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Objectif: L’obésité est caractérisée par l’expansion excessive du tissu adipeux blanc associée à une inflammation chronique à ű bas bruit Ƈ initiée par le recrutement de cellules immunes dans le tissu. L’adipogenèse et la fonction des cellules immunes sont régulés par la voie du ű mechanistic Target Of Rapamycin Ƈ (mTOR). La voie mTOR est sur-activée lors de l’obésité, participant au développement de la résistance à l’insuline et du diabète de type 2. Ainsi la rapamycine, inhibiteur sélectif de la voie mTOR, est proposée comme une nouvelle piste thérapeutique dans le traitement des maladies métaboliques. Notre objectif est de préciser les effets métaboliques de la rapamycine et de définir s’ils sont associés à des altérations immunes et/ou inflammatoires. Matériels et méthodes: Des souris mâles C57BL/6J mises sous régime hyperlipidique reçoivent des injections hebdomadaires de rapamycine ou d’excipient. Les paramètres métaboliques et immuns sont suivis par l’utilisation de techniques de biologie moléculaire (Q-PCR, transcriptomique), biochimie (ELISAs, western-blot), cytométrie en flux et histologie. Résultats: Bien que limitant la prise de poids corporel, la rapamycine aggrave l’intolérance au glucose. Le recrutement massif de macrophages dans le tissu adipeux ainsi que la production accrue de cytokines inflammatoires (IL-6, MCP-1) chez les souris traitées pourraient expliquer l’effet délétère de la drogue sur l’homéostasie glucidique. Par ailleurs, l’augmentation du nombre des cellules immuno-régulatrices Gr1+CD11b+ (Myeloïd-derived suppressor cells ou MDSC) dans le tissu adipeux, pourrait expliquer le maintien de la sensibilité à l’insuline des souris traitées ; les MDSCs ayant récemment été décrites comme modérant l’inflammation et, conséquemment, améliorant la sensibilité à l’insuline de souris obèses. Conclusion: Notre étude suggère que l’effet paradoxal de la rapamycine sur le métabolisme énergétique pourrait résulter d’un contrôle exercé par la voie mTOR sur les cellules myéloïdes régulant l’inflammation du tissu adipeux blanc (macrophages et MDSCs). [ABSTRACT FROM AUTHOR]

Background: Cancer heterogeneity is a main obstacle for the development of effective therapies, as its replication in in vitro preclinical models is challenging. Around 96% of developed drugs are estimated to fail from discovery to the clinical trial phase probably because of the unsuitability and unreliability of current preclinical models (Front Pharmacol 9:6, 2018; Nat Rev Cancer 8: 147-56, 2008) in replicating the overall biology of tumors, for instance the tumor microenvironment. Breast cancer is the most frequent cancer among women causing the greatest number of cancer-related deaths. Breast cancer can typically be modeled in vitro through the use of tumoroids; however, current approaches using mouse tumoroids fail to reproduce crucial aspect of human breast cancer, while access to human cells is limited and the focus of ethical concerns. New models of breast cancer, such as companion dogs, have emerged given the resemblance of developed spontaneous mammary tumors to human breast cancer in many clinical and molecular aspects; however, they have so far failed to replicate the tumor microenvironment. The present work aimed at developing a robust canine mammary tumor model in the form of tumoroids which recapitulate the tumor diversity and heterogeneity., Results: We conducted a complete characterization of canine mammary tumoroids through histologic, molecular, and proteomic analysis, demonstrating their strong similarity to the primary tumor. We demonstrated that these tumoroids can be used as a drug screening model. In fact, we showed that paclitaxel, a human chemotherapeutic, could kill canine tumoroids with the same efficacy as human tumoroids with 0.1 to 1 μM of drug needed to kill 50% of the cells. Due to easy tissue availability, canine tumoroids can be produced at larger scale and cryopreserved to constitute a biobank. We have demonstrated that cryopreserved tumoroids keep the same histologic and molecular features (ER, PR, and HER2 expression) as fresh tumoroids. Furthermore, two cryopreservation techniques were compared from a proteomic point of view which showed that tumoroids made from frozen material allowed to maintain the same molecular diversity as from freshly dissociated tumor., Conclusions: These findings revealed that canine mammary tumoroids can be easily generated and may provide an adequate and more reliable preclinical model to investigate tumorigenesis mechanisms and develop new treatments for both veterinary and human medicine., (© 2023. The Author(s).)

A rare but severe complication of curative-intent radiation therapy is the induction of second primary cancers. These cancers preferentially develop not inside the planning target volume (PTV) but around, over several centimeters, after a latency period of 1-40 years. We show here that normal human or mouse dermal fibroblasts submitted to the out-of-field dose scattering at the margin of a PTV receiving a mimicked patient's treatment do not die but enter in a long-lived senescent state resulting from the accumulation of unrepaired DNA single-strand breaks, in the almost absence of double-strand breaks. Importantly, a few of these senescent cells systematically and spontaneously escape from the cell cycle arrest after a while to generate daughter cells harboring mutations and invasive capacities. These findings highlight single-strand break-induced senescence as the mechanism of second primary cancer initiation, with clinically relevant spatiotemporal specificities. Senescence being pharmacologically targetable, they open the avenue for second primary cancer prevention., Competing Interests: EG, MT, CF, NM, EB, JB, Cd, JN, LS, CD, PB, AV, OM, LP, GG, OP, AP, FC, EL, NP, CA No competing interests declared, (© 2022, Goy et al.)

Background: Formalin-fixed paraffin-embedded (FFPE) tissue has been the gold standard for routine pathology for general and cancer postoperative diagnostics. Despite robust histopathology, immunohistochemistry, and molecular methods, accurate diagnosis remains difficult for certain cases. Overall, the entire process can be time consuming, labor intensive, and does not reach over 90% diagnostic sensitivity and specificity. There is a growing need in onco-pathology for adjunct novel rapid, accurate, reliable, diagnostically sensitive, and specific methods for high-throughput biomolecular identification. Lipids have long been considered only as building blocks of cell membranes or signaling molecules, but have recently been introduced as central players in cancer. Due to sample processing, which limits their detection, lipid analysis directly from unprocessed FFPE tissues has never been reported., Methods: We present a proof-of-concept with direct analysis of tissue-lipidomic signatures from FFPE tissues without dewaxing and minimal sample preparation using water-assisted laser desorption ionization mass spectrometry and deep-learning., Results: On a cohort of difficult canine and human sarcoma cases, classification for canine sarcoma subtyping was possible with 99.1% accuracy using "5-fold" and 98.5% using "leave-one-patient out," and 91.2% accuracy for human sarcoma using 5-fold and 73.8% using leave-one-patient out. The developed classification model enabled stratification of blind samples in <5 min and showed >95% probability for discriminating 2 human sarcoma blind samples., Conclusion: It is possible to create a rapid diagnostic platform to screen clinical FFPE tissues with minimal sample preparation for molecular pathology., (© American Association for Clinical Chemistry 2021.)

Purpose: F14512 is an epipodophyllotoxin derivative from etoposide, combined with a spermine moiety introduced as a cell delivery vector. The objective of this study was to compare the safety and antitumor activity of F14512 and etoposide phosphate in dogs with spontaneous non-Hodgkin lymphoma (NHL) and to investigate the potential benefit of F14512 in P-glycoprotein (Pgp) overexpressing lymphomas. Experimental Design: Forty-eight client-owned dogs with intermediate to high-grade NHL were enrolled into a randomized, double-blind trial of F14512 versus etoposide phosphate. Endpoints included safety and therapeutic efficacy. Results: Twenty-five dogs were randomized to receive F14512 and 23 dogs to receive etoposide phosphate. All adverse events (AEs) were reversible, and no treatment-related death was reported. Hematologic AEs were more severe with F14512 and gastrointestinal AEs were more frequent with etoposide phosphate. F14512 exhibited similar response rate and progression-free survival (PFS) as etoposide phosphate in the global treated population. Subgroup analysis of dogs with Pgp-overexpressing NHL showed a significant improvement in PFS in dogs treated with F14512 compared with etoposide phosphate. Conclusion: F14512 showed strong therapeutic efficacy against spontaneous NHL and exhibited a clinical benefice in Pgp-overexpressing lymphoma superior to etoposide phosphate. The results clearly justify the evaluation of F14512 in human clinical trials., Competing Interests: CONFLICTS OF INTEREST The authors declare no potential conflicts of interest., (Copyright: © 2020 Boyé et al.)

During normal mammary gland development, s-SHIP promoter expression marks a distinct type of mammary stem cells, at two different stages, puberty and early mid-pregnancy. To determine whether s-SHIP is a marker of mammary cancer stem cells (CSCs), we generated bitransgenic mice by crossing the C3(1)-SV40 T-antigen transgenic mouse model of breast cancer, and a transgenic mouse (11.5kb-GFP) expressing green fluorescent protein from the s-SHIP promoter. Here we show that in mammary tumors originating in these bitransgenic mice, s-SHIP promoter expression enriches a rare cell population with CSC activity as demonstrated by sphere-forming assays in vitro and limiting dilution transplantation in vivo. These s-SHIP-positive CSCs are characterized by lower expression of Delta-like non-canonical Notch ligand 1 (DLK1), a negative regulator of the Notch pathway. Inactivation of Dlk1 in s-SHIP-negative tumor cells increases their tumorigenic potential, suggesting a role for DLK1 in mammary cancer stemness., (Copyright © 2019 The Authors. Published by Elsevier Inc. All rights reserved.)

Background & Aims: Although the role of inflammation to combat infection is known, the contribution of metabolic changes in response to sepsis is poorly understood. Sepsis induces the release of lipid mediators, many of which activate nuclear receptors such as the peroxisome proliferator-activated receptor (PPAR)α, which controls both lipid metabolism and inflammation. We aimed to elucidate the previously unknown role of hepatic PPARα in the response to sepsis., Methods: Sepsis was induced by intraperitoneal injection of Escherichia coli in different models of cell-specific Ppara-deficiency and their controls. The systemic and hepatic metabolic response was analyzed using biochemical, transcriptomic and functional assays. PPARα expression was analyzed in livers from elective surgery and critically ill patients and correlated with hepatic gene expression and blood parameters., Results: Both whole body and non-hematopoietic Ppara-deficiency in mice decreased survival upon bacterial infection. Livers of septic Ppara-deficient mice displayed an impaired metabolic shift from glucose to lipid utilization resulting in more severe hypoglycemia, impaired induction of hyperketonemia and increased steatosis due to lower expression of genes involved in fatty acid catabolism and ketogenesis. Hepatocyte-specific deletion of PPARα impaired the metabolic response to sepsis and was sufficient to decrease survival upon bacterial infection. Hepatic PPARA expression was lower in critically ill patients and correlated positively with expression of lipid metabolism genes, but not with systemic inflammatory markers., Conclusion: During sepsis, Ppara-deficiency in hepatocytes is deleterious as it impairs the adaptive metabolic shift from glucose to FA utilization. Metabolic control by PPARα in hepatocytes plays a key role in the host defense against infection., Lay Summary: As the main cause of death in critically ill patients, sepsis remains a major health issue lacking efficacious therapies. While current clinical literature suggests an important role for inflammation, metabolic aspects of sepsis have mostly been overlooked. Here, we show that mice with an impaired metabolic response, due to deficiency of the nuclear receptor PPARα in the liver, exhibit enhanced mortality upon bacterial infection despite a similar inflammatory response, suggesting that metabolic interventions may be a viable strategy for improving sepsis outcomes., (Copyright © 2019 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.)

Background: Metastatic melanoma is one of the most aggressive forms of cancer in humans. Among its types, mucosal melanomas represent one of the most highly metastatic and aggressive forms, with a very poor prognosis. Because they are rare in Caucasian individuals, unlike cutaneous melanomas, there has been fewer epidemiological, clinical and genetic evaluation of mucosal melanomas. Moreover, the lack of predictive models fully reproducing the pathogenesis and molecular alterations of mucosal melanoma makes its treatment challenging. Interestingly, dogs are frequently affected by melanomas of the oral cavity that are characterized, as their human counterparts, by focal infiltration, recurrence, and metastasis to regional lymph nodes, lungs and other organs. In dogs, some particular breeds are at high risk, suggesting a specific genetic background and strong genetic drivers. Altogether, the striking homologies in clinical presentation, histopathological features, and overall biology between human and canine mucosal melanomas make dogs invaluable natural models with which to investigate tumor development, including tumor ætiology, and develop tailored treatments., Methods: We developed and characterized two canine oral melanoma cell lines from tumors isolated from dog patients with distinct clinical profiles; with and without lung metastases. The cells were characterized using immunohistochemistry, pharmacology and genetic studies., Results: We have developed and immunohistochemically, genetically, and pharmacologically characterized. Two cell lines (Ocr&#95;OCMM1X & Ocr&#95;OCMM2X) were produced through mouse xenografts originating from two clinically contrasting melanomas of the oral cavity. Their exhaustive characterization showed two distinct biological and genetic profiles that are potentially linked to the stage of malignancy at the time of diagnosis and sample collection of each melanoma case. These cell lines thus constitute relevant tools with which to perform genetic and drug screening analyses for a better understanding of mucosal melanomas in dogs and humans., Conclusions: The aim of this study was to establish and characterize xenograft-derived canine melanoma cell lines with different morphologies, genetic features and pharmacological sensitivities that constitute good predictive models for comparative oncology. These cell lines are relevant tools to advance the use of canine mucosal melanomas as natural models for the benefit of both veterinary and human medicine.

The genomic CDKN2A/B locus, encoding p16 INK4a among others, is linked to an increased risk for cardiovascular disease and type 2 diabetes. Obesity is a risk factor for both cardiovascular disease and type 2 diabetes. p16 INK4a is a cell cycle regulator and tumour suppressor. Whether it plays a role in adipose tissue formation is unknown. p16 INK4a knock-down in 3T3/L1 preadipocytes or p16 INK4a deficiency in mouse embryonic fibroblasts enhanced adipogenesis, suggesting a role for p16 INK4a in adipose tissue formation. p16 INK4a -deficient mice developed more epicardial adipose tissue in response to the adipogenic peroxisome proliferator activated receptor gamma agonist rosiglitazone. Additionally, adipose tissue around the aorta from p16 INK4a -deficient mice displayed enhanced rosiglitazone-induced gene expression of adipogenic markers and stem cell antigen, a marker of bone marrow-derived precursor cells. Mice transplanted with p16 INK4a -deficient bone marrow had more epicardial adipose tissue compared to controls when fed a high-fat diet. In humans, p16 INK4a gene expression was enriched in epicardial adipose tissue compared to other adipose tissue depots. Moreover, epicardial adipose tissue from obese humans displayed increased expression of stem cell antigen compared to lean controls, supporting a bone marrow origin of epicardial adipose tissue. These results show that p16 INK4a modulates epicardial adipose tissue development, providing a potential mechanistic link between the genetic association of the CDKN2A/B locus and cardiovascular disease risk.

Comparative oncology has shown that naturally occurring canine cancers are of valuable and translatable interest for the understanding of human cancer biology and the characterization of new therapies. This work was part of a comparative oncology project assessing a new, clinical-stage topoisomerase II inhibitor and comparing it with etoposide in dogs with spontaneous lymphoma with the objective to translate findings from dogs to humans. Etoposide is a topoisomerase II inhibitor widely used in various humans' solid and hematopoietic cancer, but little data is available concerning its potential antitumor efficacy in dogs. Etoposide phosphate is a water-soluble prodrug of etoposide which is expected to be better tolerated in dogs. The objectives of this study were to assess the safety, the tolerability and the efficacy of intravenous etoposide phosphate in dogs with multicentric lymphoma. Seven dose levels were evaluated in a traditional 3+3 phase I design. Twenty-seven owned-dogs with high-grade multicentric lymphoma were enrolled and treated with three cycles of etoposide phosphate IV injections every 2 weeks. Adverse effects were graded according to the Veterinary Cooperative Oncology Group criteria. A complete end-staging was realized 45 days after inclusion. The maximal tolerated dose was 300 mg/m2. At this dose level, the overall response rate was 83.3% (n = 6, 3 PR and 2 CR). Only a moderate reversible gastrointestinal toxicity, no severe myelotoxicity and no hypersensitivity reaction were reported at this dose level. Beyond the characterization of etoposide clinical efficacy in dogs, this study underlined the clinical and therapeutic homologies between dog and human lymphomas.

Background: Epidemiologic and clinical observations identify obesity as an important risk factor for asthma exacerbation, but the underlying mechanisms remain poorly understood. Type 2 innate lymphoid cells (ILC2s) and type 3 innate lymphoid cells (ILC3s) have been implicated, respectively, in asthma and adipose tissue homeostasis and in obesity-associated airway hyperresponsiveness (AHR)., Objective: We sought to determine the potential involvement of innate lymphoid cells (ILCs) in allergic airway disease exacerbation caused by high-fat diet (HFD)-induced obesity., Methods: Obesity was induced by means of HFD feeding, and allergic airway inflammation was subsequently induced by means of intranasal administration of house dust mite (HDM) extract. AHR, lung and visceral adipose tissue inflammation, humoral response, cytokines, and innate and adaptive lymphoid populations were analyzed in the presence or absence of ILCs., Results: HFD feeding exacerbated allergic airway disease features, including humoral response, airway and tissue eosinophilia, AHR, and T H 2 and T H 17 pulmonary profiles. Notably, nonsensitized obese mice already exhibited increased lung ILC counts and tissue eosinophil infiltration compared with values in lean mice in the absence of AHR. The numbers of total and cytokine-expressing lung ILC2s and ILC3s further increased in HDM-challenged obese mice compared with those in HDM-challenged lean mice, and this was accompanied by high IL-33 and IL-1β levels and decreased ILC markers in visceral adipose tissue. Furthermore, depletion of ILCs with an anti-CD90 antibody, followed by T-cell reconstitution, led to a profound decrease in allergic airway inflammatory features in obese mice, including T H 2 and T H 17 infiltration., Conclusion: These results indicate that HFD-induced obesity might exacerbate allergic airway inflammation through mechanisms involving ILC2s and ILC3s., (Copyright © 2016 American Academy of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.)

Purpose: F14512 is a new topoisomerase II inhibitor containing a spermine moiety that facilitates selective uptake by tumor cells and increases topoisomerase II poisoning. F14512 is currently in a phase I/II clinical trial in patients with acute myeloid leukemia. The aim of this study was to investigate F14512 potential in a new clinical indication. Because of the many similarities between human and dog lymphomas, we sought to determine the tolerance, efficacy, pharmacokinetic/pharmacodynamic (PK/PD) relationship of F14512 in this indication, and potential biomarkers that could be translated into human trials., Experimental Design: Twenty-three dogs with stage III-IV naturally occurring lymphomas were enrolled in the phase I dose-escalation trial, which consisted of three cycles of F14512 i.v. injections. Endpoints included safety and therapeutic efficacy. Serial blood samples and tumor biopsies were obtained for PK/PD and biomarker studies., Results: Five dose levels were evaluated to determine the recommended dose. F14512 was well tolerated, with the expected dose-dependent hematologic toxicity. F14512 induced an early decrease of tumoral lymph node cells, and a high response rate of 91% (21/23) with 10 complete responses, 11 partial responses, 1 stable disease, and 1 progressive disease. Phosphorylation of histone H2AX was studied as a potential PD biomarker of F14512., Conclusions: This trial demonstrated that F14512 can be safely administered to dogs with lymphoma resulting in strong therapeutic efficacy. Additional evaluation of F14512 is needed to compare its efficacy with standards of care in dogs, and to translate biomarker and efficacy findings into clinical trials in humans., (©2015 American Association for Cancer Research.)

Background: Remodeling of quiescent vessels with increases in permeability, vasodilatation, and edema are hallmarks of inflammatory disorders. Factors involved in this type of remodeling represent potential therapeutic targets., Objectives: We investigated whether the nuclear hormone receptor peroxisome proliferator-activated receptor (PPAR) β/δ, a regulator of metabolism, fibrosis, and skin homeostasis, is involved in regulation of this type of remodeling., Methods: Wild-type and various Pparb/d mutant mice were used to monitor dermal acute vascular hyperpermeability (AVH) and passive systemic anaphylaxis-induced hypothermia and edema. PPARβ/δ-dependent kinase activation and remodeling of endothelial cell-cell junctions were addressed by using human endothelial cells., Results: AVH and dilatation of dermal microvessels stimulated by vascular endothelial growth factor A, histamine, and thrombin are severely compromised in PPARβ/δ-deficient mice. Selective deletion of the Pparb/d-encoding gene in endothelial cells in vivo similarly limits dermal AVH and vasodilatation, providing evidence that endothelial PPARβ/δ is the major player in regulating acute dermal microvessel remodeling. Furthermore, endothelial PPARβ/δ regulatory functions are not restricted to the skin vasculature because its deletion in the endothelium, but not in smooth muscle cells, also leads to reduced systemic anaphylaxis, the most severe form of allergic reaction, in which an acute vascular response plays a key role. PPARβ/δ-dependent AVH activation likely involves the activation of mitogen-activated protein kinase and Akt pathways and leads to downstream destabilization of endothelial cell-cell junctions., Conclusion: These results unveil not only a novel function of PPARβ/δ as a direct regulator of acute vessel permeability and dilatation but also provide evidence that antagonizing PPARβ/δ represents an important strategy to consider for moderating diseases with altered endothelial integrity, such as acute inflammatory and allergic disorders., (Copyright © 2014 American Academy of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.)

Type 2 diabetes (T2D) is hallmarked by insulin resistance, impaired insulin secretion, and increased hepatic glucose production. The worldwide increasing prevalence of T2D calls for efforts to understand its pathogenesis in order to improve disease prevention and management. Recent genome-wide association studies have revealed strong associations between the CDKN2A/B locus and T2D risk. The CDKN2A/B locus contains genes encoding cell cycle inhibitors, including p16(Ink4a), which have not yet been implicated in the control of hepatic glucose homeostasis. Here, we show that p16(Ink4a) deficiency enhances fasting-induced hepatic glucose production in vivo by increasing the expression of key gluconeogenic genes. p16(Ink4a) downregulation leads to an activation of PKA-CREB-PGC1α signaling through increased phosphorylation of PKA regulatory subunits. Taken together, these results provide evidence that p16(Ink4a) controls fasting glucose homeostasis and could as such be involved in T2D development., (© 2014 by the American Diabetes Association. Readers may use this article as long as the work is properly cited, the use is educational and not for profit, and the work is not altered.)

Context: Obesity is characterized by the excessive accumulation of dysfunctional white adipose tissue (WAT), leading to a strong perturbation of metabolic regulations. However, the molecular events underlying this process are not fully understood., Objective: MicroRNAs (miRNAs) are small noncoding RNAs acting as posttranscriptional regulators of gene expression in multiple tissues and organs. However, their expression and roles in WAT cell subtypes, which include not only adipocytes but also immune, endothelial, and mesenchymal stem cells as well as preadipocytes, have not been characterized. Design/Results: By applying differential miRNome analysis, we demonstrate that the expression of several miRNAs is dysregulated in epididymal WAT from ob/ob and high-fat diet-fed mice. Adipose tissue-specific down-regulation of miR-200a and miR-200b and the up-regulation of miR-342-3p, miR-335-5p, and miR-335-3p were observed. Importantly, a similarly altered expression of miR-200a and miR-200b was observed in obese diabetic patients. Furthermore, cell fractionation of mouse adipose tissue revealed that miRNAs are differentially expressed in adipocytes and in subpopulations from the stromal vascular fraction. Finally, integration of transcriptomic data showed that bioinformatically predicted miRNA target genes rarely showed anticorrelated expression with that of targeting miRNA, in contrast to experimentally validated target genes., Conclusion: Taken together, our data indicate that the dysregulated expression of miRNAs occurs in distinct cell types and is likely to affect cell-specific function(s) of obese WAT.

Atopic dermatitis (AD) is a chronic allergic dermatosis characterized by epidermal thickening and dermal inflammatory infiltrates with a dominant Th2 profile during the acute phase, whereas a Th1 profile is characteristic of the chronic stage. Among chemokines and chemokine receptors associated with inflammation, increased levels of CX3CL1 (fractalkine) and its unique receptor, CX3CR1, have been observed in human AD. We have thus investigated their role and mechanism of action in experimental models of AD and psoriasis. AD pathology and immune responses, but not psoriasis, were profoundly decreased in CX3CR1-deficient mice and upon blocking CX3CL1-CX3CR1 interactions in wild-type mice. CX3CR1 deficiency affected neither antigen presentation nor T cell proliferation in vivo upon skin sensitization, but CX3CR1 expression by both Th2 and Th1 cells was required to induce AD. Surprisingly, unlike in allergic asthma, where CX3CL1 and CX3CR1 regulate the pathology by controlling effector CD4(+) T cell survival within inflamed tissues, adoptive transfer experiments established CX3CR1 as a key regulator of CD4(+) T cell retention in inflamed skin, indicating a new function for this chemokine receptor. Therefore, although CX3CR1 and CX3CL1 act through distinct mechanisms in different pathologies, our results further indicate their interest as promising therapeutic targets in allergic diseases., (© 2014 Staumont-Sallé et al.)

Unlabelled: Bile acid metabolism is intimately linked to the control of energy homeostasis and glucose and lipid metabolism. The nuclear receptor farnesoid X receptor (FXR) plays a major role in the enterohepatic cycling of bile acids, but the impact of nutrients on bile acid homeostasis is poorly characterized. Metabolically active hepatocytes cope with increases in intracellular glucose concentrations by directing glucose into storage (glycogen) or oxidation (glycolysis) pathways, as well as to the pentose phosphate shunt and the hexosamine biosynthetic pathway. Here we studied whether the glucose nonoxidative hexosamine biosynthetic pathway modulates FXR activity. Our results show that FXR interacts with and is O-GlcNAcylated by O-GlcNAc transferase in its N-terminal AF1 domain. Increased FXR O-GlcNAcylation enhances FXR gene expression and protein stability in a cell type-specific manner. High glucose concentrations increased FXR O-GlcNAcylation, hence its protein stability and transcriptional activity by inactivating corepressor complexes, which associate in a ligand-dependent manner with FXR, and increased FXR binding to chromatin. Finally, in vivo fasting-refeeding experiments show that FXR undergoes O-GlcNAcylation in fed conditions associated with increased direct FXR target gene expression and decreased liver bile acid content., Conclusion: FXR activity is regulated by glucose fluxes in hepatocytes through a direct posttranslational modification catalyzed by the glucose-sensing hexosamine biosynthetic pathway., (© 2014 by the American Association for the Study of Liver Diseases.)

The "mechanistic target of rapamycin" (mTOR) is a central controller of growth, proliferation and/or motility of various cell-types ranging from adipocytes to immune cells, thereby linking metabolism and immunity. mTOR signaling is overactivated in obesity, promoting inflammation and insulin resistance. Therefore, great interest exists in the development of mTOR inhibitors as therapeutic drugs for obesity or diabetes. However, despite a plethora of studies characterizing the metabolic consequences of mTOR inhibition in rodent models, its impact on immune changes associated with the obese condition has never been questioned so far. To address this, we used a mouse model of high-fat diet (HFD)-fed mice with and without pharmacologic mTOR inhibition by rapamycin. Rapamycin was weekly administrated to HFD-fed C57BL/6 mice for 22 weeks. Metabolic effects were determined by glucose and insulin tolerance tests and by indirect calorimetry measures of energy expenditure. Inflammatory response and immune cell populations were characterized in blood, adipose tissue and liver. In parallel, the activities of both mTOR complexes (e. g. mTORC1 and mTORC2) were determined in adipose tissue, muscle and liver. We show that rapamycin-treated mice are leaner, have enhanced energy expenditure and are protected against insulin resistance. These beneficial metabolic effects of rapamycin were associated to significant changes of the inflammatory profiles of both adipose tissue and liver. Importantly, immune cells with regulatory functions such as regulatory T-cells (Tregs) and myeloid-derived suppressor cells (MDSCs) were increased in adipose tissue. These rapamycin-triggered metabolic and immune effects resulted from mTORC1 inhibition whilst mTORC2 activity was intact. Taken together, our results reinforce the notion that controlling immune regulatory cells in metabolic tissues is crucial to maintain a proper metabolic status and, more generally, comfort the need to search for novel pharmacological inhibitors of the mTOR signaling pathway to prevent and/or treat metabolic diseases.

The nuclear bile acid receptor farnesoid X receptor (FXR) is an important transcriptional regulator of bile acid, lipid, and glucose metabolism. FXR is highly expressed in the liver and intestine and controls the synthesis and enterohepatic circulation of bile acids. However, little is known about FXR-associated proteins that contribute to metabolic regulation. Here, we performed a mass spectrometry-based search for FXR-interacting proteins in human hepatoma cells and identified AMPK as a coregulator of FXR. FXR interacted with the nutrient-sensitive kinase AMPK in the cytoplasm of target cells and was phosphorylated in its hinge domain. In cultured human and murine hepatocytes and enterocytes, pharmacological activation of AMPK inhibited FXR transcriptional activity and prevented FXR coactivator recruitment to promoters of FXR-regulated genes. Furthermore, treatment with AMPK activators, including the antidiabetic biguanide metformin, inhibited FXR agonist induction of FXR target genes in mouse liver and intestine. In a mouse model of intrahepatic cholestasis, metformin treatment induced FXR phosphorylation, perturbed bile acid homeostasis, and worsened liver injury. Together, our data indicate that AMPK directly phosphorylates and regulates FXR transcriptional activity to precipitate liver injury under conditions favoring cholestasis.

Objective: A genomic region near the CDKN2A locus, encoding p16(INK4a), has been associated to type 2 diabetes and atherosclerotic vascular disease, conditions in which inflammation plays an important role. Recently, we found that deficiency of p16(INK4a) results in decreased inflammatory signaling in murine macrophages and that p16(INK4a) influences the phenotype of human adipose tissue macrophages. Therefore, we investigated the influence of immune cell p16(INK4a) on glucose tolerance and atherosclerosis in mice., Methods and Results: Bone marrow p16(INK4a)-deficiency in C57Bl6 mice did not influence high fat diet-induced obesity nor plasma glucose and lipid levels. Glucose tolerance tests showed no alterations in high fat diet-induced glucose intolerance. While bone marrow p16(INK4a)-deficiency did not affect the gene expression profile of adipose tissue, hepatic expression of the alternative markers Chi3l3, Mgl2 and IL10 was increased and the induction of pro-inflammatory Nos2 was restrained on the high fat diet. Bone marrow p16(INK4a)-deficiency in low density lipoprotein receptor-deficient mice did not affect western diet-induced atherosclerotic plaque size or morphology. In line, plasma lipid levels remained unaffected and p16(INK4a)-deficient macrophages displayed equal cholesterol uptake and efflux compared to wild type macrophages., Conclusion: Bone marrow p16(INK4a)-deficiency does not affect plasma lipids, obesity, glucose tolerance or atherosclerosis in mice.

The CDKN2A locus, which contains the tumor suppressor gene p16(INK4a), is associated with an increased risk of age-related inflammatory diseases, such as cardiovascular disease and type 2 diabetes, in which macrophages play a crucial role. Monocytes can polarize toward classically (CAMϕ) or alternatively (AAMϕ) activated macrophages. However, the molecular mechanisms underlying the acquisition of these phenotypes are not well defined. Here, we show that p16(INK4a) deficiency (p16(-/-)) modulates the macrophage phenotype. Transcriptome analysis revealed that p16(-/-) BM-derived macrophages (BMDMs) exhibit a phenotype resembling IL-4-induced macrophage polarization. In line with this observation, p16(-/-) BMDMs displayed a decreased response to classically polarizing IFNγ and LPS and an increased sensitivity to alternative polarization by IL-4. Furthermore, mice transplanted with p16(-/-) BM displayed higher hepatic AAMϕ marker expression levels on Schistosoma mansoni infection, an in vivo model of AAMϕ phenotype skewing. Surprisingly, p16(-/-) BMDMs did not display increased IL-4-induced STAT6 signaling, but decreased IFNγ-induced STAT1 and lipopolysaccharide (LPS)-induced IKKα,β phosphorylation. This decrease correlated with decreased JAK2 phosphorylation and with higher levels of inhibitory acetylation of STAT1 and IKKα,β. These findings identify p16(INK4a) as a modulator of macrophage activation and polarization via the JAK2-STAT1 pathway with possible roles in inflammatory diseases.

Objective: Bile acids (BA) participate in the maintenance of metabolic homeostasis acting through different signaling pathways. The nuclear BA receptor farnesoid X receptor (FXR) regulates pathways in BA, lipid, glucose, and energy metabolism, which become dysregulated in obesity. However, the role of FXR in obesity and associated complications, such as dyslipidemia and insulin resistance, has not been directly assessed., Research Design and Methods: Here, we evaluate the consequences of FXR deficiency on body weight development, lipid metabolism, and insulin resistance in murine models of genetic and diet-induced obesity., Results: FXR deficiency attenuated body weight gain and reduced adipose tissue mass in both models. Surprisingly, glucose homeostasis improved as a result of an enhanced glucose clearance and adipose tissue insulin sensitivity. In contrast, hepatic insulin sensitivity did not change, and liver steatosis aggravated as a result of the repression of β-oxidation genes. In agreement, liver-specific FXR deficiency did not protect from diet-induced obesity and insulin resistance, indicating a role for nonhepatic FXR in the control of glucose homeostasis in obesity. Decreasing elevated plasma BA concentrations in obese FXR-deficient mice by administration of the BA sequestrant colesevelam improved glucose homeostasis in a FXR-dependent manner, indicating that the observed improvements by FXR deficiency are not a result of indirect effects of altered BA metabolism., Conclusions: Overall, FXR deficiency in obesity beneficially affects body weight development and glucose homeostasis., (© 2011 by the American Diabetes Association.)

Weight gain and metabolic disturbances, such as dyslipidemia and hyperglycaemia, are common side effects of most antipsychotic drugs, including risperidone. The aim of this study was to investigate the effects of chronic treatment with risperidone on body weight, fat accumulation, liver weight, and hepatic expression of key genes involved in lipid metabolism in female mice. We also addressed the mechanism of risperidone induction of metabolic side effects by exploring its effect on lipid and cholesterol metabolism in primary cultures of rat hepatocytes. Eleven weeks of treatment with long-acting risperidone (12.5 mpk/week) resulted in a significant weight gain associated with an increase of liver and adipose tissue weight. These effects were positively correlated with hepatic mRNA induction of two key genes involved in lipogenesis: sterol regulatory element binding protein-1c (SREBP-1c) and fatty acid synthase (FAS). Furthermore, in line with these in vivo results, risperidone elicited significant inductions of SREBP-1 maturation and FAS mRNA expression in primary cultures of rat hepatocytes associated with an increase of free fatty acid, triacylglycerol, and phospholipid synthesis as assessed by acetate incorporation. The current investigations underscore the usefulness of a mouse model to study the weight gain observed with risperidone treatment in humans. This study shows that risperidone induces similar effects in the liver (in vivo) and in hepatocyte cell cultures (in vitro) on the expression of key genes and/or proteins that control lipid metabolism. This suggests that risperidone could alter lipid metabolism in the liver and induce weight gain in a way that is partly independent of its action on the central nervous system.

The bile acid receptor farnesoid X receptor (FXR) is expressed in adipose tissue, but its function remains poorly defined. Peroxisome proliferator-activated receptor-γ (PPARγ) is a master regulator of adipocyte differentiation and function. The aim of this study was to analyze the role of FXR in adipocyte function and to assess whether it modulates PPARγ action. Therefore, we tested the responsiveness of FXR-deficient mice (FXR(-/-)) and cells to the PPARγ activator rosiglitazone. Our results show that genetically obese FXR(-/-)/ob/ob mice displayed a resistance to rosiglitazone treatment. In vitro, rosiglitazone treatment did not induce normal adipocyte differentiation and lipid droplet formation in FXR(-/-) mouse embryonic fibroblasts (MEFs) and preadipocytes. Moreover, FXR(-/-) MEFs displayed both an increased lipolysis and a decreased de novo lipogenesis, resulting in reduced intracellular triglyceride content, even upon PPARγ activation. Retroviral-mediated FXR re-expression in FXR(-/-) MEFs restored the induction of adipogenic marker genes during rosiglitazone-forced adipocyte differentiation. The expression of Wnt/β-catenin pathway and target genes was increased in FXR(-/-) adipose tissue and MEFs. Moreover, the expression of several endogenous inhibitors of this pathway was decreased early during the adipocyte differentiation of FXR(-/-) MEFs. These findings demonstrate that FXR regulates adipocyte differentiation and function by regulating two counteracting pathways of adipocyte differentiation, the PPARγ and Wnt/β-catenin pathways.

Farnesoid X receptor (FXR) is highly expressed in liver and intestine where it controls bile acid (BA), lipid and glucose homeostasis. Here we show that FXR is expressed and functional, as assessed by target gene expression analysis, in human islets and beta-cell lines. FXR is predominantly cytosolic-localized in the islets of lean mice, but nuclear in obese mice. Compared to FXR+/+ mice, FXR-/- mice display a normal architecture and beta-cell mass but the expression of certain islet-specific genes is altered. Moreover, glucose-stimulated insulin secretion (GSIS) is impaired in the islets of FXR-/- mice. Finally, FXR activation protects human islets from lipotoxicity and ameliorates their secretory index., (Copyright 2010 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.)

Obese patients have chronic, low-grade inflammation that predisposes to type 2 diabetes and results, in part, from dysregulated visceral white adipose tissue (WAT) functions. The specific signaling pathways underlying WAT dysregulation, however, remain unclear. Here we report that the PPARgamma signaling pathway operates differently in the visceral WAT of lean and obese mice. PPARgamma in visceral, but not subcutaneous, WAT from obese mice displayed increased sensitivity to activation by its agonist rosiglitazone. This increased sensitivity correlated with increased expression of the gene encoding the ubiquitin hydrolase/ligase ubiquitin carboxyterminal esterase L1 (UCH-L1) and with increased degradation of the PPARgamma heterodimerization partner retinoid X receptor alpha (RXRalpha), but not RXRbeta, in visceral WAT from obese humans and mice. Interestingly, increased UCH-L1 expression and RXRalpha proteasomal degradation was induced in vitro by conditions mimicking hypoxia, a condition that occurs in obese visceral WAT. Finally, PPARgamma-RXRbeta heterodimers, but not PPARgamma-RXRalpha complexes, were able to efficiently dismiss the transcriptional corepressor silencing mediator for retinoid and thyroid hormone receptors (SMRT) upon agonist binding. Increasing the RXRalpha/RXRbeta ratio resulted in increased PPARgamma responsiveness following agonist stimulation. Thus, the selective proteasomal degradation of RXRalpha initiated by UCH-L1 upregulation modulates the relative affinity of PPARgamma heterodimers for SMRT and their responsiveness to PPARgamma agonists, ultimately activating the PPARgamma-controlled gene network in visceral WAT of obese animals and humans.

The underlying mechanisms of the ketogenic diet (KD) remain unknown. Involvement of peroxisome proliferator-activated receptor-alpha (PPARalpha) has been suggested. The aim of this study was to assess the anticonvulsant properties of fenofibrate, a PPARalpha agonist. Wistar rats were fed at libitum during 14 days by regular diet, KD, regular diet containing 0.2% fenofibrate (F), or KD containing 0.2% fenofibrate (KD + F). Pentylenetetrazol (PTZ) threshold and latencies to the onset of status epilepticus induced by lithium-pilocarpine were used to assess diet treatments with anticonvulsive effects. Myoclonic and generalized seizure PTZ thresholds were increased in F- and KD-treated animals in comparison to control. No difference was observed between KD + F group and the others groups (control, F, KD). Latencies to the onset of status epilepticus were increased in F and KD groups compared to control. Fenofibrate exerts anticonvulsive properties comparable to KD in adult rats using PTZ and lithium-pilocarpine models. The underlying mechanisms such as PPARalpha activation and others should be investigated. These findings may provide insights into future directions to simplify KD protocols.

Androgens are major regulators of prostate cell growth and physiology. In the human prostate, androgens are inactivated in the form of hydrophilic glucuronide conjugates. These metabolites are formed by the two human UGT2B15 [UGT (UDP-glucuronosyltransferase) 2B15] and UGT2B17 enzymes. The FXR (farnesoid X receptor) is a bile acid sensor controlling hepatic and/or intestinal cholesterol, lipid and glucose metabolism. In the present study, we report the expression of FXR in normal and cancer prostate epithelial cells, and we demonstrate that its activation by chenodeoxycholic acid or GW4064 negatively interferes with the levels of UGT2B15 and UGT2B17 mRNA and protein in prostate cancer LNCaP cells. FXR activation also causes a drastic reduction of androgen glucuronidation in these cells. These results point out activators of FXR as negative regulators of androgen-conjugating UGT expression in the prostate. Finally, the androgen metabolite androsterone, which is also an activator of FXR, dose-dependently reduces the glucuronidation of androgens catalysed by UGT2B15 and UGT2B17 in an FXR-dependent manner in LNCaP cells. In conclusion, the present study identifies for the first time the activators of FXR as important regulators of androgen metabolism in human prostate cancer cells.

The role of the nuclear receptor FXR in adaptive thermogenesis was investigated using FXR-deficient mice. Despite elevated serum bile acid concentrations and increased mRNA expression profiles of thermogenic genes in brown adipose tissue, FXR-deficiency did not alter energy expenditure under basal conditions. However, FXR-deficiency accelerated the fasting-induced entry into torpor in a leptin-dependent manner. FXR-deficient mice were also extremely cold-intolerant. These altered responses may be linked to a more rapid decrease in plasma concentrations of metabolic fuels (glucose, triglycerides) thus impairing uncoupling protein 1-driven thermogenesis. These results identify FXR as a modulator of energy homeostasis.

The farnesoid X receptor (FXR) has been suggested to play a role in gluconeogenesis. To determine whether FXR modulates the response to fasting in vivo, FXR-deficient (FXR-/-) and wild-type mice were submitted to fasting for 48 h. Our results demonstrate that FXR modulates the kinetics of alterations of glucose homeostasis during fasting, with FXR-/- mice displaying an early, accelerated hypoglycaemia response. Basal hepatic glucose production rate was lower in FXR-/- mice, together with a decrease in hepatic glycogen content. Moreover, hepatic PEPCK gene expression was transiently lower in FXR-/- mice after 6h of fasting and was decreased in FXR-/- hepatocytes. FXR therefore plays an unexpected role in the control of fuel availability upon fasting.

Optimal treatment of non operable localized non small cell lung carcinoma (NSCLC) continues to evolve. Increasing overall survival must evolute through improving local tumoral control and eradication of probable occult metastasis. Historically, median survival varies between 7 and 10 months with a standard conventional fractionated radiotherapy (RT). Induction chemotherapy (CT) followed by RT has demonstrated its superiority over RT alone, modality which is widely utilised. Other studies revealed best results with decreasing metastatic relapses. Three independent meta-analysis confirmed benefit obtained with cisplatin based CT followed by RT that allowed to consider this association as a gold standard. Other authors demonstrated an improvement of local control and survival with concomitant RT-CT or hyperfractionated accelerated RT. Results of all of these new therapeutic modalities still poor. Implication of new CT drugs has conducted for an emergence of new studies finding to demonstrate more encouraging results. Randomized trials are conducted in this way.