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Background To determine whether adipocyte-derived lipids could be transferred into breast cancer cells and investigate the underlying mechanisms of subsequent lipolysis and fatty acid trafficking in breast cancer cells. Methods A Transwell co-culture system was used in which human breast cancer cells were cultured in the absence or presence of differentiated murine 3 T3-L1 adipocytes. Migration/invasion and proliferation abilities were compared between breast cancer cells that were cultivated alone and those co-cultivated with mature adipocytes. The ability of lipolysis in breast cancer cells were measured, as well as the expression of the rate-limiting lipase ATGL and fatty acid transporter FABP5. ATGL and FABP5 were then ablated to investigate their impact on the aggressiveness of breast cancer cells that were surrounded by adipocytes. Further, immunohistochemistry was performed to detect differential expression of ATGL and FABP5 in breast cancer tissue sections. Results The migration and invasion abilities of cancer cells were significantly enhanced after co-culture with adipocytes, accompanied by elevated lipolysis and expression of ATGL and FABP5. Abrogation of ATGL and FABP5 sharply attenuated the malignancy of co-cultivated breast cancer cells. However, this phenomenon was not observed if a lipid emulsion was added to the culture medium to substitute for adipocytes. Furthermore, epithelial-mesenchymal transaction was induced in co-cultivated breast cancer cells. That may partially due to the stimulation of PPARβ/δ and MAPK, which was resulted from upregulation of FABP5. As evidenced by immunohistochemistry, ATGL and FABP5 also had higher expression levels at the invasive front of the breast tumor, in where the adipocytes abound, compared to the central area in tissue specimens. Conclusions Lipid originating from tumor-surrounding adipocytes could be transferred into breast cancer cells. Adipocyte-cancer cell crosstalk rather than lipids alone induced upregulation of lipases and fatty acid transport protein in cancer cells to utilize stored lipids for tumor progression. The increased expression of the key lipase ATGL and intracellular fatty acid trafficking protein FABP5 played crucial roles in this process via fueling or signaling. Electronic supplementary material The online version of this article (10.1186/s12964-018-0221-6) contains supplementary material, which is available to authorized users.

Globally, approximately 10-25% of women smoke during pregnancy. Since nicotine is highly addictive, women may use nicotine containing products like nicotine replacement therapies for smoking cessation, but the long-term consequences of early life exposure to nicotine remain poorly defined. Our laboratory has previously demonstrated that maternal nicotine exposed (MNE) rat offspring exhibit hypertriglyceridemia due to increased hepatic de novo lipogenesis. Hypertriglyceridemia may also be attributed to impaired white adipose tissue (WAT) lipid storage; however, the effects of MNE on WAT are not completely understood. We hypothesize that nicotine-induced alterations in adipose function (e.g. lipid storage) underlie dyslipidemia in MNE adults. Female 6 month old rats exposed to nicotine during gestation and lactation exhibited significantly decreased visceral adipocyte cell area by 40%, attributed, in part, to a 3-fold increase in adipose triglyceride lipase (ATGL) protein expression compared to vehicle. Given ATGL has antioxidant properties and in utero nicotine exposure promotes oxidative stress in various tissues, we next investigated if there was evidence of increased oxidative stress in MNE WAT. At both 3 weeks and 6 months, MNE offspring expressed 37-48% higher protein levels of SOD1 and SOD2 in WAT. Since oxidative stress can induce inflammation, we examined the inflammatory profile of WAT and found increased expression of cytokines (IL-1β, TNFα, and IL-6) by 44-61% at 6 months. Collectively, this suggests that the expression of WAT ATGL may be induced to counter MNE-induced oxidative stress and inflammation. However, higher levels of ATGL would further promote lipolysis in WAT, culminating in impaired lipid storage and long-term dyslipidemia.

Introduction Cardiac metabolism undergoes changes in response to pathological left ventricular hypertrophy (LVH) and heart failure (HF), characterized by increased reliance on glycolysis, decreased fatty acid (FA) oxidation and a loss of metabolic flexibility. In addition to the heart, non-cardiac organs are involved in these metabolic changes. In this context, the influence of white adipose tissue (WAT) and its release of FA on the development of LVH and HF are not well investigated. The aim of this study was to investigate the effect of Adipose Triglyceride Lipase (ATGL) in WAT on the development of LVH and HF in a pressure overload-induced cardiac hypertrophy model in mice. Methods Male adipose tissue-specific ATGL knock-out (atATGL-KO) and wild-type mice (WT) underwent transverse aortic constriction (TAC-surgery) or sham- surgery. Echocardiography 1 week before, 5 and 11 weeks after Sham/TAC-surgery were performed to evaluate wall thickness, left ventricular mass (LVM) and cardiac function. In addition, histological and gene expression analysis of hearts were performed. Insulin sensitivity, body composition, glucose tolerance and FA serum levels were measured to investigate metabolic changes. Results TAC-surgery caused a significant increase in LVM compared to Sham- surgery in atATGL-KO and WT mice 5 weeks post-surgery. However, there were no significant differences in LVM between atATGL-KO and WT mice. 11 weeks after TAC-surgery, LVM in WT was significantly larger compared to atATGL-KO (LVM [mg] WT-TAC: 211.16±19.73; atATGL-KO-TAC: 124.32±9.06; p, Einleitung Der kardiale Metabolismus ändert sich während der Entwicklung einer linksventrikulären Hypertrophie (LVH) und einer Herzinsuffizienz (HF), welche gekennzeichnet ist durch eine erhöhte Abhängigkeit von Glykolyse, einer erniedrigten Oxidation von Fettsäuren (FA) und einem Verlust der metabolischen Flexibilität. Neben dem Herzen sind auch nicht-kardiale Organe von diesen metabolischen Veränderungen betroffen. In diesem Zusammenhang ist der Einfluss von weißen Fettgewebe (WAT) und seiner Freisetzung von FA auf die Entwicklung der LVH und HF wenig untersucht. Das Ziel dieser Studie war es, den Effekt von Adipozyten Triglycerid Lipase (ATGL) im WAT auf die Entwicklung einer LVH und HF in einem Model der druckinduzierenten kardialen Hypertrophie in der Maus zu untersuchen. Methoden Männliche fettgewebsspezifische ATGL knock-out (atATGL- KO) und Wildtyp Mäuse (WT) wurden einer transversen Aortenkonstriktion (TAC- OP) oder Scheinoperation (Sham-OP) unterzogen. 1 Woche vor, 5 und 11 Wochen nach der Sham/TAC-OP wurde die linksventrikuläre Masse (LVM) und die kardiale Funktion echokardiographisch untersucht. Zusätzlich wurden histologische und Genexpressions-Analysen der Herzen durchgeführt. Die Insulinsensitivität, Glukosetoleranz, Körperzusammensetzung und FA-Serumspiegel wurden gemessen, um metabolische Veränderungen zu untersuchen. Ergebnisse Die TAC-OP resultierte in einer signifikanten Zunahme der LVM im Vergleich zur Sham-OP bei atATGL-KO und WT Mäusen 5 Wochen nach OP. Es lagen jedoch keine signifikanten Unterschiede in der LVM zwischen atATGL-KO und WT Mäusen vor. 11 Wochen nach TAC-OP, LVM in WT war signifikant größer im Vergleich zu atATGL-KO (LVM [mg] WT-TAC: 211,16±19,73; atATGL-KO-TAC: 124,32±9,06; p

// Xiaoying Zhou 1, 2, * , Jiazhang Wei 1, * , Fu Chen 3 , Xue Xiao 1 , Tingting Huang 1 , Qian He 1 , Shumin Wang 1 , Chunping Du 1 , Yingxi Mo 1, 4 , Longde Lin 5 , Ying Xie 1 , Lili Wei 1 , Ying Lan 1 , Mairiko Murata 4 , Guangwu Huang 1 , Ingemar Ernberg 2 , Liudmila Matskova 2 , Zhe Zhang 1 1 Department of Otolaryngology-Head & Neck Surgery, First Affiliated Hospital of Guangxi Medical University, Guangxi, China 2 Department of Microbiology, Tumor and Cell Biology, Karolinska Institutet, Solna, Sweden 3 Department of Radiation Oncology, Eye Ear Nose & Throat Hospital of Fudan University, Shanghai, China 4 Department of Environmental and Molecular Medicine, Mie University Graduate School of Medicine, Mie, Japan 5 School of Public Health, Guangxi Medical University, Guangxi, China * These authors have contributed equally to this work Correspondence to: Ingemar Ernberg e-mail: ingemar.ernberg@ki.se Zhe Zhang e-mail: zhangzhe@gxmu.edu.cn Keywords: UBE2L6, nasopharyngeal carcinoma, DNA methylation, tumor suppressor gene, ATGL Received: April 30, 2015 Accepted: August 20, 2015 Published: October 12, 2015 ABSTRACT We identified the UBE2L6 gene, encoding the ISG15-conjugating enzyme UbcH8, as one gene significantly downregulated by promoter hypermethylation in nasopharyngeal carcinoma (NPC). Reduced expression of the UbcH8 protein correlated with poor outcome in NPC patients. Restored expression of UBE2L6 suppressed proliferation and colony formation in NPC cells, while inducing apoptosis. Of particular interest, we found that aberrant lipid turnover was controlled by UbcH8 in NPC through ISG15-conjugation of valosin-containing protein (VCP). Tumor tissue and NPC cell lines showed conspicuously strong accumulation of lipid droplets (LDs) compared to control nasopharyngeal epithelium and non-cancerous cell lines. We demonstrated that UbcH8 counteracts degradation of adipocyte triglyceride lipase (ATGL), a key enzyme in lipid catabolism.

The G0/G1 switch gene 2 (G0S2) is methylated and silenced in a wide range of human cancers. The protein encoded by G0S2 is an endogenous inhibitor of lipid catabolism that directly binds adipose triglyceride lipase (ATGL). ATGL is the rate-limiting step in triglyceride metabolism. Although the G0S2 gene is silenced in cancer, the impact of ATGL in the growth and survival of cancer cells has never been addressed. Here we show that ectopic expression of G0S2 in non-small cell lung carcinomas (NSCL) inhibits triglyceride catabolism and results in lower cell growth. Similarly, knockdown of ATGL increased triglyceride levels, attenuated cell growth and promoted apoptosis. Conversely, knockdown of endogenous G0S2 enhanced the growth and invasiveness of cancer cells. G0S2 is strongly induced in acute promyelocytic leukemia (APL) cells in response to all trans retinoic acid (ATRA) and we show that inhibition of ATGL in these cells by G0S2 is required for efficacy of ATRA treatment. Our data uncover a novel tumor suppressor mechanism by which G0S2 directly inhibits activity of a key intracellular lipase. Our results suggest that elevated ATGL activity may be a general property of many cancer types and potentially represents a novel target for chemotherapy.

Lipolysis is the biochemical pathway responsible for the catabolism of cellular triacylglycerol (TG). Lipolytic TG breakdown is a central metabolic process leading to the generation of free fatty acids (FA) and glycerol, thereby regulating lipid, as well as energy homeostasis. The precise tuning of lipolysis is imperative to prevent lipotoxicity, obesity, diabetes and other related metabolic disorders. Here, we present our finding that miR-124a attenuates RNA and protein expression of the major TG hydrolase, adipose triglyceride lipase (ATGL/PNPLA2) and its co-activator comparative gene identification 58 (CGI-58/ABHD5). Ectopic expression of miR-124a in adipocytes leads to reduced lipolysis and increased cellular TG accumulation. This phenotype, however, can be rescued by overexpression of truncated Atgl lacking its 3'UTR, which harbors the identified miR-124a target site. In addition, we observe a strong negative correlation between miR-124a and Atgl expression in various murine tissues. Moreover, miR-124a regulates the expression of Atgl and Cgi-58 in murine white adipose tissue during fasting as well as the expression of Atgl in murine liver, during fasting and re-feeding. Together, these results point to an instrumental role of miR-124a in the regulation of TG catabolism. Therefore, we suggest that miR-124a may be involved in the regulation of several cellular and organismal metabolic parameters, including lipid storage and plasma FA concentration.

Hepatic lipid droplet (LD) catabolism is thought to occur via cytosolic lipases such as adipose triglyceride lipase (ATGL) or through autophagy of LDs, a process known as lipophagy. We tested the potential interplay between these metabolic processes and its effects on hepatic lipid metabolism. We show that hepatic ATGL is both necessary and sufficient to induce both autophagy and lipophagy. Moreover, lipophagy is required for ATGL to promote LD catabolism and the subsequent oxidation of hydrolyzed fatty acids (FAs). Following previous work showing that ATGL promotes sirtuin 1 (SIRT1) activity, studies in liver-specific SIRT1−/− mice and in primary hepatocytes reveal that SIRT1 is required for ATGL-mediated induction of autophagy and lipophagy. Taken together, these studies show that ATGL-mediated signaling via SIRT1 promotes autophagy/lipophagy as a primary means to control hepatic LD catabolism and FA oxidation.

cope In recent years, several studies reported the role of eIF4E-binding proteins (4E-BPs) on the development of diet-induced obesity and insulin resistance. Our aim was to investigate the effect of 4E-BP protein deletion on lipid accumulation and metabolism in skeletal muscle in response to a high fat diet-induced obesity in 4E-BP1/2 DKO mice. Methods and results Diet-induced obesity engendered increased ectopic accumulation of lipotoxic species in skeletal muscle of 4E-BP1 and 4E-BP2 double-knockout mice (4E-BP1/2 DKO), namely diacylglycerols and ceramides. Increased lipid accumulation was associated with alterations in the expression of genes involved in fatty acid transport (FATP, CD36), diacylglycerol/triacylglycerol biosynthesis (GPAT1, AGPAT1, DGAT1) and β-oxidation (CPT1b, MCAD). Diet-induced obesity resulted in increased lean mass and muscle in 4E-BP1/2 DKO mice despite the development of a more severe systemic insulin resistance. Since increased expression of genes of several proteolytic systems (MuRF1, Atrogin/MAFbx and Cathepsin-L) in 4EBP1/2 DKO skeletal muscle was reported, the increased of skeletal muscle mass in 4E-BP1/2 DKO mice suggests that ablation of 4E-BPs compensate with activation of muscle anabolism. Conclusions These findings indicate that 4E-BP proteins may prevent excess lipid accumulation in skeletal muscle and suggest that 4E-BPs are key regulators of muscle homeostasis regardless of insulin sensitivity. This article is protected by copyright. All rights reserved

Synucleins are a family of homologous, predominantly neuronal proteins known for their involvement in synaptic transmission and neurodegeneration. γ-synuclein is predominantly localized in axons and presynaptic terminals of selected populations of peripheral and central neurons but is also highly expressed in human white adipose tissue (WAT) and increased in obesity. We have recently shown that γ-synuclein is nutritionally regulated in murine adipocytes while its loss protects mice from high fat diet (HFD)-induced obesity and associated metabolic complications. This protection was coupled with increased adipocyte lipolysis, lipid oxidation, and energy expenditure in HFD-fed γ-synuclein-null mutant compared with wild-type mice. Cellular studies suggest that relocalization of ATGL to the lipid droplet in γ-synuclein-deficient adipocytes may contribute to increased lipolysis in these cells. Loss of γ-synuclein in adipocytes also attenuates the assembly of SNARE complexes, an important component of lipid droplet fusion machinery, possibly due to reduced chaperoning of SNAP-23 to the assembling SNARE complex by γ-synuclein. Together our data suggests that not only is γ-synuclein a novel regulator of lipid handling in adipocytes but also that the deficiency of this protein has a significant effect on whole body energy expenditure.

My PhD research work was focused on the role of muscle lipids in the regulation of energy metabolism and insulin sensitivity. Lipids can be found under two different forms in skeletal muscle: adipocytes located between muscle fibers/bundles and lipid droplets inside muscle fibers (i.e. intramyocellular triacylglycerols or IMTG). These depots, when present in excess, have both been associated with insulin-resistance in humans, mainly because of intracellular lipotoxic lipid accumulation known to impair insulin signaling for IMTG, and through a yet unknown mechanism for adipocytes. First, we isolated and characterized two distinct populations of progenitor cells from human muscle biopsies. The first population is composed of satellite cells (muscle progenitor cells) and display a myogenic differentiation potential in vitro. The second population is composed of cells that acquire the phenotypic and metabolic properties of functional white adipocytes, called fibro/adipogenic progenitors (FAPs). By using these cell models, we showed that FAPs-derived adipocytes secretions are able to impair insulin signaling and action in human skeletal muscle fibers in vitro. This paracrine effect could explain, at least partly, the inverse relationship observed between intramuscular adipocyte content and insulin sensitivity in humans. Secondly, we studied the role of two proteins, G0/G1 Switch Gene 2 (G0S2) and perilipin 5 (PLIN5), in lipid droplets dynamics as well as their impact on lipid metabolism and insulin sensitivity. We showed in vitro that these two proteins play a key role in the control of muscle lipolysis (i.e. IMTG hydrolysis) via the adipose triglyceride lipase (ATGL, catalyzing the limiting step of muscle lipolysis), and that G0S2 and PLIN5 inhibit ATGL activity through direct and indirect mechanisms, respectively. Furthermore, our data showed that G0S2 and PLIN5 invalidation in vivo in mouse skeletal muscle activates lipolysis, increases lipotoxicity and impairs insulin sensitivity. We have also highlighted an important role for PLIN5 in the regulation of fatty acids oxidation, by finely adjusting their availability to energy demand. Overall, these results clearly show on one hand that a crosstalk between adipocytes and fibers within skeletal muscle can lead to an alteration of insulin sensitivity in humans, and on the other hand that G0S2 and PLIN5, two lipid droplet proteins, play a central role in the control of muscle lipid homeostasis and insulin sensitivity. These data help to develop our current understanding of the link between muscle lipids and insulin sensitivity in humans.; Mon travail de thèse a été axé sur l'étude du rôle des lipides musculaires dans la régulation du métabolisme énergétique et la sensibilité à l'insuline. Les lipides sont présents sous deux formes au sein du muscle squelettique : soit sous forme d'adipocytes insérés entre les fibres/faisceaux musculaires, soit sous forme de gouttelettes lipidiques à l'intérieur des fibres musculaires (i.e. triglycérides intramyocellulaires ou IMTG). Ces deux dépôts de lipides, lorsqu'ils sont présents en excès, sont associés à la mise en place de l'insulino-résistance musculaire chez l'homme, via l'accumulation intracellulaire d'espèces lipidiques lipotoxiques altérant la signalisation insulinique pour les IMTG, et par un mécanisme inconnu pour les adipocytes. Dans un premier temps, nous avons isolé et mieux caractérisé, à partir de biopsies musculaires humaines, deux populations de cellules progénitrices. La première population présente un potentiel de différenciation myogénique en culture, il s'agit des cellules satellites (cellules progénitrices musculaires). La deuxième population est composée de cellules capables d'acquérir les propriétés phénotypiques et métaboliques d'adipocytes blancs matures, il s'agit des progéniteurs fibro/adipocytaires (FAPs). Grace à ces modèles d'étude, nous avons mis en évidence que les sécrétions des adipocytes dérivés des FAPs sont capables d'altérer la voie de signalisation et les effets de l'insuline sur des fibres musculaires humaines in vitro. Cet effet paracrine pourrait en partie expliquer la corrélation négative observée entre le contenu en adipocytes intramusculaires et la sensibilité à l'insuline chez l'homme. Dans un second temps, nous avons étudié le rôle de deux protéines, G0/G1 Switch Gene 2 (G0S2) et la périlipine 5 (PLIN5), dans la dynamique des gouttelettes lipidiques ainsi que leur impact sur le métabolisme des lipides et la sensibilité à l'insuline. Nous avons montré in vitro que ces deux protéines jouent un rôle clé dans le contrôle de la lipolyse musculaire (i.e. hydrolyse des IMTG) via l'adipose triglyceride lipase (ATGL, enzyme limitante de la lipolyse musculaire), et que G0S2 et PLIN5 inhibent l'activité de l'ATGL par des mécanismes directs et indirects, respectivement. Par ailleurs, nos données ont montré que l'invalidation de G0S2 et PLIN5 dans le muscle squelettique active la lipolyse, augmente la lipotoxicité et diminue la sensibilité à l'insuline in vivo chez la souris. Nous avons également démontré un rôle important de PLIN5 dans la régulation de l'oxydation des acides gras en ajustant finement leur disponibilité aux besoins énergétiques des cellules. En résumé, ces travaux démontrent d'une part qu'une communication entre adipocytes et fibres au sein du muscle peut entraîner une altération de la sensibilité à l'insuline musculaire chez l'homme, et d'autre part que G0S2 et PLIN5, deux protéines de la gouttelette lipidique, sont au centre du contrôle de l'homéostasie lipidique et du maintien de l'insulino-sensibilité musculaire. Ces données permettent ainsi d'élargir les connaissances existantes sur le lien entre les lipides musculaires et la sensibilité à l'insuline chez l'homme.

Background Obesity is closely related to the metabolism of triacylglycerol (TG) in adipocytes. Adipose triglyceride lipase (ATGL) and hormone-sensitive lipase (HSL) are rate-limiting enzymes that control the hydrolysis of TG. Effects on ATGL and HSL to increase lipolysis may counteract obesity. Berberine (BBR) is a compound derived from the Chinese medicine plant Coptis chinensis. In the present study we show the effects of BBR on ATGL and HSL and explore the potential underlying mechanisms of these effects. Methods The TG content in cells was measured using a colorimetric assay. The expressions of HSL, ATGL and GPAT3 were evaluated by Western-blotting. The expression of ATGL was also evaluated by real-time PCR and radioimmunoassay. Compound C, an inhibitor of AMP-activated protein kinase (AMPK), was used to explore the possible pathway that involved in the effect of BBR on ATGL. Results TG content of differentiated 3T3-L1 cells was significantly decreased by more than 10% after treated with BBR. In differentiated 3T3-L1 adipocytes, BBR increased the expression of p-HSL and ATGL, and these effects were time-depended (p

Triglycerides (TG) hydrolysis in adipose tissue is initialized by the action of the adipose TG lipase (ATGL) and its cofactor, the CGI-58 protein. Mutations in the gene coding the CGI-58 or ATGL proteins are the cause of the syndrome of Chanarin-Dorfman and of neutral lipids storage disease, respectively. CGI-58 protein belongs to the a/ß-hydrolase family, harboring the catalytic triad Ser-Asp/Glu-His, characteristic of carboxylester hydrolases, as well as the HX4D motif, characteristic of acyltransferase activity. Nowadays, the role and the enzymatic activity of the CGI-58 protein, in mammalians and plants, are not quite clear. In order to better understand the function of CGI-58 protein in lipid metabolism, we developed a new strategy using a new set of plasmids that enabled us, with the use of pET28b(+) plasmid, to express and purify the CGI-58 protein of mice and plants as well as the murine ATGL. These proteins were expressed in different E. coli strains, to test in vitro their activity. In addition, we have set up, using the generated plasmids and different E. coli strains, a system that allowed us to test the in vivo acyltransferase activity (LPAAT and/or LPGAT) of the CGI-58 proteins expressed in E. coli. Using these techniques, we demonstrated in vivo and in vitro that both mice and plant CGI-58 proteins, are neither able to catalyze a LPAAT or a LPGAT reaction, and that the plant CGI-58 protein is devoid of TG lipase or phospholipase activity. However we have shown, by analyzing lipid extracts, by thin layer chromatography and by mass spectrometry of different E. coli strains expressing the CGI-58 protein, that the expression of the plant CGI-58 protein, but not the mice one, results in a decrease of phosphatidylglycerol (PG) content in the different strains tested. However the mutation of the serine or histidine residue of the putative catalytic triad restores the wild-typephenotype. These results allowed us to propose that plant CGI-58 protein is involved in the metabolism of PG, L’hydrolyse des triglycérides (TG) du tissu adipeux est initialisée par l’action de l’adipocyte TG lipase (ATGL), activée par son cofacteur la protéine CGI-58. Des mutations du gène codant les protéines CGI-58 ou ATGL sont respectivement à l’origine du syndrome de Chanarin-Dorfmann et de maladies de stockagede lipides neutres. La protéine CGI-58 appartient à la famille des a/ß-hydrolases, elle en possède la triade catalytique Ser-Asp/Glu-His caractéristique des carboxylester hydrolases, ainsi que le motif HX4D, caractéristique d’une activité acyltransférase. A ce jour, le rôle de la protéine CGI-58, chez les mammifères ou chez les plantes, n’est pas totalement éclairci, de même que son activité enzymatique. Dans le but de mieux comprendre le rôle de cette protéine CGI-58 dans le métabolisme lipidique, nous avons développé une nouvelle stratégie, en générant notamment de nouveaux plasmides qui nous ont permis d’exprimer et de purifier la protéine CGI-58 de souris et de plantes, ainsi que l’ATGL murine, dans différents souches d’E. coli, pour tester l’activité in vitro de ces protéines. De plus, nous avons mis en place, en utilisant ces plasmides générés et différentes souches E. coli, un système qui nous a permis de tester in vivo, dans E. coli, l’activité acyltransférase (LPAAT et/ou LPGAT) de la protéine CGI-58. En utilisant ces différentes techniques, nous avons pu montrer, aussi bien in vivo qu’in vitro, que la protéine CGI-58 de plante, ainsique celle de mammifère, ne possède ni activité LPAAT ni activité LPGAT, et que la protéine CGI-58 de plante est dépourvue d’activité TG lipase ou phospholipase. Cependant, nous avons montré, en analysant, par chromatographie sur couche mince et par spectrométrie de masse, des extraits lipidiques des différentes souches d’E. coli exprimant la protéine CGI-58, que l’expression de la protéine de plante, et non celle de mammifère, aboutit à une diminution du taux de phosphatidylglycérol (PG) dans les différentes souchestestées, et a contrario nous avons montré que la mutation de la sérine, ainsi que la mutation de l’histidine de la triade catalytique potentielle, restaure le phénotype sauvage. Ces résultats nous ont permis de proposer que la protéine CGI-58 de plante est impliquée dans le métabolisme du PG

Fatty acids are derived from the hydrolysis of triacylglycerol (TG) found in white adipose tissue, muscle tissue and circulating lipoproteins. The mobilization of free fatty acids (FFA) from white adipose tissue contributes to about 50% of the FFA utilized during moderate-intensity exercise. The delivery of FFA from white adipose tissue is improved by hormone-stimulated lipolytic events in white adipocytes (WA). Thus, the lipolysis in WA that provides fuel for metabolism has been a highly conserved function throughout the course of evolution. This short review outlines our current understanding of the molecular regulation of TG lipases via the lipolytic cascade in WA, as well as provides an account of our recent findings concerning changes in the lipolytic molecules of WA that result from acute and habitual exercise.

Introduction: Trans-10, cis-12 conjugated linoleic acid (CLA) and resveratrol have been shown to reduce TG content in cultured 3T3-L1 adipocyte acting on different pathways. In recent years, the method of simultaneously targeting several signal transduction pathways with multiple natural products in order to achieve additive or synergistic effects has been tested. However, the combined effect of both molecules on lipid metabolism has not been described before. Objective: The aim of the present work was to analyze the effect of the combination of trans-10, cis-12 CLA and resveratrol on TG accumulation as well as on FAS, HSL and ATGL expression in 3T3-L1 mature adipocytes, in order to assess a potential interaction between both molecules. Methods: For this purpose, 3T3-L1 mature adipocytes were treated with the two molecules, both separately and combined, in 10 and 100 μM for 20 hours. TG content and FAS, ATGL and HSL expression were measured by spectrophotometry and Real Time RT-PCR respectively. Results: Both doses of CLA and 100 M resveratrol decreased TG content in mature adipocytes. The combination of both molecules reduced TG accumulation to the same extent as each one separately. No change in FAS and HSL mRNA levels after CLA and resveratrol treatment was observed. ATGL was not modified by CLA but it was increased by resveratrol and by the combination. This combination did not increase the effect caused by resveratrol on its own. Conclusion: Lipolysis increase via ATGL is involved in the TG reduction induced by resveratrol and the combination of both molecules. The combination of these two molecules does not increase the efficacy of each molecule separately in mature adipocytes and thus it does not represent an advantage for obesity treatment or prevention.Introducción: Se ha demostrado que el ácido linoleico trans-10, cis-12 conjugado (ALC) y el resveratrol reducen el contenido de TG en el adipocito 3T3-L1 cultivado actuando sobre distintas vías. En los últimos años, se ha probado el método de llegar a diferentes vías de transducción de señal simultáneamente con múltiples productos naturales con el fin de alcanzar efectos aditivos o sinérgicos. Sin embargo, el efecto combinado de ambas moléculas sobre el metabolismo de los lípidos no se ha descrito previamente. Objetivo: El objetivo de este estudio fue analizar el efecto de la combinación del ALC trans-10, cis-12 y el resveratrol sobre la acumulación de TG así como sobre la expresión de FAS, HSL y ATGL en los adipocitos maduros 3T3-L1 con el propósito de evaluar la interacción potencial entre ambas moléculas. Métodos: Para este propósito, se trató a adipocitos maduros 3T3-L1 con las dos moléculas, de forma separada y combinada, en 10 y 100 μM durante 20 horas. El contenido de TG y la expresión de FAS, ATGL y HSL se midieron con espectrofotometría y en RT-PCR en tiempo real, respectivamente. Resultados: ambas dosis de ALC y 100 μM de resveratrol disminuyeron el contenido de TG en los adipocitos maduros. La combinación de ambas moléculas redujo la acumulación de TG en el mismo grado que cada una de ellas por separado. No se observaron cambios en los niveles de ARNm de FAS y HSL tras el tratamiento con ALC y resveratrol. El ATGL no se modificó por el ALC pero se incrementó por el resveratrol y la combinación. Esta combinación no aumentó el afecto causado por el resveratrol solo. Conclusión: el aumento de la lipólisis vía ATGL está implicado en la reducción de TG inducida por resveratrol y la combinación de ambas moléculas. Esta combinación no aumenta la eficacia de cada una de las moléculas por separado en los adipocitos maduros y, por lo tanto, no representa una ventaja en la prevención ni el tratamiento de la obesidad.

Keratinocyte differentiation is essential for skin development and the formation of the skin permeability barrier. This process involves an orchestrated remodeling of lipids. The cleavage of precursor lipids from lamellar bodies by β-glucocerebrosidase, sphingomyelinase, phospholipases and sterol sulfatase generates ceramides, non-esterified fatty acids and cholesterol for the lipid-containing extracellular matrix, the lamellar membranes in the stratum corneum. The importance of triacylglycerol (TAG) hydrolysis for the formation of a functional permeability barrier was only recently appreciated. Mice with defects in TAG synthesis (acyl-CoA:diacylglycerol acyltransferase-2-knock-out) or TAG catabolism (comparative gene identification-58, -CGI-58-knock-out) develop severe permeability barrier defects and die soon after birth because of desiccation. In humans, mutations in the CGI-58 gene also cause (non-lethal) neutral lipid storage disease with ichthyosis. As a result of defective TAG synthesis or catabolism, humans and mice lack ω-(O)-acylceramides, which are essential lipid precursors for the formation of the corneocyte lipid envelope. This structure plays an important role in linking the lipid-enriched lamellar membranes to highly cross-linked corneocyte proteins. This review focuses on the current knowledge of biochemical mechanisms that are essential for epidermal neutral lipid metabolism and the formation of a functional skin permeability barrier.

The human genome expresses nine patatin-like phospholipase domain containing proteins (PNPLA1–9). Members of this family share a protein domain discovered initially in patatin, the most abundant protein of the potato tuber. Patatin is a lipid hydrolase with an unusual folding topology that differs from classical lipases. Mammalian PNPLAs include lipid hydrolases with specificities for diverse substrates such as triacylglycerols, phospholipids, and retinol esters. Analysis of induced mutant mouse models and the clinical phenotype of patients with mutations revealed important insights into the physiological role of several members of the PNPLA family. This review aims to summarize current knowledge of PNPLA proteins and to document their emerging importance in lipid and energy homeostasis.

Innledning: Overvekt skyldtes høyt inntak av energi i kombinasjon med lite fysisk aktivitet og er ofte forekommende i den norske befolkningen. Ekstra energi (for eksempel fettsyre) tilført kroppen blir omdannet til triglyserider (TG) og lagret i lipiddråper (LDer) i fettvevet som utgjør det primære energilaget. Vedvarende overvekt nedsetter fettvevets evne til å lagre TG og kroppen kompenserer med å lagre LDer i andre av kroppens organer og vev (lever, muskel, hjerte, bukspyttkjertel og karsystemet). Denne lagringen i andre organer og vev er sterkt assosiert med forhøyet risiko for utvikling av metabolsk syndrom, type 2 diabetes (T2D) og hjertekarsykdom. Disse sykdommene blir stadig mer vanlig i befolkningen. LD er organeller som befinner seg i cytoplasma i celler. De består av en hydrofob kjerne av nøytrale lipider (hovedsakelig TG) som er omkranset av et enkelt lag med fosfolipider hvor en rekke ulike proteiner bindes. Perilipiner (Plin) er en familie av proteiner kodet av fem Plin gener (1-5) som binder seg til overflaten av LDer hvor binding til overflaten hindrer degradering av de nøytrale lipidene lagret i kjernen. Plin2 er høyt uttrykt i skjelettmuskel og spiller en rolle i energiomsetningen, men mekanismen er ikke enda helt klarlagt. I en pågående studie i forskningsgruppen er det blitt vist at muskelceller som mangler Plin2 (Plin2-/-) oksiderer mindre glukose men mer fettsyrer, akkumulerer færre LDer noe som tyder på at muskelceller er mindre insulinsensitive i forhold til muskelceller som har tilstede Plin2 (Plin2+/+). I andre studier er det vist at overuttrykk av Plin2 i skjelettmuskel er forbundet med insulinfølsomhet ved å begrense interaksjonen av lipaser (HSL, Atgl) med TGer inne i LDer og hemmer da lipolyse prosessen. Forskningsgruppen ønsker nå å kartlegge den kjemiske sammensetningen av LDer ved isolering av LDer ved hjelp av ultrasentrifugeringen, for dernest å identifisere lipidkomponenter med tynnsjiktkromatografi (TLC) eller proteiner på LDer med Western analyse. Metode: Det ble dyrket myoblast kulturer isolert fra Plin2+/+ og Plin2-/- mus, som differensiert til flerkjernede myotuber over 4 dager før det ble utført forsøk på dem. Myotubene ble inkubert med ulike konsentrasjoner av oljesyre (OA) i 24 timer og behandlet med bærer (DMSO 0,1 %) eller inhibitor av fettvev triglyserid lipase (Atglistatin, 10 µM i DMSO). Kvantitativ sanntids-polymerasekjedereaksjon (qRT-PCR) ble brukt for å undersøke mRNA uttrykket av Plin2 og andre gener i fravær av Plin2 (Plin2-/-) genet. Lagring av LDer i Plin2+/+ og Plin2-/- myotuber ble undersøkt ved mikroskopering og fotografering, kvantifisering av triglyserider og proteiner etter 24 timers forbehandling. For å undersøke effekten av inhibering av Atgl på lagring av LDer ble myotubene også behandlet med Atglistatin. Inhibitoren ble tilsatt samme dag som myotubene ble forbehandlet med OA. Det ble gjennomført etablering av teknikk for isolering av LDer i Plin2+/+ celler. Deretter ble isolering av LDer utført i Plin2+/+ og Plin2-/- celler for å undersøke om tilsetning av Atglistatin førte til at Plin2-/- celler akkumulerte nok LDer til at de kunne isoleres. Resultater: Det ble funnet at proliferering og differensiering av Plin2 celler ikke skjer når cellene ble sådd ut i petriskål med glassbunn sammenlignet med 12-brønners brett med plastbunn. Det ble funnet signifikant lavere triglyserid (TG) akkumulering i Plin2-/- myotuber sammenlignet med Plin2+/+ myotuber, ved hjelp av TG og protein kvantifisering, og mikroskopering ved å farge LDer med Bodipy fargestoff etter 24 timers forbehandling med OA. Behandling med inhibitor av Atgl resulterte i en signifikant høyere akkumulering av TG i myotubene i begge donortyper. Det ble observert lavere akkumulering av TG i Plin2-/- celler forbehandlet med 400 µM OA sammenlignet med 100 µM tilsatt Atglistatin. Dette ble observert både under mikroskopering og kvantifisering av TG. For etablering av teknikk for isolering av LDer ble det funnet at nitrogen kavitasjon for lysering av celler, etterfulgt av en kortvarig lavhastighets sentrifugering og ultrasentrifugering med en tetthetsgradient med sukrose var en effektiv teknikk for å isolere relativt rene LD fra Plin2 celler. Det ble observert at tilsetning av Atglistatin førte til høyere akkumulering av LDer i Plin2-/- celler slik at disse kunne isoleres. Antallet isolert LDer fra Plin2-/- celler var færre sammenlignet med Plin2+/+ celler med og uten lipasehemmeren Atglistatin. Genekspresjonsstudier viste redusert uttrykk av endret Plin2 (Plin2-/- mRNA) og Plin3 i Plin2-/- myotuber sammenlignet med Plin2+/+ myotuber, mens mRNA-uttrykket av Plin4 var høyere i Plin2-/- myotuber sammenlignet med Plin2+/+ myotuber. Diskusjon/konklusjon: Mangel på Plin2 proteinet i myotuber fører til redusert lagring av FFA som TG i LDer. Tilsetning av Atglistatin fører til høyere lagring av TG i LDer i begge donortyper, særlig i Plin2-/- myotuber. Nitrogen kavitasjon er en effektiv teknikk for lysering av celler, mens kortvarig lavhastighetssentrifugering og tetthetsgradient ultrasentrifugering med sukrose er en effektiv teknikk for opprensning av LDer. Tilsetning av Atglistatin i Plin2-/- celler fører til høyere akkumulering av LDer slik at de kunne isoleres.

Lipid droplets (LDs) provide an "on-demand" source of fatty acids (FAs) that can be mobilized in response to fluctuations in nutrient abundance. Surprisingly, the amount of LDs increases during prolonged periods of nutrient deprivation. Why cells store FAs in LDs during an energy crisis is unknown. Our data demonstrate that mTORC1-regulated autophagy is necessary and sufficient for starvation-induced LD biogenesis. The ER-resident diacylglycerol acyltransferase 1 (DGAT1) selectively channels autophagy-liberated FAs into new, clustered LDs that are in close proximity to mitochondria and are lipolytically degraded. However, LDs are not required for FA delivery to mitochondria but instead function to prevent acylcarnitine accumulation and lipotoxic dysregulation of mitochondria. Our data support a model in which LDs provide a lipid buffering system that sequesters FAs released during the autophagic degradation of membranous organelles, reducing lipotoxicity. These findings reveal an unrecognized aspect of the cellular adaptive response to starvation, mediated by LDs.

Obesity is a global epidemic resulting from increased energy intake, which alters energy homeostasis and results in an imbalance in fat storage and breakdown. G0/G1 switch gene 2 (G0s2) has been recently characterised in vitro as an inhibitor of adipose triglyceride lipase (ATGL), the rate-limiting step in fat catabolism. In the current study we aim to functionally characterise G0s2 within the physiological context of a mouse model. We generated a mouse model in which G0s2 was deleted. The homozygous G0s2 knockout (G0s2 −/−) mice were studied over a period of 22 weeks. Metabolic variables were measured including body weight and body composition, food intake, glucose and insulin tolerance tests, energy metabolism and thermogenesis. We report that G0s2 inhibits ATGL and regulates lipolysis and energy metabolism in vivo. G0s2 −/− mice are lean, resistant to weight gain induced by a high-fat diet and are glucose tolerant and insulin sensitive. The white adipose tissue of G0s2 −/− mice has enhanced lipase activity and adipocytes showed enhanced stimulated lipolysis. Energy metabolism in the G0s2 −/− mice is shifted towards enhanced lipid metabolism and increased thermogenesis. G0s2 −/− mice showed enhanced cold tolerance and increased expression of thermoregulatory and oxidation genes within white adipose tissue, suggesting enhanced ‘browning’ of the white adipose tissue. Our data show that G0s2 is a physiological regulator of adiposity and energy metabolism and is a potential target in the treatment of obesity and insulin resistance.

Graphical abstract, Background & Aims Adipose tissue (AT)-derived fatty acids (FAs) are utilized for hepatic triacylglycerol (TG) generation upon fasting. However, their potential impact as signaling molecules is not established. Herein we examined the role of exogenous AT-derived FAs in the regulation of hepatic gene expression by investigating mice with a defect in AT-derived FA supply to the liver. Methods Plasma FA levels, tissue TG hydrolytic activities and lipid content were determined in mice lacking the lipase co-activator comparative gene identification-58 (CGI-58) selectively in AT (CGI-58-ATko) applying standard protocols. Hepatic expression of lipases, FA oxidative genes, transcription factors, ER stress markers, hormones and cytokines were determined by qRT-PCR, Western blotting and ELISA. Results Impaired AT-derived FA supply upon fasting of CGI-58-ATko mice causes a marked defect in liver PPARα-signaling and nuclear CREBH translocation. This severely reduced the expression of respective target genes such as the ATGL inhibitor G0/G1 switch gene-2 (G0S2) and the endocrine metabolic regulator FGF21. These changes could be reversed by lipid administration and raising plasma FA levels. Impaired AT-lipolysis failed to induce hepatic G0S2 expression in fasted CGI-58-ATko mice leading to enhanced ATGL-mediated TG-breakdown strongly reducing hepatic TG deposition. On high fat diet, impaired AT-lipolysis counteracts hepatic TG accumulation and liver stress linked to improved systemic insulin sensitivity. Conclusions AT-derived FAs are a critical regulator of hepatic fasting gene expression required for the induction of G0S2-expression in the liver to control hepatic TG-breakdown. Interfering with AT-lipolysis or hepatic G0S2 expression represents an effective strategy for the treatment of hepatic steatosis.