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Tissue clearing and fluorescent microscopy are powerful tools for unbiased organ-scale protein expression studies. Critical for interpreting expression patterns of large imaged volumes are reliable quantification methods. Here, we present DELiVR a deep learning pipeline that uses virtual reality (VR)-generated training data to train deep neural networks, and quantify c-Fos as marker for neuronal activity in cleared mouse brains and map its expression at cellular resolution. VR annotation significantly accelerated the speed of generating training data compared to conventional 2D slice based annotation. DELiVR detects cells with much higher precision than current threshold-based pipelines, and provides an extensive toolbox for data visualization, inspection and comparison. We applied DELiVR to profile cancer-related mouse brain activity, and discovered a novel activation pattern that distinguishes between weight-stable cancer and cancer-associated weight loss. Thus, DELiVR provides a robust mouse brain analysis pipeline at cellular scale that can be used to study brain activity patterns in health and disease.The DELiVR software, Fiji plugin and documentation can be found athttps://www.DISCOtechnologies.org/DELiVR/.Graphical AbstractHighlightsDELiVR detects labelled cells in cleared brains with deep learningDELiVR is trained by annotating ground-truth data in virtual reality (VR)DELiVR is launched via a FIJI plugin anywhere from PCs to clustersUsing DELiVR, we found new brain activity patterns in weight-stable vs. cachectic cancerSupplementary Videos can be seen at:https://www.DISCOtechnologies.org/DELiVR/

Background Cancer cachexia (CCx) is a multifactorial wasting disorder characterized by involuntary loss of body weight that affects many cancer patients and implies a poor prognosis, reducing both tolerance to and efficiency of anticancer therapies. Actual challenges in management of CCx remain in the identification of tumour‐derived and host‐derived mediators involved in systemic inflammation and tissue wasting and in the discovery of biomarkers that would allow for an earlier and personalized care of cancer patients. The aim of this study was to identify new markers of CCx across different species and tumour entities. Methods Quantitative secretome analysis was performed to identify specific factors characteristic of cachexia‐inducing cancer cell lines. To establish the subsequently identified phospholipase PLA2G7 as a marker of CCx, plasma PLA2G7 activity and/or protein levels were measured in well‐established mouse models of CCx and in different cohorts of weight‐stable and weight‐losing cancer patients with different tumour entities. Genetic PLA2G7 knock‐down in tumours and pharmacological treatment using the well‐studied PLA2G7 inhibitor darapladib were performed to assess its implication in the pathogenesis of CCx in C26 tumour‐bearing mice. Results High expression and secretion of PLA2G7 were hallmarks of cachexia‐inducing cancer cell lines. Circulating PLA2G7 activity was increased in different mouse models of CCx with various tumour entities and was associated with the severity of body wasting. Circulating PLA2G7 levels gradually rose during cachexia development. Genetic PLA2G7 knock‐down in C26 tumours only partially reduced plasma PLA2G7 levels, suggesting that the host is also an important contributor. Chronic treatment with darapladib was not sufficient to counteract inflammation and tissue wasting despite a strong inhibition of the circulating PLA2G7 activity. Importantly, PLA2G7 levels were also increased in colorectal and pancreatic cancer patients with CCx. Conclusions Overall, our data show that despite no immediate pathogenic role, at least when targeted as a single entity, PLA2G7 is a consistent marker of CCx in both mice and humans. The early increase in circulating PLA2G7 levels in pre‐cachectic mice supports future prospective studies to assess its potential as biomarker for cancer patients.

Background Cancer cachexia (CCx) is a multifactorial energy‐wasting syndrome reducing the efficiency of anti‐cancer therapies, quality of life, and survival of cancer patients. In the past years, most studies focused on the identification of tumour and host‐derived proteins contributing to CCx. However, there is still a lack of studies addressing the changes in bioactive lipids. The aim of this study was to identify specific lipid species as a hallmark of CCx by performing a broad range lipid analysis of plasma from well‐established CCx mouse models as well as cachectic and weight stable cancer patients. Methods Plasma from non‐cachectic (PBS‐injected mice, NC26 tumour‐bearing mice), pre‐cachectic and cachectic mice (C26 and LLC tumour‐bearing mice, ApcMin/+ mutant mice), and plasma from weight stable and cachectic patients with gastrointestinal cancer, were analysed using the Lipidyzer™ platform. In total, 13 lipid classes and more than 1100 lipid species, including sphingolipids, neutral and polar glycerolipids, were covered by the analysis. Correlation analysis between specific lipid species and readouts of CCx were performed. Lipidomics data were confirmed by gene expression analysis of metabolic organs to analyse enzymes involved in sphingolipid synthesis and degradation. Results A decrease in several lysophosphatidylcholine (LPC) species and an increase in numerous sphingolipids including sphingomyelins (SMs), ceramides (CERs), hexosyl‐ceramides (HCERs) and lactosyl‐ceramides (LCERs), were mutual features of CCx in both mice and cancer patients. Notably, sphingolipid levels gradually increased during cachexia development. Key enzymes involved in ceramide synthesis were elevated in liver but not in adipose, muscle, or tumour tissues, suggesting that ceramide turnover in the liver is a major contributor to elevated sphingolipid levels in CCx. LPC(16:1), LPC(20:3), SM(16:0), SM(24:1), CER(16:0), CER(24:1), HCER(16:0), and HCER(24:1) were the most consistently affected lipid species between mice and humans and correlated negatively (LPCs) or positively (SMs, CERs and HCERs) with the severity of body weight loss. Conclusions High levels of sphingolipids, specifically ceramides and modified ceramides, are a defining feature of murine and human CCx and may contribute to tissue wasting and skeletal muscle atrophy through the inhibition of anabolic signals. The progressive increase in sphingolipids during cachexia development supports their potential as early biomarkers for CCx.

International audience; In mammals, the white adipocyte is a cell type that is specialized for storage of energy (in the form of triacylglycerols) and for energy mobilization (as fatty acids). White adipocyte metabolism confers an essential role to adipose tissue in whole-body homeostasis. Dysfunction in white adipocyte metabolism is a cardinal event in the development of insulin resistance and associated disorders. This Review focuses on our current understanding of lipid and glucose metabolic pathways in the white adipocyte. We survey recent advances in humans on the importance of adipocyte hypertrophy and on the in vivo turnover of adipocytes and stored lipids. At the molecular level, the identification of novel regulators and of the interplay between metabolic pathways explains the fine-tuning between the anabolic and catabolic fates of fatty acids and glucose in different physiological states. We also examine the metabolic alterations involved in the genesis of obesity-associated metabolic disorders, lipodystrophic states, cancers and cancer-associated cachexia. New challenges include defining the heterogeneity of white adipocytes in different anatomical locations throughout the lifespan and investigating the importance of rhythmic processes. Targeting white fat metabolism offers opportunities for improved patient stratification and a wide, yet unexploited, range of therapeutic opportunities.

By accentuating drug efficacy and impeding resistance mechanisms, combinatorial, multi‐agent therapies have emerged as key approaches in the treatment of complex diseases, most notably cancer. Using high‐throughput drug screens, we uncovered distinct metabolic vulnerabilities and thereby identified drug combinations synergistically causing a starvation‐like lethal catabolic response in tumor cells from different cancer entities. Domperidone, a dopamine receptor antagonist, as well as several tricyclic antidepressants (TCAs), including imipramine, induced cancer cell death in combination with the mitochondrial uncoupler niclosamide ethanolamine (NEN) through activation of the integrated stress response pathway and the catabolic CLEAR network. Using transcriptome and metabolome analyses, we characterized a combinatorial response, mainly driven by the transcription factors CHOP and TFE3, which resulted in cell death through enhanced pyrimidine catabolism as well as reduced pyrimidine synthesis. Remarkably, the drug combinations sensitized human organoid cultures to the standard‐of‐care chemotherapy paclitaxel. Thus, our combinatorial approach could be clinically implemented into established treatment regimen, which would be further facilitated by the advantages of drug repurposing., This study identifies novel combinatorial drug treatments to induce death of different tumor cells, and defines the mechanisms of synergism between a mitochondrial uncoupler and antidepressants or dopamine receptor antagonists.

International audience; No abstract available

Simple Summary Cachexia is a deadly disease that accompanies many different types of cancers. Animal studies on cachexia have so far mostly been conducted using young mice, while cancer in humans is a disease of high age. Mouse models used to date may therefore not be suitable to study cachexia with relevance to patients. By comparing young and old mice of three different strains and two different tumor types, we here show that the age of mice has a substantial effect on cachexia progression (specifically body weight, tissue weight, fiber size, molecular markers) that is dependent on the mouse strain studied. This is independent of glucose tolerance. The cachexia markers IL6 and GDF15 differ between ages in both mice and patients. Future studies on cachexia should consider the age and strain of mice. Abstract Background: Cancer is primarily a disease of high age in humans, yet most mouse studies on cancer cachexia are conducted using young adolescent mice. Given that metabolism and muscle function change with age, we hypothesized that aging may affect cachexia progression in mouse models. Methods: We compare tumor and cachexia development in young and old mice of three different strains (C57BL/6J, C57BL/6N, BALB/c) and with two different tumor cell lines (Lewis Lung Cancer, Colon26). Tumor size, body and organ weights, fiber cross-sectional area, circulating cachexia biomarkers, and molecular markers of muscle atrophy and adipose tissue wasting are shown. We correlate inflammatory markers and body weight dependent on age in patients with cancer. Results: We note fundamental differences between mouse strains. Aging aggravates weight loss in LLC-injected C57BL/6J mice, drives it in C57BL/6N mice, and does not influence weight loss in C26-injected BALB/c mice. Glucose tolerance is unchanged in cachectic young and old mice. The stress marker GDF15 is elevated in cachectic BALB/c mice independent of age and increased in old C57BL/6N and J mice. Inflammatory markers correlate significantly with weight loss only in young mice and patients. Conclusions: Aging affects cachexia development and progression in mice in a strain-dependent manner and influences the inflammatory profile in both mice and patients. Age is an important factor to consider for future cachexia studies.

Obesity-induced insulin resistance is a major risk factor for the development of type 2 diabetes. Basal fat cell lipolysis (i.e., fat cell triacylglycerol breakdown into fatty acids and glycerol in the absence of stimulatory factors) is elevated during obesity and is closely associated with insulin resistance. Inhibition of adipocyte lipolysis may therefore be a promising therapeutic strategy for treating insulin resistance and preventing obesity-associated type 2 diabetes. In this review, we explore the relationship between adipose lipolysis and insulin sensitivity. After providing an overview of the components of fat cell lipolytic machinery, we describe the hypotheses that may support the causality between lipolysis and insulin resistance. Excessive circulating fatty acids may ectopically accumulate in insulin-sensitive tissues and impair insulin action. Increased basal lipolysis may also modify the secretory profile of adipose tissue, influencing whole body insulin sensitivity. Finally, excessive fatty acid release may also worsen adipose tissue inflammation, a well-known parameter contributing to insulin resistance. Partial genetic or pharmacologic inhibition of fat cell lipases in mice as well as short term clinical trials using antilipolytic drugs in humans support the benefit of fat cell lipolysis inhibition on systemic insulin sensitivity and glucose metabolism, which occurs without an increase of fat mass. Modulation of fatty acid fluxes and, putatively, of fat cell secretory pattern may explain the amelioration of insulin sensitivity whereas changes in adipose tissue immune response do not seem involved.

Introduction Bien qu'une association robuste entre une diminution des niveaux circulants de peptides natriuretiques (PN) et un risque accru de diabete de type 2 a ete rapportee chez l'homme, il n'y a, a ce jour, aucune evidence directe d'une effet des PN sur la sensibilite a l'insuline globale ou dans un tissu cible. Materiels et Methodes L'expression genique des recepteurs aux PN a ete mesuree par RT-qPCR dans des biopsies de tissu adipeux sous-cutane de deux cohortes europeennes independantes. Le transport de glucose a ete mesure sur des adipocytes humains isoles et sur des adipocytes hMADS ( human multipotent adipose-derived stem cell ) en culture. Les experiences de flux metaboliques et de signalisation ont ete effectuees sur hMADS en culture. Resultats Nous demontrons ici que l'expression du recepteur biologiquement actif aux PN de type A (NPRA) diminue tandis que celle du recepteur de clairance de type C (NPRC) augmente avec le degre d'obesite. L'expression du NPRA correle negativement avec la glycemie a jeun et est diminue chez des individus pre-diabetiques et diabetiques de type 2. L'expression du NPRA correle aussi negativement avec l'insulino-resistance (r=− 0,65, p Conclusions Collectivement, nos donnees supportent un role des PN dans la regulation du metabolisme du glucose et de la sensibilite a l'insuline en stimulant le transport de glucose dans l'adipocyte humain. La signalisation aux PN pourrait constituer une cible potentielle pour ameliorer l'insulinoresistance associee a l'obesite.

Introduction L’inhibition de la lipolyse adipocytaire ameliore la sensibilite a l’insuline in vivo et in vitro. Au vu des connaissances actuelles, nous avons pose l’hypothese que l’amelioration de la reponse a l’insuline observee lors d’une inhibition de la Lipase hormono-sensible (LHS) est dirigee par l’augmentation de la voie de la lipogenese de novo (LDN), sous le controle du facteur de transcription ChREBP. Materiels et methodes Dans des preadipocytes humains en culture (cellules hMADS), l’inhibition de l’expression de la LHS (siLHS), du facteur ChREBP (siChREBP) ou des deux (siLHS/siChREBP) est realisee a l’aide de petits ARN interferents. Le metabolisme du glucose est evalue a l’aide de glucose radiomarque. L’expression des differentes enzymes dirigeant la LDN a ete mesuree par RT-qPCR. La composition intra-cellulaire en acides gras a ete determinee par chromatographie en phase gazeuse. Enfin, la transmission du signal insulinique a ete evaluee par Western Blot. Resultats Les adipocytes siLHS presentent une amelioration du transport et de l’oxydation du glucose ainsi que de la LDN en reponse a l’insuline en parallele a une forte augmentation de l’expression du facteur ChREBP. Ces cellules ont aussi une meilleure transmission du signal insulinique. La perte de ces effets dans les adipocytes siLHS/siChREBP suggere un role important de ce facteur dans les effets medies par l’inhibition de la LHS. L’amelioration de la reponse a l’insuline des adipocytes siLHS pourrait s’expliquer par une augmentation de l’oleate au detriment du palmitoleate qui est partiellement perdue dans les cellules siLHS/siChREBP. Une expression differentielle de l’elongase ELOVL6 pourrait notamment expliquer ces differences. Conclusion Ces resultats mettent en evidence l’implication de la LDN, induite par l’augmentation du facteur ChREBP, dans l’amelioration de la sensibilite a l’insuline adipocytaire observee lors d’une inhibition de la LHS. Declaration d’interet Les auteurs declarent ne pas avoir d’interet direct ou indirect (financier ou en nature) avec un organisme prive, industriel ou commercial en relation avec le sujet presente.