Glutamine depletion by crisantaspase hinders the growth of human hepatocellular carcinoma xenografts

Background: A subset of human hepatocellular carcinomas (HCC) exhibit mutations of β-catenin gene CTNNB1 and overexpress Glutamine synthetase (GS). The CTNNB1-mutated HCC cell line HepG2 is sensitive to glutamine starvation induced in vitro with the antileukemic drug Crisantaspase and the GS inhibitor methionine-L-sulfoximine (MSO). Methods: Immunodeficient mice with subcutaneous xenografts of the CTNNB1-mutated HCC cell lines HepG2 and HC-AFW1 were treated with Crisantaspase and/or MSO, and tumour growth was monitored. At the end of treatment, tumour weight and histology were assessed. Serum and tissue amino acids were determined by HPLC. Gene and protein expression were estimated with RT-PCR and western blot and GS activity with a colorimetric method. mTOR activity was evaluated from the phosphorylation of p70S6K1. Results: Crisantaspase and MSO depleted serum glutamine, lowered glutamine in liver and tumour tissue, and inhibited liver GS activity. HepG2 tumour growth was significantly reduced by either Crisantaspase or MSO, and completely suppressed by the combined treatment. The combined treatment was also effective against xenografts of the HC-AFW1 cell line, which is Crisantaspase resistant in vitro. Conclusions: The combination of Crisantaspase and MSO reduces glutamine supply to CTNNB1-mutated HCC xenografts and hinders their growth

Problemáticas del cambio climático en la Ciudad Autónoma de Buenos Aires - aportes de las cubiertas vegetadas en la regulación térmica

197-209La urbanización se define como el crecimiento físico de las áreas urbanas. Alrededor de la mitad de la población del mundo vive en ciudades. Se espera que esta aumente al 61 por ciento para el 2030. Las ciudades, pese a su diversidad presentan características ambientales comunes en muchas partes del mundo, como por ejemplo la presencia de la Isla de Calor Urbana (ICU). La ICU evidencia el impacto del hábitat construido sobre el medio físico y el aumento de temperatura que produce. La mayoría de las emisiones de gas de efecto invernadero que contribuyen al cambio climático global, provienen de zonas urbanas. El cambio climático está afectando el clima a escala global y en las ciudades no solo modifica las condiciones del ciclo hidrológico, sino también afecta la ICU. Para Buenos Aires se está pronosticando, en el período 2020-2029, un aumento de las temperaturas promedio máximas en 0,6°C hasta más de 2°C. La ley N° 3.871, de adaptación y mitigación al cambio climático en Buenos Aires, propone medidas mínimas de adaptación. Una de ellas son las cubiertas verdes, las cuales, ayudarían principalmente a amortiguar las temperaturas extremas y la isla de calor urbano. El objetivo del presente trabajo fue determinar si, para las condiciones climáticas de la Ciudad Autónoma de Buenos Aires (CABA), las cubiertas vegetadas (con las características del sitio de estudio), pueden significar un aporte a la regulación de temperatura. El estudio permite afirmar, para las condiciones climáticas, período y tipo de cubierta estudiada, que las mismas pueden ser un aporte a la regulación térmica de las edificaciones

Problemáticas del cambio climático en la Ciudad Autónoma de Buenos Aires - aportes de las cubiertas vegetadas en la regulación térmica

197-209La urbanización se define como el crecimiento físico de las áreas urbanas. Alrededor de la mitad de la población del mundo vive en ciudades. Se espera que esta aumente al 61 por ciento para el 2030. Las ciudades, pese a su diversidad presentan características ambientales comunes en muchas partes del mundo, como por ejemplo la presencia de la Isla de Calor Urbana (ICU). La ICU evidencia el impacto del hábitat construido sobre el medio físico y el aumento de temperatura que produce. La mayoría de las emisiones de gas de efecto invernadero que contribuyen al cambio climático global, provienen de zonas urbanas. El cambio climático está afectando el clima a escala global y en las ciudades no solo modifica las condiciones del ciclo hidrológico, sino también afecta la ICU. Para Buenos Aires se está pronosticando, en el período 2020-2029, un aumento de las temperaturas promedio máximas en 0,6°C hasta más de 2°C. La ley N° 3.871, de adaptación y mitigación al cambio climático en Buenos Aires, propone medidas mínimas de adaptación. Una de ellas son las cubiertas verdes, las cuales, ayudarían principalmente a amortiguar las temperaturas extremas y la isla de calor urbano. El objetivo del presente trabajo fue determinar si, para las condiciones climáticas de la Ciudad Autónoma de Buenos Aires (CABA), las cubiertas vegetadas (con las características del sitio de estudio), pueden significar un aporte a la regulación de temperatura. El estudio permite afirmar, para las condiciones climáticas, período y tipo de cubierta estudiada, que las mismas pueden ser un aporte a la regulación térmica de las edificaciones

ALL blasts drive primary mesenchymal stromal cells to increase asparagine availability during asparaginase treatment

Mechanisms underlying the resistance of acute lymphoblastic leukemia (ALL) blasts to L-asparaginase are still incompletely known. Here we demonstrate that human primary bone marrow mesenchymal stromal cells (MSCs) successfully adapt to L-asparaginase and markedly protect leukemic blasts from the enzyme-dependent cytotoxicity through an amino acid tradeoff. ALL blasts synthesize and secrete glutamine, thus increasing extracellular glutamine availability for stromal cells. In turn, MSCs use glutamine, either synthesized through glutamine synthetase (GS) or imported, to produce asparagine, which is then extruded to sustain asparagine-auxotroph leukemic cells. GS inhibition prevents mesenchymal cells adaptation to L-asparaginase, lowers glutamine secretion by ALL blasts, and markedly hinders the protection exerted by MSCs on leukemic cells. The pro-survival amino acid exchange is hindered by the inhibition or silencing of the asparagine efflux transporter SNAT5, which is induced in mesenchymal cells by ALL blasts. Consistently, primary MSCs from ALL patients express higher levels of SNAT5 (P <.05), secrete more asparagine (P <.05), and protect leukemic blasts (P <.05) better than MSCs isolated from healthy donors. In conclusion, ALL blasts arrange a pro-leukemic amino acid trade-off with bone marrow mesenchymal cells, which depends on GS and SNAT5 and promotes leukemic cell survival during L-asparaginase treatment

Brain Region–Specific Decrease in the Activity and Expression of Protein Kinase A in the Frontal Cortex of Regressive Autism

Autism is a severe neurodevelopmental disorder that is characterized by impaired language, communication, and social skills. In regressive autism, affected children first show signs of normal social and language development but eventually lose these skills and develop autistic behavior. Protein kinases are essential in G-protein-coupled, receptor-mediated signal transduction and are involved in neuronal functions, gene expression, memory, and cell differentiation. We studied the activity and expression of protein kinase A (PKA), a cyclic AMP–dependent protein kinase, in postmortem brain tissue samples from the frontal, temporal, parietal, and occipital cortices, and the cerebellum of individuals with regressive autism; autistic subjects without a clinical history of regression; and age-matched developmentally normal control subjects. The activity of PKA and the expression of PKA (C-α), a catalytic subunit of PKA, were significantly decreased in the frontal cortex of individuals with regressive autism compared to control subjects and individuals with non-regressive autism. Such changes were not observed in the cerebellum, or the cortices from the temporal, parietal, and occipital regions of the brain in subjects with regressive autism. In addition, there was no significant difference in PKA activity or expression of PKA (C-α) between non-regressive autism and control groups. These results suggest that regression in autism may be associated, in part, with decreased PKA-mediated phosphorylation of proteins and abnormalities in cellular signaling

Physiologically relevant culture medium Plasmax improves human placental trophoblast stem cell function

Human trophoblast cultures provide powerful tools to model key processes of placental development. In vitro trophoblast studies to date have relied on commercial media which contains non-physiological levels of nutrients, and the impact of these conditions on trophoblast metabolism and function is unknown. Here we show that the physiological medium (Plasmaxä) with nutrient and metabolite concentrations recapitulating human plasma improves human trophoblast stem cell (hTSC) proliferation and differentiation compared to standard medium (DMEM-F12). hTSCs cultured in Plasmax-based medium also show altered glycolytic and mitochondrial metabolism, as well as reduced S-adenosylmethionine/S-adenosyl-homosysteine ratio compared to DMEM-F12-based medium. These findings demonstrate the importance of the nutritional environment for phenotyping cultured human trophoblasts

Chronic treatment with agomelatine or venlafaxine reduces depolarization-evoked glutamate release from hippocampal synaptosomes

Background: Growing compelling evidence from clinical and preclinical studies has demonstrated the primary role of alterations of glutamatergic transmission in cortical and limbic areas in the pathophysiology of mood disorders. Chronic antidepressants have been shown to dampen endogenous glutamate release from rat hippocampal synaptic terminals and to prevent the marked increase of glutamate overflow induced by acute behavioral stress in frontal/prefrontal cortex. Agomelatine, a new antidepressant endowed with MT1/MT2 agonist and 5-HT2C serotonergic antagonist properties, has shown efficacy at both preclinical and clinical levels. Results: Chronic treatment with agomelatine, or with the reference drug venlafaxine, induced a marked decrease of depolarization-evoked endogenous glutamate release from purified hippocampal synaptic terminals in superfusion. No changes were observed in GABA release. This effect was accompanied by reduced accumulation of SNARE protein complexes, the key molecular effector of vesicle docking, priming and fusion at presynaptic membranes. Conclusions: Our data suggest that the novel antidepressant agomelatine share with other classes of antidepressants the ability to modulate glutamatergic transmission in hippocampus. Its action seems to be mediated by molecular mechanisms located on the presynaptic membrane and related with the size of the vesicle pool ready for release

Antidepressant stimulation of CDP-diacylglycerol synthesis does not require monoamine reuptake inhibition

<p>Abstract</p> <p>Background</p> <p>Recent studies demonstrate that diverse antidepressant agents increase the cellular production of the nucleolipid CDP-diacylglycerol and its synthetic derivative, phosphatidylinositol, in depression-relevant brain regions. Pharmacological blockade of downstream phosphatidylinositide signaling disrupted the behavioral antidepressant effects in rats. However, the nucleolipid responses were resistant to inhibition by serotonin receptor antagonists, even though antidepressant-facilitated inositol phosphate accumulation was blocked. Could the neurochemical effects be additional to the known effects of the drugs on monoamine transmitter transporters? To examine this question, we tested selected agents in serotonin-depleted brain tissues, in PC12 cells devoid of serotonin transporters, and on the enzymatic activity of brain CDP-diacylglycerol synthase - the enzyme that catalyzes the physiological synthesis of CDP-diacylglycerol.</p> <p>Results</p> <p>Imipramine, paroxetine, and maprotiline concentration-dependently increased the levels of CDP-diacylglycerol and phosphatidylinositides in PC12 cells. Rat forebrain tissues depleted of serotonin by pretreatment with <it>p</it>-chlorophenylalanine showed responses to imipramine or maprotiline that were comparable to respective responses from saline-injected controls. With fluoxetine, nucleolipid responses in the serotonin-depleted cortex or hippocampus were significantly reduced, but not abolished. Each drug significantly increased the enzymatic activity of CDP-diacylglycerol synthase following incubations with cortical or hippocampal brain tissues.</p> <p>Conclusion</p> <p>Antidepressants probably induce the activity of CDP-diacylglycerol synthase leading to increased production of CDP-diacylglycerol and facilitation of downstream phosphatidylinositol synthesis. Phosphatidylinositol-dependent signaling cascades exert diverse salutary effects in neural cells, including facilitation of BDNF signaling and neurogenesis. Hence, the present findings should strengthen the notion that modulation of brain phosphatidylinositide signaling probably contributes to the molecular mechanism of diverse antidepressant medications.</p

Differential and converging molecular mechanisms of antidepressants' action in the hippocampal dentate gyrus

Major depression is a highly prevalent, multidimensional disorder. Although several classes of antidepressants (ADs) are currently available, treatment efficacy is limited, and relapse rates are high; thus, there is a need to find better therapeutic strategies. Neuroplastic changes in brain regions such as the hippocampal dentate gyrus (DG) accompany depression and its amelioration with ADs. In this study, the unpredictable chronic mild stress (uCMS) rat model of depression was used to determine the molecular mediators of chronic stress and the targets of four ADs with different pharmacological profiles (fluoxetine, imipramine, tianeptine, and agomelatine) in the hippocampal DG. All ADs, except agomelatine, reversed the depression-like behavior and neuroplastic changes produced by uCMS. Chronic stress induced significant molecular changes that were generally reversed by fluoxetine, imipramine, and tianeptine. Fluoxetine primarily acted on neurons to reduce the expression of pro-inflammatory response genes and increased a set of genes involved in cell metabolism. Similarities were found between the molecular actions and targets of imipramine and tianeptine that activated pathways related to cellular protection. Agomelatine presented a unique profile, with pronounced effects on genes related to Rho-GTPase-related pathways in oligodendrocytes and neurons. These differential molecular signatures of ADs studied contribute to our understanding of the processes implicated in the onset and treatment of depression-like symptoms.Patricia Patricio, Antonio Mateus-Pinheiro, Monica Morais, and Nuno Dinis Alves received fellowships from the Portuguese Foundation for Science and Technology (FCT). Michal Korostynski and Marcin Piechota were funded by the POIG De-Me-Ter 3.1 and NCN 2011/03/D/NZ3/01686 grants. This study was co-funded by the Life and Health Sciences Research Institute (ICVS) and ON. 2-O NOVO NORTE-North Portugal Regional Operational Programme 2007/2013, of the National Strategic Reference Framework (NSRF) 2007/ 2013, through the European Regional Development Fund (ERDF) and by the SwitchBox Consortium (Contract FP7-Health-F2-2010-259772 from the European Union). The authors declare no conflict of interest