano ao DNA , Peroxidação de Lipídeos , Mitocôndrias/metabolismo , Músculo Esquelético/química , Carbonilação Proteica , Proteínas/metabolismo , Espécies Reativas de Oxigênio , Hormônios Tireóideos/metabolismoRESUMO
Environmental enrichment of laboratory animals influences brain plasticity, stimulates neurogenesis, increases neurotrophic factor expression, and protects against the effects of brain insult. However, these positive effects are not constantly observed, probably because standardized procedures of environmental enrichment are lacking. Therefore, we engineered an enriched cage (the Marlau™ cage), which offers: (1) minimally stressful social interactions; (2) increased voluntary exercise; (3) multiple entertaining activities; (4) cognitive stimulation (maze exploration), and (5) novelty (maze configuration changed three times a week). The maze, which separates food pellet and water bottle compartments, guarantees cognitive stimulation for all animals. Compared to rats raised in groups in conventional cages, rats housed in Marlau™ cages exhibited increased cortical thickness, hippocampal neurogenesis and hippocampal levels of transcripts encoding various genes involved in tissue plasticity and remodeling. In addition, rats housed in Marlau™ cages exhibited better performances in learning and memory, decreased anxiety-associated behaviors, and better recovery of basal plasma corticosterone level after acute restraint stress. Marlau™ cages also insure inter-experiment reproducibility in spatial learning and brain gene expression assays. Finally, housing rats in Marlau™ cages after severe status epilepticus at weaning prevents the cognitive impairment observed in rats subjected to the same insult and then housed in conventional cages. By providing a standardized enriched environment for rodents during housing, the Marlau™ cage should facilitate the uniformity of environmental enrichment across laboratories.
Assuntos
Encéfalo/fisiologia , Encéfalo/fisiopatologia , Cognição , Abrigo para Animais/normas , Plasticidade Neuronal , Estado Epiléptico/patologia , Estado Epiléptico/fisiopatologia , Adaptação Psicológica/fisiologia , Animais , Ansiedade/complicações , Peso Corporal , Encéfalo/citologia , Encéfalo/patologia , Região CA1 Hipocampal/citologia , Região CA1 Hipocampal/patologia , Região CA1 Hipocampal/fisiologia , Região CA1 Hipocampal/fisiopatologia , Ingestão de Alimentos , Comportamento Exploratório/fisiologia , Saúde , Metabolismo dos Lipídeos , Masculino , Neurogênese/genética , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Ratos , Padrões de Referência , Reprodutibilidade dos Testes , Restrição Física/psicologia , Estado Epiléptico/psicologia , Estresse Psicológico/complicações , Estresse Psicológico/patologia , Estresse Psicológico/fisiopatologia , Sinapses/patologiaRESUMO
Low-grade inflammation observed in obesity is a risk factor for cardiovascular disease. Recent studies revealed that this would be linked to gut-derived endotoxemia during fat digestion in high-fat diets, but nothing is known about the effect of lipid composition. The study was designed to test the impact of oil composition of high-fat diets on endotoxin metabolism and inflammation in mice. C57/Bl6 mice were fed for 8 wk with chow or isocaloric isolipidic diets enriched with oils differing in fatty acid composition: milk fat, palm oil, rapeseed oil, or sunflower oil. In vitro, adipocytes (3T3-L1) were stimulated or not with lipopolysaccharide (LPS; endotoxin) and incubated with different fatty acids. In mice, the palm group presented the highest level of IL-6 in plasma (P < 0.01) together with the highest expression in adipose tissue of IL-1ß and of LPS-sensing TLR4 and CD14 (P < 0.05). The higher inflammation in the palm group was correlated with a greater ratio of LPS-binding protein (LBP)/sCD14 in plasma (P < 0.05). The rapeseed group resulted in higher sCD14 than the palm group, which was associated with lower inflammation in both plasma and adipose tissue despite higher plasma endotoxemia. Taken together, our results reveal that the palm oil-based diet resulted in the most active transport of LPS toward tissues via high LBP and low sCD14 and the greatest inflammatory outcomes. In contrast, a rapeseed oil-based diet seemed to result in an endotoxin metabolism driven toward less inflammatory pathways. This shows that dietary fat composition can contribute to modulate the onset of low-grade inflammation through the quality of endotoxin receptors.
Assuntos
Tecido Adiposo Branco/imunologia , Citocinas/metabolismo , Dieta Hiperlipídica/efeitos adversos , Doenças Metabólicas/etiologia , Doenças Metabólicas/imunologia , Receptores Imunológicos/metabolismo , Células 3T3-L1 , Proteínas de Fase Aguda , Tecido Adiposo Branco/metabolismo , Animais , Biomarcadores/sangue , Biomarcadores/metabolismo , Proteínas de Transporte/sangue , Citocinas/sangue , Ácidos Graxos Monoinsaturados , Ácidos Graxos não Esterificados/efeitos adversos , Ácidos Graxos não Esterificados/sangue , Bactérias Gram-Negativas/imunologia , Bactérias Gram-Negativas/isolamento & purificação , Bactérias Gram-Positivas/imunologia , Bactérias Gram-Positivas/isolamento & purificação , Intestinos/imunologia , Intestinos/microbiologia , Intestinos/patologia , Receptores de Lipopolissacarídeos/sangue , Receptores de Lipopolissacarídeos/metabolismo , Masculino , Glicoproteínas de Membrana/sangue , Doenças Metabólicas/metabolismo , Doenças Metabólicas/microbiologia , Camundongos , Camundongos Endogâmicos C57BL , Óleo de Palmeira , Óleos de Plantas/efeitos adversos , Distribuição Aleatória , Óleo de Brassica napus , Óleo de Girassol , Receptor 4 Toll-Like/metabolismoRESUMO
We previously demonstrated the importance of upregulation of phosphatidylethanolamine N-methylation pathway in euryhaline fish and crustaceans facing hyperosmotic conditions. In marine molluscs phosphatidylcholine synthesis through N-methylation of phosphatidylethanolamine has not been described until now. In vivo labeling of the mussel Mytilus galloprovincialis with [1-(3)H]-ethanolamine showed that the digestive gland is the tissue expressing the highest incorporation into lipids. A sustained increase in lipid labeling was observed up to 72 h following label injection with 79-92% of radioactivity concentrated into phosphatidylethanolamine and phosphatidylcholine. A direct correlation (r = 0.47, p < 0.01) between the specific radioactivities of phosphatidylcholine in plasma and the digestive gland was observed. Moreover, the phosphatidylcholine fatty acid compositions of plasma and the digestive gland were similar but differed from those of phosphatidylcholine purified from other tissues. In vitro incubation of tissues with [1-(3)H]-ethanolamine or L-[3-(3)H]-serine showed that a significant labeling of the choline moiety of phosphatidylcholine was observed in the digestive gland and hemocytes. Pulse-chase experiments with [1-(3)H]-ethanolamine also demonstrated that hemocytes are exchanging the newly formed phospholipids with plasma. Finally, phosphatidylethanolamine N-methyltransferase assays demonstrated salinity-dependent activities in the digestive gland and hemocytes. We conclude that in M. galloprovincialis an active phosphatidylcholine synthesis through N-methylation of phosphatidylethanolamine occurs in the digestive gland and hemocytes and that this newly formed phosphatidylcholine is partly exchanged with plasma.
Assuntos
Organismos Aquáticos/fisiologia , Trato Gastrointestinal/metabolismo , Hemócitos/metabolismo , Mytilus/fisiologia , Fosfatidilcolinas/biossíntese , Fosfatidiletanolaminas/metabolismo , Animais , Etanolamina/metabolismo , Hidrogênio/metabolismo , Metilação , Concentração Osmolar , Fosfatidiletanolamina N-Metiltransferase/metabolismo , Radioisótopos/metabolismo , Água do Mar , Serina/metabolismoRESUMO
Phosphatidylcholine (PC), the main phospholipid in eukaryotes, is synthesized via two different routes, the phosphatidylethanolamine N-methyl transferase (PEMT) and the CDP-choline pathways. We previously showed in euryhaline fish that salinity impacts the relative contribution of the two pathways for PC biosynthesis, with PEMT pathway being activated in the liver of sea water (SW)-adapted animals. To address the occurrence of such phenomenon in other animals we performed in vivo metabolic studies in two crustacean species: the Chinese crab (Eriocheir sinensis) and the green crab (Carcinus maenas). In both species, the levels of PC and phosphatidylethanolamine in hepatopancreas and hemolymph were not modified by SW-adaptation. In E. sinensis, SW-adaptation activated PC labeling from L-(U-(14)C)-serine in the hepatopancreas and resulted in an increased ratio of PC specific activities between hemolymph and hepatopancreas. In C. maenas, incorporation of L-(3-(3)H)-serine and L-(2-(14)C)-ethanolamine into PC of hepatopancreas was strongly inhibited after acclimation to fresh water (FW). The results show that PC synthesis via the PEMT pathway and its subsequent release into hemolymph are both activated in SW- compared to FW-adapted animals. SW-adaptation also resulted in increased tissue concentrations of betaine and labeling from L-(U-(14)C)-serine, suggesting that the PEMT-derived PC is used for the synthesis of organic osmolytes. The physiological relevance of these observations is discussed.
Assuntos
Crustáceos/metabolismo , Hemolinfa/metabolismo , Hepatopâncreas/metabolismo , Fosfatidilcolinas/metabolismo , Fosfatidiletanolaminas/metabolismo , Salinidade , Adaptação Fisiológica , Animais , Metilação , Fosfatidilcolinas/biossínteseRESUMO
The cannabinoid Delta(9)-tetrahydrocannabinol inhibits the growth of some pathogenic amoebae in vitro and exacerbates amoebic encephalitis in animal models. However, the effects of endogenous cannabinoids on amoebae remain unknown. Therefore, we tested several endocannabinoids (N-acyl ethanolamines and 2-O-acyl glycerol) on different genera of amoebae. The results showed that all of the endocannabinoids tested inhibit amoebic growth at subpharmacological doses, with 50% inhibitory concentrations ranging from 15 to 20 microM. A nonhydrolyzable endocannabinoid had similar effects, showing that the inhibition seen results from endocannabinoids per se rather than from a catabolic product.
Assuntos
Amoeba/efeitos dos fármacos , Amoeba/crescimento & desenvolvimento , Moduladores de Receptores de Canabinoides/farmacologia , Endocanabinoides , Animais , Dronabinol/farmacologiaRESUMO
Erythropoietin receptor (EpoR) binding mediates neuroprotection by endogenous Epo or by exogenous recombinant human (rh)Epo. The level of EpoR gene expression may determine tissue responsiveness to Epo. Thus, harnessing the neuroprotective power of Epo requires an understanding of the Epo-EpoR system and its regulation. We tested the hypothesis that neuronal expression of EpoR is required to achieve optimal neuroprotection by Epo. The ventral limbic region (VLR) in the rat brain was used because we determined that its neurons express minimal EpoR under basal conditions, and they are highly sensitive to excitotoxic damage, such as occurs with pilocarpine-induced status epilepticus (Pilo-SE). We report that (i) EpoR expression is significantly elevated in nearly all VLR neurons when rats are subjected to 3 moderate hypoxic exposures, with each separated by a 4-day interval; (ii) synergistic induction of EpoR expression is achieved in the dorsal hippocampus and neocortex by the combination of hypoxia and exposure to an enriched environment, with minimal increased expression by either treatment alone; and (iii) rhEpo administered after Pilo-SE cannot rescue neurons in the VLR, unless neuronal induction of EpoR is elicited by hypoxia before Pilo-SE. This study thus demonstrates using environmental manipulations in normal rodents, the strict requirement for induction of EpoR expression in brain neurons to achieve optimal neuroprotection. Our results indicate that regulation of EpoR gene expression may facilitate the neuroprotective potential of rhEpo.
Assuntos
Eritropoetina/farmacologia , Neurônios/metabolismo , Fármacos Neuroprotetores/farmacologia , Receptores da Eritropoetina/metabolismo , Animais , Ensaio de Imunoadsorção Enzimática , Eritropoetina/metabolismo , Regulação da Expressão Gênica , Hipóxia/metabolismo , Masculino , Pilocarpina/farmacologia , RNA Mensageiro/genética , Ratos , Ratos Sprague-Dawley , Receptores da Eritropoetina/genética , Receptores da Eritropoetina/fisiologia , Proteínas Recombinantes/farmacologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Estado Epiléptico/induzido quimicamente , Estado Epiléptico/patologiaRESUMO
Brain effects of erythropoietin (Epo) are proposed to involve a heteromeric receptor comprising the classical Epo receptor (Epo-R) and the common beta chain (betac). However, data documenting the pattern of betac gene expression in the healthy brain, in comparison with that of the Epo-R gene, are still lacking. The present study is the first to investigate at the same time betac, Epo-R, and Epo gene expression within different rat brain areas throughout the life span, from neonatal to elderly stages, using quantitative RT-PCR for transcripts. Corresponding proteins were localized by using immunohistochemistry. We demonstrate that the betac transcript level does not correlate with that of Epo-R or Epo, whereas the Epo-R transcript level strongly correlates with that of Epo throughout the life span in all brain structures analyzed. Both Epo and Epo-R were detected primarily in neurons. In the hippocampus, the greatest Epo-R mRNA levels were measured during the early postnatal period and in middle-aged rats, associated with an intense neuronal immunolabeling. Conversely, betac protein was barely detectable in the brain at all ages, even in neurons expressing high levels of Epo-R. Finally, betac transcript could not be detected in PC12 cells, even after nerve growth factor-induced neuritogenesis, which is a condition that dramatically enhances Epo-R transcript level. Altogether, our data suggest that most neurons are likely to express high levels of Epo-R but low, if not null, levels of betac. Given that Epo protects extended populations of neurons after injury, a yet-to-be-identified receptor heterocomplex including Epo-R may exist in the large population of brain neurons that does not express betac.
Assuntos
Encéfalo/crescimento & desenvolvimento , Encéfalo/metabolismo , Eritropoetina/metabolismo , Receptores da Eritropoetina/metabolismo , Envelhecimento/metabolismo , Análise de Variância , Animais , Astrócitos/metabolismo , Expressão Gênica , Imuno-Histoquímica , Masculino , Microglia/metabolismo , Neuritos/metabolismo , Neurônios/metabolismo , Células PC12 , RNA Mensageiro/metabolismo , Ratos , Ratos Sprague-Dawley , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
PURPOSE: To define which intracellular pools of sphingomyelin and ceramide are involved in the triggering of apoptosis of Jurkat leukemia cells in response to gamma-ray exposure. METHODS AND MATERIALS: We examined the kinetics of ceramide generation at the whole-cell level and in different subcellular compartments (plasma membrane rafts, mitochondria, and endoplasmic reticulum) after irradiation with photons. Ceramide was measured by high-performance liquid chromatography or after pulse labeling experiments, and the presence of sphingomyelinase within mitochondria was assessed by electron microscopy. RESULTS: Irradiation of Jurkat leukemia cells resulted in the sequential triggering of sphingomyelin hydrolysis, followed by de novo synthesis that led to a late ceramide response (from 24 h) correlated with the triggering of apoptosis. At the subcellular level, pulse-label experiments, using [(3)H]-palmitate as a precursor, strengthened the involvement of the radiation-induced sphingomyelin breakdown and revealed a very early peak (15 min) of ceramide in plasma membrane rafts. A second peak in mitochondria was measured 4 h after irradiation, resulting from an increase of the sphingomyelin content relating to the targeting of acid sphingomyelinase toward this organelle. CONCLUSION: These data confirm that ceramide is a major determinant in the triggering of radiation-induced apoptosis and highlight the complexity of the sequential compartment-specific ceramide-mediated response of Jurkat leukemia cells to gamma-rays.
Assuntos
Apoptose/fisiologia , Ceramidas/biossíntese , Células Jurkat/efeitos da radiação , Esfingomielinas/biossíntese , Membrana Celular/metabolismo , Membrana Celular/efeitos da radiação , Retículo Endoplasmático/metabolismo , Retículo Endoplasmático/efeitos da radiação , Raios gama , Humanos , Hidrólise , Células Jurkat/metabolismo , Mitocôndrias/metabolismo , Mitocôndrias/efeitos da radiação , Palmitatos , Esfingomielina Fosfodiesterase/metabolismo , Fatores de TempoRESUMO
Legionella pneumophila is known as a facultative intracellular parasite of free-living soil and freshwater amoebae, of which several species have been shown to support the growth of the pathogenic bacteria. We report for the first time the behaviour of two strains (c2c and Z503) of the amoeba Willaertia magna towards different strains of L. pneumophila serogroup 1 and compared it with Acanthamoeba castellanii and Hartmannella vermiformis, known to be L. pneumophila permissive. In contrast to the results seen with other amoebae, W. magna c2c inhibited the growth of one strain of Legionella (L. pneumophila, Paris), but not of others belonging to the same serogroup (L. pneumophila, Philadelphia and L. pneumophila, Lens). Also, the different L. pneumophila inhibited cell growth and induced cell death in A. castellanii, H. vermiformis and W. magna Z503 within 3-4 days while W. magna c2c strain remained unaffected even up to 7 days. Electron microscopy demonstrated that the formation of numerous replicative phagosomes observed within Acanthamoeba and Hartmannella is rarely seen in W. magna c2c cocultured with L. pneumophila. Moreover, the morphological differences were observed between L. pneumophila cultured either with Willaertia or other amoebae. These observations show that amoebae are not all equally permissive to L. pneumophila and highlight W. magna c2c as particularly resistant towards some strains of this bacterium.
Assuntos
Acanthamoeba castellanii/microbiologia , Eucariotos/microbiologia , Hartmannella/microbiologia , Interações Hospedeiro-Patógeno , Legionella pneumophila/fisiologia , Acanthamoeba castellanii/crescimento & desenvolvimento , Acanthamoeba castellanii/ultraestrutura , Animais , Técnicas de Cocultura , Eucariotos/crescimento & desenvolvimento , Eucariotos/ultraestrutura , Água Doce , Hartmannella/crescimento & desenvolvimento , Hartmannella/ultraestrutura , Humanos , Legionella pneumophila/patogenicidade , Microscopia Eletrônica de Transmissão , Especificidade da EspécieRESUMO
BACKGROUND: Increasing evidence supports n-3 fatty acid (FA) supplementation for patients with psychiatric disorders, such as attention deficit hyperactivity disorder. However, the exact metabolic fate of dietary eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) on different glyceride carriers remains unclear. OBJECTIVE: We investigated whether conjugation of EPA and DHA to phospholipid (PL-n-3) or to triacylglycerol (fish oil; FO) affects their incorporation in blood compartments and influences executive functioning. DESIGN: Children aged 8-13 y with impaired visual sustained attention performance received placebo, 250 mg/d EPA + DHA esterified to PL-n-3 (300 mg/d phosphatidylserine), or FO for 3 mo in a randomized double-blind manner. Main outcome measures included plasma and erythrocyte FA profile and continuous performance test results (Test of Variables of Attention; TOVA). RESULTS: Sixty of the 83 children enrolled completed the interventions (n = 18-21 per group). There was an enrichment of EPA (1.5-2.2-fold), docosapentaenoic acid (DPA; 1.2-fold), and DHA (1.3-fold) in the PL fraction in the plasma of FO- and PL-n-3-fed children. In erythrocytes, only PL-n-3 resulted in a significant reduction (approximately 30%) of very-long-chain saturated FAs (C20-24) and in an increase (1.2- and 2.2-fold, respectively) in linoleic acid and DPA. Total TOVA scores increased in the PL-n-3 (mean +/- SD: 3.35 +/- 1.86) and FO (1.72 +/- 1.67) groups but not in the placebo group (-0.42 +/- 2.51) (PL-n-3 > FO > placebo; P < 0.001). A significant correlation between the alterations in FAs and increased TOVA scores mainly occurred in the PL-n-3 group. CONCLUSION: Consumption of EPA+DHA esterified to different carriers had different effects on the incorporation of these FAs in blood fractions and on the visual sustained attention performance in children. This trial was registered at clinicaltrials.gov as NCT00382616.