38 research outputs found
Dynamic Changes in Lipid Droplet-Associated Proteins in the "Browning" of White Adipose Tissues
No full text08.02.13 KB. Accepted version ok to add to Spiral. Elsevier ok while mandate is not enforced.The morphological and functional differences between lipid droplets (LDs) in brown (BAT) and white (WAT) adipose tissue will largely be determined by their associated proteins. Analysing mRNA expression in mice fat depots we have found that most LD protein genes are expressed at higher levels in BAT, with the greatest differences observed for Cidea and Plin5. Prolonged cold exposure, which induces the appearance of brown-like adipocytes in mice WAT depots, was accompanied with the potentiation of the lipolytic machinery, with changes in ATGL, CGI-58 and G0S2 gene expression. However the major change detected in WAT was the enhancement of Cidea mRNA. Together with the increase in Cidec, it indicates that LD enlargement through LD-LD transference of fat is an important process during WAT browning. To study the dynamics of this phenotypic change, we have applied 4D confocal microscopy in differentiated 3T3-L1 cells under sustained beta-adrenergic stimulation. Under these conditions the cells experienced a LD remodelling cycle, with progressive reduction on the LD size by lipolysis, followed by the formation of new LDs, which were subjected to an enlargement process, likely to be CIDE-triggered, until the cell returned to the basal state. This transformation would be triggered by the activation of a thermogenic futile cycle of lipolysis/lipogenesis and could facilitate the molecular mechanism for the unilocular to multilocular transformation during WAT browning
Estudi dels efectes del liti sobre el metabolisme dels fosfolípids i l'alliberació de senyals químics en cultius d'astròcits
No full textDescripció del recurs: el 26 de març de 2010El desenvolupament de noves teràpies per als trastorns bipolars, caracteritzats per l'alternança d'episodis de depressió i d'eufòria exacerbada, es veu dificultat pel desconeixement de l'etiologia de la malaltia i del mecanisme d'acció dels fàrmacs que estabilitzen l'estat d'ànim del pacient, com el valproat, la carbamazepina, la lamotrigina, i el liti. En aquest sentit, una possibilitat que ha estat poc explorada és que les cèl·lules diana d'aquests fàrmacs siguin els astròcits, un tipus de glia que darrerament està emergent com a element actiu en els processos de transmissió sinàptica, fet que ens ha portat a estudiar els efectes del liti en cultius d'astròcits. Hem caracteritzat l'increment que produeix el liti en la velocitat de síntesi de fosfatidilcolina (PC) en astròcits, que ja apareix amb el tractament agut amb liti, però és especialment significatiu a partir de les 24 hores de tractament. El liti potencia la ruta de Kennedy per a la síntesi de PC, un efecte que es veu parcialment revertit per l'addició d'inositol, indicant que la inhibició de la IMPasa per part del liti hi estaria implicada. Els efectes del liti no es limiten a la PC sinó que es produeix una alteració general del metabolisme lipídic dels astròcits. El liti inhibeix la síntesi de fosfatidilinositol i de fosfatidiletanolamina, un efecte que es dóna de manera similar amb valproat o carbamazepina que, de manera oposada al liti, també provoquen una forta inhibició en la síntesi de PC. Paral·lelament, el tractament crònic amb liti sembla reduïr la síntesi de novo d'àcids grassos i de colesterol. Globalment, els efectes observats ens fan pensar que en incrementar la síntesi de PC el liti podria produir una reducció en els nivells de diacilglicerol cel·lulars, fet que podria estar implicat amb el mecanisme terapèutic d'aquest ió. Si la diana terapèutica del liti es troba en els astròcits, és probable que actuï a nivell de les molècules que aquests alliberen per tal de modular la transmissió sinàptica. En aquest sentit hem comprovat que el tractament amb liti, a dosis lleugerament superiors a les terapèutiques, incrementa l'alliberació de TNF-, òxid nítric, i prostaglandina E2 (PGE2) en cultius d'astròcits estimulats amb lipopolisacàrid bacterià (LPS). Sorprenentment en emprar una dosi de liti inferior, dins del rang terapèutic del fàrmac, s'observa una reducció en l'alliberació de PGE2, un efecte reproduït pel valproat, la carbamazepina i la lamotrigina. L'efecte dual del liti en funció de la concentració aplicada, es deu a una reducció en la producció d'àcid araquidònic en resposta a senyals de Ca2+ citosòlic, paral·lela a un increment en la inducció de la ciclooxigenasa-2 (COX-2) en astròcits estimulats amb LPS. Aquest fet podria estar relacionat amb l'estreta finestra terapèutica d'aquest fàmac, i contribueix a relacionar els astròcits amb el mecanisme d'acció dels estabilitzadors de l'estat d'ànim.Bipolar disorder is a common disease characterized by an alternating pattern of depression and mania episodes. Bipolar patients are treated with the mood stabilizing drugs, like lithium, valproate and carbamazepine, however the action mechanism of these drugs remains unknown. As most of the studies on this issue have been focused in neurons, we decided to analyze the effects of lithium in cultured astrocytes, a type of glial cells that are emerging as active elements in the regulation of synaptic transmission. We have characterized the increase on phosphatidylcholine (PC) synthesis induced by lithium in cultured astrocytes, an effect that appears with the acute treatment and is enhanced after 24 hours of treatment with lithium. Lithium potentiates the Kennedy pathway for the synthesis of PC, in an IMPase (inositol monophosphatase) inhibition dependent manner. Lithium effects are not limited to PC but it produces a broad alteration on astrocytic lipid metabolism. Lithium inhibits phosphatidylinositol and phosfatidylethanolamine synthesis, an effect also induced by the treatment with valproate or carbamazepine, which don't stimulate PC synthesis as lithium but inhibit it. On the other hand, chronic lithium treatment reduces fatty acids and cholesterol "de novo" synthesis. Overall, we hypothesized that lithium, increasing PC synthesis, induces a reduction on cellular diacylglycerol levels, which in turn could be related with the therapeutic mechanism of this ion
Lipid droplet growth : regulation of a dynamic organelle
No full textIntracellular lipid droplets (LDs) are remarkably dynamic and complex organelles that enact regulated storage and release of lipids to fulfil their fundamental roles in energy metabolism, membrane synthesis and provision of lipid-derived signaling molecules. Although small LDs are observed in all types of eukaryotic cells, it is adipocytes that present the widest range of sizes up to the massive unilocular droplet of a white adipocyte. Our knowledge of the proteins and associated processes that control LD dynamics is improving. The dynamic expression of LD-associated proteins is vital for controlling LD biology and is most apparent during adipocyte differentiation. Recent findings on the molecular mechanisms of lipid droplet enlargement reveal the importance of distinct functional groups of proteins and phospholipids
Fàrmacs estabilitzadors de l'estat d'ànim en cultius d'astròcits: estudi de la secreció vesicular i de l'alliberament de mol·lecules senyalitzadores
No full textProjecte de recerca elaborat a partir d’una estada al Institut Conzorcio Mario Negri Sud, entre març i setembre del 2007. Els trastorns bipolars es caracteritzen per l’alternança d’episodis de depressió i d’eufòria exacerbada. Tot i el desconeixement que hi ha sobre la patogènesi d’aquesta malaltia, els pacients responen positivament a fàrmacs que els estabilitzen l’estat d’ànim, com ara les sals de liti i els anticonvulsius valproat, carbamazepina i lamotrigina. S’ha descrit que aquests fàrmacs produeixen múltiples efectes bioquímics però encara se’n desconeix el seu mecanisme d’acció. Els astròcits poden modular la transmissió sinàptica, per la qual cosa creiem que la patogènesi i la teràpia de molts trastorns psiquiàtrics, poden estar relacionats amb alteracions de la seva activitat. Considerant aquesta hipòtesi, la recerca s’ha centrat en els efectes dels estabilitzadors de l’estat d’ànim sobre els astròcits, especialment pel que fa l’alliberació de molècules senyalitzadores que puguin actuar com a neuromoduladors
Hallmarks of cancer stem cell metabolism.
No full textCancer cells adapt cellular metabolism to cope with their high proliferation rate. Instead of primarily using oxidative phosphorylation (OXPHOS), cancer cells use less efficient glycolysis for the production of ATP and building blocks (Warburg effect). However, tumours are not uniform, but rather functionally heterogeneous and harbour a subset of cancer cells with stemness features. Such cancer cells have the ability to repopulate the entire tumour and thus have been termed cancer stem cells (CSCs) or tumour-initiating cells (TICs). As opposed to differentiated bulk tumour cells relying on glycolysis, CSCs show a distinct metabolic phenotype that, depending on the cancer type, can be highly glycolytic or OXPHOS dependent. In either case, mitochondrial function is critical and takes centre stage in CSC functionality. Remaining controversies in this young and emerging research field may be related to CSC isolation techniques and/or the use of less suitable model systems. Still, the apparent dependence of CSCs on mitochondrial function, regardless of their primary metabolic phenotype, represents a previously unrecognised Achilles heel amendable for therapeutic intervention. Elimination of highly chemoresistant CSCs as the root of many cancers via inhibition of mitochondrial function bears the potential to prevent relapse from disease and thus improve patients' long-term outcome.ERC Advanced Investigator Grant (Pa-CSC 233460 to CH), the European Community's Seventh Framework Programme (FP7/2007-2013) under grant agreement no 256974 (EPC-TM-NET to C.H.) and no 602783 (CAM-PaC to C.H.), the 2015 SU2C Lustgarten CRUK Pancreatic Cancer Dream Team Award (to CH), Pancreatic Cancer UK RIF2014_04 and RIF2015_03 (both to CH) and the Pancreatic Cancer Research Fund (to PS)
A century later, still fighting back: antigen‐specific immunotherapies for type 1 diabetes
No full textDevelopment of isotope-enriched phosphatidylinositol-4-and 5-phosphate cellular mass spectrometry probes
No full textSynthetic phosphatidylinositol phosphate (PtdInsPn) derivatives play a pivotal role in broadening our understanding of PtdInsPn metabolism. However, the development of such tools is reliant on efficient enantioselective and regioselective synthetic strategies. Here we report the development of a divergent synthetic route applicable to the synthesis of deuterated PtdIns4P and PtdIns5P derivatives. The synthetic strategy developed involves a key enzymatic desymmetrisation step using Lipozyme TL-IM®. In addition, we optimised the large-scale synthesis of deuterated myo-inositol, allowing for the preparation of a series of saturated and unsaturated deuterated PtdIns4P and PtdIns5P derivatives. Experiments in MCF7 cells demonstrated that these deuterated probes enable quantification of the corresponding endogenous phospholipids in a cellular setting. Overall, these deuterated probes will be powerful tools to help improve our understanding of the role played by PtdInsPn in physiology and disease
FAIM-L regulation of XIAP degradation modulates synaptic long-term depression and axon degeneration
No full textCaspases have recently emerged as key regulators of axonal pruning and degeneration and of long-term depression (LTD), a long-lasting form of synaptic plasticity. However, the mechanism underlying these functions remains unclear. In this context, XIAP has been shown to modulate these processes. The neuron-specific form of FAIM protein (FAIM-L) is a death receptor antagonist that stabilizes XIAP protein levels, thus preventing death receptor-induced neuronal apoptosis. Here we show that FAIM-L modulates synaptic transmission, prevents chemical-LTD induction in hippocampal neurons, and thwarts axon degeneration after nerve growth factor (NGF) withdrawal. Additionally, we demonstrate that the participation of FAIM-L in these two processes is dependent on its capacity to stabilize XIAP protein levels. Our data reveal FAIM-L as a regulator of axonal degeneration and synaptic plasticity
The synthetic lethal interaction between CDS1 and CDS2 is a vulnerability in uveal melanoma and across multiple tumor types.
No full textMetastatic uveal melanoma is an aggressive disease with limited effective therapeutic options. To comprehensively map monogenic and digenic dependencies, we performed CRISPR-Cas9 screening in ten extensively profiled human uveal melanoma cell line models. Analysis involved genome-wide single-gene and combinatorial paired-gene CRISPR libraries. Among our 76 uveal melanoma-specific essential genes and 105 synthetic lethal gene pairs, we identified and validated the CDP-diacylglycerol synthase 2 gene (CDS2) as a genetic dependency in the context of low CDP-diacylglycerol synthase 1 gene (CDS1) expression. We further demonstrate that CDS1/CDS2 forms a synthetic lethal interaction in vivo and reveal that CDS2 knockout results in the disruption of phosphoinositide synthesis and increased cellular apoptosis and that re-expression of CDS1 rescues this cell fitness defect. We extend our analysis using pan-cancer data, confirming increased CDS2 essentiality in diverse tumor types with low CDS1 expression. Thus, the CDS1/CDS2 axis is a therapeutic target across a range of cancers
Group IVA phospholipase A2 is necessary for the biogenesis of lipid droplets
No full textLipid droplets (LD) are organelles present in all cell types, consisting of a hydrophobic core of triacylglycerols and cholesteryl esters, surrounded by a monolayer of phospholipids and cholesterol. This work shows that LD biogenesis induced by serum, by long-chain fatty acids, or the combination of both in CHO-K1 cells was prevented by phospholipase A2 inhibitors with a pharmacological profile consistent with the implication of group IVA cytosolic phospholipase A2 (cPLA2α). Knocking down cPLA2α expression with short interfering RNA was similar to pharmacological inhibition in terms of enzyme activity and LD biogenesis. A Chinese hamster ovary cell clone stably expressing an enhanced green fluorescent protein-cPLA2α fusion protein (EGFP-cPLA2) displayed higher LD occurrence under basal conditions and upon LD induction. Induction of LD took place with concurrent phosphorylation of cPLA2α at Ser505. Transfection of a S505A mutant cPLA2α showed that phosphorylation at Ser505 is key for enzyme activity and LD formation. cPLA2α contribution to LD biogenesis was not because of the generation of arachidonic acid, nor was it related to neutral lipid synthesis. cPLA2α inhibition in cells induced to form LD resulted in the appearance of tubulo-vesicular profiles of the smooth endoplasmic reticulum, compatible with a role of cPLA2α in the formation of nascent LD from the endoplasmic reticulum
