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Leukotrienes (LTs) are potent proinflammatory mediators, and many important aspects of innate and adaptive immune responses are regulated by LTs. Key members of the LT synthesis pathway are overexpressed in adipose tissue (AT) during obesity, resulting in increased LT levels in this tissue. We observed that several mouse adipocyte cell lines and primary adipocytes from mice and humans both can secrete large amounts of LTs. Furthermore, this production increases with a high-fat diet (HFD) and positively correlates with adipocyte size. LTs produced by adipocytes play an important role in attracting macrophages and T cells in in vitro chemotaxis assays. Mice that are deficient for the enzyme 5-lipoxygenase (5-LO), and therefore lack LTs, exhibit a decrease in HFD-induced AT macrophage and T-cell infiltration and are partially protected from HFD-induced insulin resistance. Similarly, treatment of HFD-fed wild-type mice with the 5-LO inhibitor Zileuton also results in a reduction of AT macrophages and T cells, accompanied by a decrease in insulin resistance. Together, these findings suggest that LTs represent a novel target in the prevention or treatment of obesity-associated inflammation and insulin resistance. [ABSTRACT FROM AUTHOR]

Type 2 diabetes (T2D) and obesity are strongly associated with low natriuretic peptide (NP) plasma levels and a down-regulation of NP guanylyl cyclase receptor-A (GCA) in skeletal muscle and adipose tissue. However, no study has so far provided evidence for a causal link between atrial NP (ANP)/GCA deficiency and T2D pathogenesis. Here, we show that both systemic and skeletal muscle ANP/GCA deficiencies in mice promote metabolic disturbances and prediabetes. Skeletal muscle insulin resistance is further associated with altered mitochondrial function and impaired endurance running capacity. ANP/GCA-deficient mice exhibit increased proton leak and reduced content of mitochondrial oxidative phosphorylation proteins. We further show that GCA is related to several metabolic traits in T2D and positively correlates with markers of oxidative capacity in human skeletal muscle. Together, these results indicate that ANP/GCA signaling controls muscle mitochondrial integrity and oxidative capacity in vivo and plays a causal role in the development of prediabetes.

The 5/6 nephrectomy and adenine-induced nephropathy mouse models have been extensively used to study Chronic Kidney Disease (CKD)-related cachexia. One common caveat of these CKD models is the cross-sectional nature of comparisons made versus controls. We here performed a comprehensive longitudinal assessment of body composition and energy metabolism in both models. The most striking finding is that weight loss is largely driven by reduced food intake which promotes rapid loss of lean and fat mass. However, in both models, mice catch up weight and lean mass a few days after the surgery or when they are switched back to standard chow diet. Muscle force and mass are fully recovered and no sign of cachexia is observed. Our data demonstrate that the time-course of kidney failure and weight loss are unrelated in these common CKD models. These data highlight the need to reconsider the relative contribution of direct and indirect mechanisms to muscle wasting observed in CKD., (© 2024. The Author(s).)

Skeletal muscle plays a central role in the regulation of systemic metabolism during lifespan. With aging, this function is perturbed, initiating multiple chronic diseases. Our knowledge of mechanisms responsible for this decline is limited. Glycerophosphocholine phosphodiesterase 1 (Gpcpd1) is a highly abundant muscle enzyme that hydrolyzes glycerophosphocholine (GPC). The physiological functions of Gpcpd1 remain largely unknown. Here we show, in mice, that the Gpcpd1-GPC metabolic pathway is perturbed in aged muscles. Further, muscle-specific, but not liver- or fat-specific, inactivation of Gpcpd1 resulted in severely impaired glucose metabolism. Western-type diets markedly worsened this condition. Mechanistically, Gpcpd1 muscle deficiency resulted in accumulation of GPC, causing an 'aged-like' transcriptomic signature and impaired insulin signaling in young Gpcpd1-deficient muscles. Finally, we report that the muscle GPC levels are markedly altered in both aged humans and patients with type 2 diabetes, displaying a high positive correlation between GPC levels and chronological age. Our findings reveal that the muscle GPCPD1-GPC metabolic pathway has an important role in the regulation of glucose homeostasis and that it is impaired during aging, which may contribute to glucose intolerance in aging., (© 2024. The Author(s), under exclusive licence to Springer Nature America, Inc.)

Fibro-adipogenic progenitors (FAPs) play a crucial role in skeletal muscle regeneration, as they generate a favorable niche that allows satellite cells to perform efficient muscle regeneration. After muscle injury, FAP content increases rapidly within the injured muscle, the origin of which has been attributed to their proliferation within the muscle itself. However, recent single-cell RNAseq approaches have revealed phenotype and functional heterogeneity in FAPs, raising the question of how this differentiation of regenerative subtypes occurs. Here we report that FAP-like cells residing in subcutaneous adipose tissue (ScAT), the adipose stromal cells (ASCs), are rapidly released from ScAT in response to muscle injury. Additionally, we find that released ASCs infiltrate the damaged muscle, via a platelet-dependent mechanism and thus contribute to the FAP heterogeneity. Moreover, we show that either blocking ASCs infiltration or removing ASCs tissue source impair muscle regeneration. Collectively, our data reveal that ScAT is an unsuspected physiological reservoir of regenerative cells that support skeletal muscle regeneration, underlining a beneficial relationship between muscle and fat., (© 2023. The Author(s).)

The number of older obese adults is increasing worldwide. Whether obese adults show similar health benefits in response to lifestyle interventions at different ages is unknown. The study enrolled 25 obese men (body mass index: 31-39 kg/m 2 ) in two arms according to age (30-40 and 60-70 yr old). Participants underwent an 8-wk intervention with moderate calorie restriction (∼20% below individual energy requirements) and supervised endurance training resulting in ∼5% weight loss. Body composition was measured using dual energy X-ray absorptiometry. Insulin sensitivity was assessed during a hypersinsulinemic-euglycemic clamp. Cardiometabolic profile was derived from blood parameters. Subcutaneous fat and vastus lateralis muscle biopsies were used for ex vivo analyses. Two-way repeated-measure ANOVA and linear mixed models were used to evaluate the response to lifestyle intervention and comparison between the two groups. Fat mass was decreased and bone mass was preserved in the two groups after intervention. Muscle mass decreased significantly in older obese men. Cardiovascular risk (Framingham risk score, plasma triglyceride, and cholesterol) and insulin sensitivity were greatly improved to a similar extent in the two age groups after intervention. Changes in adipose tissue and skeletal muscle transcriptomes were marginal. Analysis of the differential response to the lifestyle intervention showed tenuous differences between age groups. These data suggest that lifestyle intervention combining calorie restriction and exercise shows similar beneficial effects on cardiometabolic risk and insulin sensitivity in younger and older obese men. However, attention must be paid to potential loss of muscle mass in response to weight loss in older obese men. NEW & NOTEWORTHY Rise in obesity and aging worldwide are major trends of critical importance in public health. This study addresses a current challenge in obesity management. Do older obese adults respond differently to a lifestyle intervention composed of moderate calorie restriction and supervised physical activity than younger ones? The main conclusion of the study is that older and younger obese men similarly benefit from the intervention in terms of cardiometabolic risk.

Atrial natriuretic peptide (ANP) is a cardiac hormone controlling blood volume and pressure in mammals. It is still unclear whether ANP controls cold-induced thermogenesis in vivo. Here, we show that acute cold exposure induces cardiac ANP secretion in mice and humans. Genetic inactivation of ANP promotes cold intolerance and suppresses half of cold-induced brown adipose tissue (BAT) activation in mice. While white adipocytes are resistant to ANP-mediated lipolysis at thermoneutral temperature in mice, cold exposure renders white adipocytes fully responsive to ANP to activate lipolysis and a thermogenic program, a physiological response that is dramatically suppressed in ANP null mice. ANP deficiency also blunts liver triglycerides and glycogen metabolism, thus impairing fuel availability for BAT thermogenesis. ANP directly increases mitochondrial uncoupling and thermogenic gene expression in human white and brown adipocytes. Together, these results indicate that ANP is a major physiological trigger of BAT thermogenesis upon cold exposure in mammals., Competing Interests: Declaration of Interests The authors declare no competing interests., (Copyright © 2020 The Author(s). Published by Elsevier Inc. All rights reserved.)

We hypothesized that skeletal muscle contraction produces a cellular stress signal, triggering adipose tissue lipolysis to sustain fuel availability during exercise. The present study aimed at identifying exercise-regulated myokines, also known as exerkines, able to promote lipolysis. Human primary myotubes from lean healthy volunteers were submitted to electrical pulse stimulation (EPS) to mimic either acute intense or chronic moderate exercise. Conditioned media (CM) experiments with human adipocytes were performed. CM and human plasma samples were analyzed using unbiased proteomic screening and/or ELISA. Real-time qPCR was performed in cultured myotubes and muscle biopsy samples. CM from both acute intense and chronic moderate exercise increased basal lipolysis in human adipocytes. Growth and differentiation factor 15 (GDF15) gene expression and secretion increased rapidly upon skeletal muscle contraction. GDF15 protein was upregulated in CM from both acute and chronic exercise-stimulated myotubes. We further showed that physiological concentrations of recombinant GDF15 protein increased lipolysis in human adipose tissue, while blocking GDF15 with a neutralizing antibody abrogated EPS CM-mediated lipolysis. We herein provide the first evidence to our knowledge that GDF15 is a potentially novel exerkine produced by skeletal muscle contraction and able to target human adipose tissue to promote lipolysis.

Ectopic lipid deposition (ELD) is defined by excess fat storage in locations not classically associated with adipose tissue (AT) storage. ELD is positively correlated with insulin resistance and increased risk of metabolic disorders. ELD appears as lipid droplets or adipocytes, whose cell origin is unknown. We previously showed that subcutaneous AT (ScAT) releases adipocyte progenitors into the circulation. Here, we demonstrate that triggering or preventing the release of adipocyte precursors from ScAT directly promoted or limited ectopic adipocyte formation in skeletal muscle in mice. Importantly, obesity-associated metabolic disorders could be mimicked by causing adipocyte precursor release without a high-fat diet. Finally, during nutrient overload, adipocyte progenitors exited ScAT, where their retention signals (CXCR4/CXCL12 axis) were greatly decreased, and further infiltrated skeletal muscles. These data provide insights into the formation of ELD associated with calorie overload and highlight adipocyte progenitor trafficking as a potential target in the treatment of metabolic diseases., (Copyright © 2019 The Author(s). Published by Elsevier Inc. All rights reserved.)

We describe here a collective and experimental research project-based learning (ERPBL) for master's students that can be used to illustrate some basic concepts on glucose/lipid homeostasis and renal function around a topical issue. The primary objective of this ERPBL was to strengthen students' knowledge and understanding of physiology and pathophysiology. The secondary objectives were to help students to develop technical/practical abilities and acquire transversal skills with real-world connections. Obesity is a worldwide public health problem that increases the risk for developing type 2 diabetes and nephropathies. To study the impact of western dietary habits, students evaluated the effects of a diet enriched with fat and cola [high-fat and cola diet (HFCD)] on metabolism and renal function in mice. Students mainly worked in tandem to prepare and perform experiments, but also collectively to compile, analyze, and discuss data. Students showed that HFCD-fed mice 1 ) developed obesity; 2 ) exhibited glucose homeostasis impairments associated to ectopic fat storage; and 3 ) displayed reduced glomerular filtration. The educational benefit of the program was estimated using three evaluation metrics: a conventional multicriteria assessment by teachers, a pre-/posttest, and a self-evaluation questionnaire. They showed that the current approach successfully strengthened scientific student knowledge and understanding of physiology/pathophysiology. In addition, it helped students develop new skills, such as technical and transversal skills. We concluded that this ERPBL dealing with the pathophysiology of obesity was strongly beneficial for master's students, thereby appearing as an efficient and performing educational tool., (Copyright © 2017 the American Physiological Society.)

Lipid droplets (LD) play a central role in lipid homeostasis by controlling transient fatty acid (FA) storage and release from triacylglycerols stores, while preventing high levels of cellular toxic lipids. This crucial function in oxidative tissues is altered in obesity and type 2 diabetes. Perilipin 5 (PLIN5) is a LD protein whose mechanistic and causal link with lipotoxicity and insulin resistance has raised controversies. We investigated here the physiological role of PLIN5 in skeletal muscle upon various metabolic challenges. We show that PLIN5 protein is elevated in endurance-trained (ET) subjects and correlates with muscle oxidative capacity and whole-body insulin sensitivity. When overexpressed in human skeletal muscle cells to recapitulate the ET phenotype, PLIN5 diminishes lipolysis and FA oxidation under basal condition, but paradoxically enhances FA oxidation during forskolin- and contraction- mediated lipolysis. Moreover, PLIN5 partly protects muscle cells against lipid-induced lipotoxicity. In addition, we demonstrate that down-regulation of PLIN5 in skeletal muscle inhibits insulin-mediated glucose uptake under normal chow feeding condition, while paradoxically improving insulin sensitivity upon high-fat feeding. These data highlight a key role of PLIN5 in LD function, first by finely adjusting LD FA supply to mitochondrial oxidation, and second acting as a protective factor against lipotoxicity in skeletal muscle.

Objective: Recent data suggest that adipose triglyceride lipase (ATGL) plays a key role in providing energy substrate from triglyceride pools and that alterations of its expression/activity relate to metabolic disturbances in skeletal muscle. Yet little is known about its regulation. We here investigated the role of the protein G0/G1 Switch Gene 2 (G0S2), recently described as an inhibitor of ATGL in white adipose tissue, in the regulation of lipolysis and oxidative metabolism in skeletal muscle., Methods: We first examined G0S2 protein expression in relation to metabolic status and muscle characteristics in humans. We next overexpressed and knocked down G0S2 in human primary myotubes to assess its impact on ATGL activity, lipid turnover and oxidative metabolism, and further knocked down G0S2 in vivo in mouse skeletal muscle., Results: G0S2 protein is increased in skeletal muscle of endurance-trained individuals and correlates with markers of oxidative capacity and lipid content. Recombinant G0S2 protein inhibits ATGL activity by about 40% in lysates of mouse and human skeletal muscle. G0S2 overexpression augments (+49%, p < 0.05) while G0S2 knockdown strongly reduces (-68%, p < 0.001) triglyceride content in human primary myotubes and mouse skeletal muscle. We further show that G0S2 controls lipolysis and fatty acid oxidation in a strictly ATGL-dependent manner. These metabolic adaptations mediated by G0S2 are paralleled by concomitant changes in glucose metabolism through the modulation of Pyruvate Dehydrogenase Kinase 4 (PDK4) expression (5.4 fold, p < 0.001). Importantly, downregulation of G0S2 in vivo in mouse skeletal muscle recapitulates changes in lipid metabolism observed in vitro., Conclusion: Collectively, these data indicate that G0S2 plays a key role in the regulation of skeletal muscle ATGL activity, lipid content and oxidative metabolism.

Circulating natriuretic peptide (NP) levels are reduced in obesity and predict the risk of type 2 diabetes (T2D). Since skeletal muscle was recently shown as a key target tissue of NP, we aimed to investigate muscle NP receptor (NPR) expression in the context of obesity and T2D. Muscle NPRA correlated positively with whole-body insulin sensitivity in humans and was strikingly downregulated in obese subjects and recovered in response to diet-induced weight loss. In addition, muscle NP clearance receptor (NPRC) increased in individuals with impaired glucose tolerance and T2D. Similar results were found in obese diabetic mice. Although no acute effect of brain NP (BNP) on insulin sensitivity was observed in lean mice, chronic BNP infusion improved blood glucose control and insulin sensitivity in skeletal muscle of obese and diabetic mice. This occurred in parallel with a reduced lipotoxic pressure in skeletal muscle due to an upregulation of lipid oxidative capacity. In addition, chronic NP treatment in human primary myotubes increased lipid oxidation in a PGC1α-dependent manner and reduced palmitate-induced lipotoxicity. Collectively, our data show that activation of NPRA signaling in skeletal muscle is important for the maintenance of long-term insulin sensitivity and has the potential to treat obesity-related metabolic disorders., (© 2015 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.)

The aim of the present study was to investigate the influence of substrate availability on fuel selection during exercise. Eight endurance-trained male cyclists performed 90-min exercise at 70% of their maximal oxygen uptake in a cross-over design, either in rested condition (CON) or the day after 2-h exercise practised at 70% of maximal oxygen uptake (EX). Subjects were given a sucrose load (0.75 g kg(-1) body weight) 45 min after the beginning of the 90-min exercise test. Lipolysis was measured in subcutaneous abdominal adipose tissue (SCAT) by microdialysis and substrate oxidation by indirect calorimetry. Lipid oxidation increased during exercise and tended to decrease during sucrose ingestion in both conditions. Lipid oxidation was higher during the whole experimental period in the EX group (p = 0.004). Interestingly, fuel selection, assessed by the change in respiratory exchange ratio (RER), was increased in the EX session (p = 0.002). This was paralleled by a higher rate of SCAT lipolysis reflected by dialysate glycerol, plasma glycerol, and fatty acids (FA) levels (p < 0.001). Of note, we observed a significant relationship between whole-body fat oxidation and dialysate glycerol in both sessions (r (2) = 0.33, p = 0.02). In conclusion, this study highlights the limiting role of lipolysis and plasma FA availability to whole-body fat oxidation during exercise in endurance-trained subjects. This study shows that adipose tissue lipolysis is a determinant of fuel selection during exercise in healthy subjects.

Adipose tissue fibrosis development blocks adipocyte hypertrophy and favors ectopic lipid accumulation. Here, we show that adipose tissue fibrosis is associated with obesity and insulin resistance in humans and mice. Kinetic studies in C3H mice fed a high-fat diet show activation of macrophages and progression of fibrosis along with adipocyte metabolic dysfunction and death. Adipose tissue fibrosis is attenuated by macrophage depletion. Impairment of Toll-like receptor 4 signaling protects mice from obesity-induced fibrosis. The presence of a functional Toll-like receptor 4 on adipose tissue hematopoietic cells is necessary for the initiation of adipose tissue fibrosis. Continuous low-dose infusion of the Toll-like receptor 4 ligand, lipopolysaccharide, promotes adipose tissue fibrosis. Ex vivo, lipopolysaccharide-mediated induction of fibrosis is prevented by antibodies against the profibrotic factor TGFβ1. Together, these results indicate that obesity and endotoxemia favor the development of adipose tissue fibrosis, a condition associated with insulin resistance, through immune cell Toll-like receptor 4., (Copyright © 2014 The Authors. Published by Elsevier Inc. All rights reserved.)

Context: Skeletal muscle lipase and intramyocellular triglyceride (IMTG) play a role in obesity-related metabolic disorders., Objectives: The aim of the present study was to investigate the impact of 8 weeks of endurance exercise training on IMTG content and lipolytic proteins in obese male subjects., Design and Volunteers: Ten obese subjects completed an 8-week supervised endurance exercise training intervention in which vastus lateralis muscle biopsy samples were collected before and after training., Main Outcome Measures: Clinical characteristics and ex vivo substrate oxidation rates were measured pre- and posttraining. Skeletal muscle lipid content and lipolytic protein expression were also investigated., Results: Our data show that exercise training reduced IMTG content by 42% (P < .01) and increased skeletal muscle oxidative capacity, whereas no change in total diacylglycerol content and glucose oxidation was found. Exercise training up-regulated adipose triglyceride lipase, perilipin (PLIN) 3 protein, and PLIN5 protein contents in skeletal muscle despite no change in mRNA levels. Training also increased hormone sensitive-lipase Ser660 phosphorylation. No significant changes in comparative gene identification 58, G₀/G₁ switch gene 2, and PLIN2 protein and mRNA levels were observed in response to training. Interestingly, we noted a strong relationship between skeletal muscle comparative gene identification 58 and mitochondrial respiratory chain complex I protein contents at baseline (r = 0.87, P < .0001)., Conclusions: Endurance exercise training coordinately up-regulates fat oxidative capacity and lipolytic protein expression in skeletal muscle of obese subjects. This physiological adaptation probably favors fat oxidation and may alleviate the lipotoxic lipid pressure in skeletal muscle. Enhancement of IMTG turnover may be required for the beneficial metabolic effects of exercise in obesity.

Context: It was suggested that human cultured primary myotubes retain the metabolic characteristics of their donor in vitro., Objectives: The aim of the present study was to investigate whether the metabolic responses to endurance training are also conserved in culture., Design and Volunteers: Middle-aged obese subjects completed an 8-week supervised aerobic exercise training program in which vastus lateralis muscle biopsies were collected before and after training., Main Outcome Measures: Anthropometric and blood parameters, as well as aerobic capacity, were assessed before and after training. Muscle biopsies were either used for Western blot analysis or digested to harvest myogenic progenitors that were differentiated into myotubes. Glucose oxidation, palmitate oxidation, and glycogen synthesis assays were performed on myotubes before and after training. Gene expression was assessed by real-time quantitative PCR., Results: Our data indicate that in parallel of in vivo improvement of whole-body aerobic capacity and glucose metabolism, biopsy-derived primary myotubes showed similar patterns in vitro. Indeed, glucose oxidation, glycogen synthesis, and inhibition of palmitate oxidation by glucose were enhanced in myotubes after training. This was associated with consistent changes in the expression of metabolism-linked genes such as GLUT1, PDK4, and PDHA1. Interestingly, no difference in myogenic differentiation capacity was observed before and after training., Conclusion: Aerobic exercise training is associated with metabolic adaptations in vivo that are preserved in human cultured primary myotubes. It can be hypothesized that skeletal muscle microenvironmental changes induced by endurance training lead to metabolic imprinting on myogenic progenitor cells.

Elevated expression/activity of adipose triglyceride lipase (ATGL) and/or reduced activity of hormone-sensitive lipase (HSL) in skeletal muscle are causally linked to insulin resistance in vitro. We investigated here the effect of high-fat feeding on skeletal muscle lipolytic proteins, lipotoxicity, and insulin signaling in vivo. Five-week-old C3H mice were fed normal chow diet (NCD) or 45% kcal high-fat diet (HFD) for 4 weeks. Wild-type and HSL knockout mice fed NCD were also studied. Whole-body and muscle insulin sensitivity, as well as lipolytic protein expression, lipid levels, and insulin signaling in skeletal muscle, were measured. HFD induced whole-body insulin resistance and glucose intolerance and reduced skeletal muscle glucose uptake compared with NCD. HFD increased skeletal muscle total diacylglycerol (DAG) content, protein kinase Cθ and protein kinase Cε membrane translocation, and impaired insulin signaling as reflected by a robust increase of basal Ser1101 insulin receptor substrate 1 phosphorylation (2.8-fold, P < .05) and a decrease of insulin-stimulated v-Akt murine thymoma viral oncogene homolog Ser473 (-37%, P < .05) and AS160 Thr642 (-47%, P <.01) phosphorylation. We next showed that HFD strongly reduced HSL phosphorylation at Ser660. HFD significantly up-regulated the muscle protein content of the ATGL coactivator comparative gene identification 58 and triacylglycerol hydrolase activity, despite a lower ATGL protein content. We further show a defective skeletal muscle insulin signaling and DAG accumulation in HSL knockout compared with wild-type mice. Together, these data suggest a pathophysiological link between altered skeletal muscle lipase expression and DAG-mediated insulin resistance in mice.

Cardiac natriuretic peptides (NP) are major activators of human fat cell lipolysis and have recently been shown to control brown fat thermogenesis. Here, we investigated the physiological role of NP on the oxidative metabolism of human skeletal muscle. NP receptor type A (NPRA) gene expression was positively correlated to mRNA levels of PPARγ coactivator-1α (PGC1A) and several oxidative phosphorylation (OXPHOS) genes in human skeletal muscle. Further, the expression of NPRA, PGC1A, and OXPHOS genes was coordinately upregulated in response to aerobic exercise training in human skeletal muscle. In human myotubes, NP induced PGC-1α and mitochondrial OXPHOS gene expression in a cyclic GMP-dependent manner. NP treatment increased OXPHOS protein expression, fat oxidation, and maximal respiration independent of substantial changes in mitochondrial proliferation and mass. Treatment of myotubes with NP recapitulated the effect of exercise training on muscle fat oxidative capacity in vivo. Collectively, these data show that activation of NP signaling in human skeletal muscle enhances mitochondrial oxidative metabolism and fat oxidation. We propose that NP could contribute to exercise training-induced improvement in skeletal muscle fat oxidative capacity in humans.

T-cell regulation in adipose tissue provides a link between inflammation and insulin resistance. Because of alterations in adipose tissue T-cell composition in obesity, we aimed to identify the antigen-presenting cells in adipose tissue of obese mice and patients with insulin resistance. Dendritic cells (DCs) and T cells were studied in mice and in two cohorts of obese patients. In lean mice, only CD11c(+) DCs were detected in adipose tissue. Adoptive transfer of naive CD4(+) T cells in Rag1(-/-) mice led to a predominant Th1 response in adipose tissue. In contrast, during obesity DCs (human CD11c(+)CD1c(+) and mouse CD11c(high)F4/80(low)) accumulated in adipose tissue. CD11c(high)F4/80(low) DCs from obese mice induced Th17 differentiation. In patients, the presence of CD11c(+)CD1c(+) DCs correlated with the BMI and with an elevation in Th17 cells. In addition, these DCs led to ex vivo Th17 differentiation. CD1c gene expression further correlated with homeostatic model assessment-insulin resistance in the subcutaneous adipose tissue of obese patients. We show for the first time the presence and accumulation of specific DCs in adipose tissue in mouse and human obesity. These DCs were functional and could be important regulators of adipose tissue inflammation by regulating the switch toward Th17 cell responses in obesity-associated insulin resistance.

We investigated here the specific role of CGI-58 in the regulation of energy metabolism in skeletal muscle. We first examined CGI-58 protein expression in various muscle types in mice, and next modulated CGI-58 expression during overexpression and knockdown studies in human primary myotubes and evaluated the consequences on oxidative metabolism. We observed a preferential expression of CGI-58 in oxidative muscles in mice consistent with triacylglycerol hydrolase activity. We next showed by pulse-chase that CGI-58 overexpression increased by more than 2-fold the rate of triacylglycerol (TAG) hydrolysis, as well as TAG-derived fatty acid (FA) release and oxidation. Oppositely, CGI-58 silencing reduced TAG hydrolysis and TAG-derived FA release and oxidation (-77%, P < 0.001), whereas it increased glucose oxidation and glycogen synthesis. Interestingly, modulations of CGI-58 expression and FA release are reflected by changes in pyruvate dehydrogenase kinase 4 gene expression. This regulation involves the activation of the peroxisome proliferator activating receptor-δ (PPARδ) by lipolysis products. Altogether, these data reveal that CGI-58 plays a limiting role in the control of oxidative metabolism by modulating FA availability and the expression of PPARδ-target genes, and highlight an important metabolic function of CGI-58 in skeletal muscle.

Objective: The present study was undertaken to characterize the remodeling phenotype of human adipose tissue (AT) macrophages (ATM) and to analyze their paracrine effects on AT progenitor cells., Research Design and Methods: The phenotype of ATM, immunoselected from subcutaneous (Sc) AT originating from subjects with wide range of body mass index and from paired biopsies of Sc and omental (Om) AT from obese subjects, was studied by gene expression analysis in the native and activated states. The paracrine effects of ScATM on the phenotype of human ScAT progenitor cells (CD34(+)CD31(-)) were investigated., Results: Two main ATM phenotypes were distinguished based on gene expression profiles. For ScAT-derived ATM, obesity and adipocyte-derived factors favored a pro-fibrotic/remodeling phenotype whereas the OmAT location and hypoxic culture conditions favored a pro-angiogenic phenotype. Treatment of native human ScAT progenitor cells with ScATM-conditioned media induced the appearance of myofibroblast-like cells as shown by expression of both α-SMA and the transcription factor SNAIL, an effect mimicked by TGFβ1 and activinA. Immunohistochemical analyses showed the presence of double positive α-SMA and CD34 cells in the stroma of human ScAT. Moreover, the mRNA levels of SNAIL and SLUG in ScAT progenitor cells were higher in obese compared with lean subjects., Conclusions: Human ATM exhibit distinct pro-angiogenic and matrix remodeling/fibrotic phenotypes according to the adiposity and the location of AT, that may be related to AT microenvironment including hypoxia and adipokines. Moreover, human ScAT progenitor cells have been identified as target cells for ScATM-derived TGFβ and as a potential source of fibrosis through their induction of myofibroblast-like cells.

Objective: Growth of white adipose tissue takes place in normal development and in obesity. A pool of adipose progenitors is responsible for the formation of new adipocytes and for the potential of this tissue to expand in response to chronic energy overload. However, factors controlling self-renewal of human adipose progenitors are largely unknown. We investigated the expression profile and the role of activin A in this process., Research Design and Methods: Expression of INHBA/activin A was investigated in three types of human adipose progenitors. We then analyzed at the molecular level the function of activin A during human adipogenesis. We finally investigated the status of activin A in adipose tissues of lean and obese subjects and analyzed macrophage-induced regulation of its expression., Results: INHBA/activin A is expressed by adipose progenitors from various fat depots, and its expression dramatically decreases as progenitors differentiate into adipocytes. Activin A regulates the number of undifferentiated progenitors. Sustained activation or inhibition of the activin A pathway impairs or promotes, respectively, adipocyte differentiation via the C/EBPβ-LAP and Smad2 pathway in an autocrine/paracrine manner. Activin A is expressed at higher levels in adipose tissue of obese patients compared with the expression levels in lean subjects. Indeed, activin A levels in adipose progenitors are dramatically increased by factors secreted by macrophages derived from obese adipose tissue., Conclusions: Altogether, our data show that activin A plays a significant role in human adipogenesis. We propose a model in which macrophages that are located in adipose tissue regulate adipose progenitor self-renewal through activin A.

Objective: Adipose tissue (AT) plays a major role in the low-grade inflammatory state associated with obesity. The aim of the present study was to characterize the human AT lymphocytes (ATLs) and to analyze their interactions with adipocytes., Methods and Results: Human ATL subsets were characterized by flow cytometry in subcutaneous ATs from 92 individuals with body mass index (BMI) ranging from 19 to 43 kg/m(2) and in paired biopsies of subcutaneous and visceral AT from 45 class II/III obese patients. CD3(+) ATLs were composed of effector and memory CD4(+) helper and CD8(+) cytotoxic T cells. The number of ATLs correlated positively with BMI and was higher in visceral than subcutaneous AT. Mature adipocytes stimulated the migration of ATLs and released the chemokine CCL20, the receptor of which (CCR6) was expressed in ATLs. The expression of adipocyte CCL20 was positively correlated with BMI and increased in visceral compared to subcutaneous adipocytes. ATLs expressed inflammatory markers and released interferon gamma (IFN gamma). Progenitor and adipocyte treatment with ATL-conditioned media reduced the insulin-mediated upregulation of lipogenic enzymes, an effect involving IFN gamma., Conclusions: Therefore, crosstalk occurs between adipocytes and lymphocytes within human AT involving T cell chemoattraction by adipocytes and modulation of lipogenesis by ATLs.

Context: Adipocyte formation in human adult adipose tissue (hAT) originates from resident progenitor cell differentiation in the stroma vascular fraction of the AT. The processes involved in the self-renewal of this cell population remain to be defined., Objective: The objective was to study in situ and in vitro hAT progenitor cell (defined as CD34(+)/CD31(-) cells) proliferation., Design and Participants: In situ progenitor cell proliferation was assessed by immunohistochemistry and flow cytometry analyses on hAT from lean to obese subjects using the proliferation marker Ki-67. The effects of adipokines, hypoxia, and conditioned media (CM) from adipocytes, capillary endothelial cells, and macrophages isolated by an immunoselection approach were studied on hAT progenitor cell growth. Cell death in hAT was assessed by the terminal deoxynucleotidyl transferase-mediated dUTP-fluorescein end labeling method., Results: Ki-67-positive staining was observed in AT progenitor cells. Fat mass enlargement in obese patients was associated with an increased Ki-67(+) progenitor cell population together with a new fraction of small adipocytes and increased cell death. HIF-1alpha mRNA expression in freshly harvested progenitor cells was positively correlated with body mass index. Adipocyte- and capillary endothelial cell-CM, hypoxia, leptin, IL-6, lysophosphatidic acid, and vascular endothelial growth factor, all increased hAT progenitor cell proliferation in vitro. Macrophage-CM had an antiproliferative effect that was suppressed by an antioxidant., Conclusions: The fraction of proliferative progenitor cells in adult hAT is modulated by the degree of adiposity. Changes in the progenitor cell microenvironment involving adipokines, hypoxia, and oxidative stress might play a key role in the control of the self-renewal of the local pool of AT progenitor cells.

In a previous publication, we reported that human immunodeficiency virus (HIV) protease inhibitors (PIs) inhibited the differentiation of human preadipocytes in primary culture, reducing the expression and secretion of matrix metalloproteinase 9 (MMP-9). The present work was performed to clarify this mechanism. Interestingly, HIV-PIs have been reported to be inhibitors of the proteasome complex, which is known to regulate nuclear factor (NF)-kappaB activation and transcription of its target genes, among them MMP-9. We thus investigated the potential involvement of the proteasome in the antiadipogenic effects of HIV-PIs. The effect of four HIV-PIs was tested on preadipocyte proteasomal activity, and chronic treatment with the specific proteasome inhibitor lactacystin was performed to evaluate alterations of adipogenesis and MMP-9 expression/secretion. Finally, modifications of the NF-kappaB pathway induced by either HIV-PIs or lactacystin were studied. We demonstrated that preadipocyte proteasomal activity was decreased by several HIV-PIs and that chronic treatment with lactacystin mimicked the effects of HIV-PIs by reducing adipogenesis and MMP-9 expression/secretion. Furthermore, we observed an intracellular accumulation of the NF-kappaB inhibitor, IkappaBbeta, with chronic treatment with HIV-PIs or lactacystin as well as a decrease in MMP-9 expression induced by acute tumor necrosis factor-alpha stimulation. These results indicate that inhibition of the proteasome by specific (lactacystin) or nonspecific (HIV-PIs) inhibitors leads to a reduction of human adipogenesis, and they therefore implicate deregulation of the NF-kappaB pathway and the related decrease of the key adipogenic factor, MMP-9. This study adds significantly to recent reports that have linked HIV-PI-related lipodystrophic syndrome with altered proteasome function, endoplasmic reticulum stress, and metabolic disorders.

We previously showed that human and murine 3T3-F442A preadipocytes produced and released matrix metalloproteinases (MMPs) 2 and 9 and that a treatment by MMP inhibitors resulted in the blockade of murine fat cell adipose conversion. In parallel, investigators reported that other protease inhibitors, the human immunodeficiency virus (HIV) protease inhibitors (PIs) involved in lipodystrophy in humans, also reduced the adipocyte differentiation process of several murine cell lines. The present work was performed to define the effects of MMP inhibitors and HIV-PIs on the human adipocyte differentiation process, to clarify the involvement of MMPs in the control of human adipogenesis, and to determine whether HIV-PIs interact with MMPs in the control of this process. The effect of two MMP inhibitor and four HIV-PI treatments on the differentiation of primary culture human preadipocytes, as well as the putative relationships between HIV-PIs and MMP-2 and -9 expression, release, or activity were investigated. We showed that MMP inhibitors and HIV-PIs reduced the human adipocyte differentiation process as assessed by the decrease of cell protein and/or triglyceride contents and expression of fatty acid binding protein and hormone-sensitive lipase, two adipocyte markers. Unlike MMP inhibitors, HIV-PIs were devoid of any effect per se on recombinant MMP-2 and 9 activities but reduced the expression and release of MMP-9 by human preadipocytes. Thus, the present study indicates that the modulation of the extracellular matrix components through the production and/or activity of MMPs, and, more precisely, MMP-9 might be a key factor in the regulation of human adipose tissue development.

Bourlier, V., Diserbo, M., Gourmelon, P. and Verdetti, J. Prolonged Effects of Acute Gamma Irradiation on Acetylcholine-Induced Potassium Currents in Human Umbilical Vein Endothelial Cells. Radiat. Res. 155, 748-752 (2001). We have recently reported an acute effect of gamma irradiation (15 Gy, 1 Gy/min) on acetylcholine-mediated endothelium-dependent relaxation in rat aortic rings. Given the importance of permeability to K+ to endothelium-dependent relaxation, we have evaluated the effect of the same radiation on K+ currents in human endothelial cells in culture using the patch-clamp technique in the whole-cell recording configuration. Our results indicate that, in resting cells, gamma irradiation has no effect on endothelial permeability to K+. However, irradiation during stimulation of endothelial cells with acetylcholine reduces the sustained increase in permeability to K+ observed in the acetylcholine-stimulated, nonirradiated cells. Additional experiments using K+ channel inhibitors (TEA, charybdotoxin, apamin) suggest that irradiation may in part decrease the prolonged activation of Ca2+-activated K+ channels by acetylcholine. Taken together with our previous finding that irradiation inhibits the acute relaxing effects of acetylcholine, these results show that gamma irradiation also affects the delayed effects of acetylcholine on permeability to K+.

Septic shock is characterized by vasodilation and decreased responsiveness to vasoconstrictors. Recent studies suggest this results from nitric oxide (NO) overproduction after expression of the calcium-independent isoform of NO synthase (iNOS) in smooth muscle cells. However, direct evidence linking iNOS (NOS2) expression and decreased microvascular responsiveness after septic stimuli is lacking. In the present study, we determined the effect of bacterial lipopolysaccharide (LPS, 20 mg/kg, IP) on smooth muscle contraction and endothelial relaxation in mesenteric resistance arteries from wild-type and iNOS knockout mice. Four hours after challenge with LPS or saline in vivo, concentration-dependent responses to norepinephrine (NE) and acetylcholine (NE+ACh) were measured in cannulated, pressurized vessels ex vivo. In vessels from wild-type mice, NE-induced contraction was markedly impaired after LPS, and pretreatment with the iNOS inhibitor aminoguanidine (AG) partly restored the NE contraction. In contrast, NE contraction in microvessels from iNOS knockout mice was unaffected by LPS. ACh-induced relaxation was unaffected by LPS in vessels from either genotype. These data provide direct evidence that iNOS gene expression mediates the LPS-induced decrease in microvascular responsiveness to vasoconstrictors. Moreover, the observation that AG did not fully restore NE contraction after LPS, whereas iNOS gene deficiency did, suggests that iNOS expression plays a central role in the development of the NO-independent effect of LPS on microvascular responsiveness. Finally, our data indicate that LPS or iNOS expression has little effect on endothelium-dependent relaxation, and eNOS activity does not appear to play a role in the decreased smooth muscle responsiveness after LPS in this model. The full text of this article is available at http://www.circresaha.org.

Background: Noninvasive techniques used to determine the changes in cerebral blood volume in response to carbon dioxide are hampered by their limited spatial or temporal resolution or both. Using steady state contrast-enhanced magnetic resonance imaging, the authors determined regional changes in cerebral plasma volume (CPV) induced by hypercapnia in halothane-anesthetized rats., Methods: Cerebral plasma volume was determined during normocapnia, hypercapnia and recovery in the dorsoparietal neocortex and striatum of each hemisphere, in cerebellum, and in extracerebral tissue of rats with either intact carotid arteries (group 1) or unilateral common carotid ligation (group 2). Another group was studied without injection of a contrast agent (group 3)., Results: Hypercapnia (partial pressure of carbon dioxide in arterial blood [PaCO2] approximately 65 mmHg) resulted in a significant increase in CPV in the striatum (+42 +/- 8%), neocortex (+34 +/- 6%), and cerebellum (+49 +/- 12%) compared with normocapnic CPV values (group 1). Carotid ligation (group 2) led to a marked reduction of the CPV response to hypercapnia in the ipsilateral striatum (+23 +/- 14%) and neocortex (+27 +/- 17%) compared with the unclamped side (+34 +/- 15% and +38 +/- 16%, respectively). No significant changes in CPV were found in extracerebral tissue. In both groups, the CPV changes were reversed by the carbon dioxide washout period. Negligible changes in contrast imaging were detected during hypercapnia without administration of the contrast agent (group 3)., Conclusions: The contrast-enhanced magnetic resonance imaging technique is sensitive to detect noninvasively regional CPV changes induced by hypercapnia in rat brain. This could be of clinical interest for determining the cerebrovascular reactivity among different brain regions.

1. To determine the acute effects of irradiation on the functionality of vessel, rat aortic rings were mounted in an organ bath for isometric tension measurements and irradiated (60Co, 1 Gy min(-1), 15 min). 2. Irradiation, which is without effect on non-contracted or endothelium-denuded vessels, led to an immediate and reversible increase in vascular tone on (-)-phenylephrine (1 microM)-precontracted aortic rings. The tension reached a plateau about 5 min after the beginning of irradiation. 3. The maximal radiation-induced contraction occurred on aortic rings relaxed by acetylcholine (ACh) (1 microM). In this condition, the addition of catalase (1000 u ml(-1)), which reduces hydrogen peroxide, and DMSO (0.1% v/v), which scavenges hydroxyl radical, had no influence on tension level while superoxide dismutase (SOD) (100 u ml(-1)), a superoxide anion scavenger, reduced the observed contraction. A similar result was obtained in the presence of indomethacin (10 microM), a cyclo-oxygenase blocker. 4. Pretreatment of rings with the nitric oxide synthase inhibitor, N(omega)-nitro-L-arginine methyl ester (L-NAME) (10-100 microM) inhibited the radiation-induced contraction. 5. This effect was dose rate-dependent and even occurred for a very low dose rate (0.06 Gy min(-1)). 6. The present results indicate that gamma-radiation induces an instantaneous vascular tone increase that is endothelium and dose rate-dependent. This effect is (i) maximal when nitric oxide (NO) is produced, (ii) greatly reduced by SOD and (iii) inhibited by L-NAME, suggesting a major involvement of complexes between NO and superoxide anion.

The protective effect of heat stress against mechanical dysfunction and myocardial necrosis after prolonged ischemia is well known. We have investigated whether the protective effect of heat stress extends to reperfusion arrhythmias in the isolated perfused rat heart. Rats were exposed to 20 min of 42 degrees C hyperthermia. Twenty-four h later their hearts were isolated, perfused and subjected to a 5-min period of occlusion of the left coronary artery. The incidence and duration of reperfusion arrhythmias were assessed in the 30-min reperfusion period. Prior heat stress led to a reduction in the incidence (from 100 to 60%, P