Transition from metabolic adaptation to maladaptation of the heart in obesity: role of apelin.

BACKGROUND/OBJECTIVES: Impaired energy metabolism is the defining characteristic of obesity-related heart failure. The adipocyte-derived peptide apelin has a role in the regulation of cardiovascular and metabolic homeostasis and may contribute to the link between obesity, energy metabolism and cardiac function. Here we investigate the role of apelin in the transition from metabolic adaptation to maladaptation of the heart in obese state. METHODS: Adult male C57BL/6J, apelin knock-out (KO) or wild-type mice were fed a high-fat diet (HFD) for 18 weeks. To induce heart failure, mice were subjected to pressure overload after 18 weeks of HFD. Long-term effects of apelin on fatty acid (FA) oxidation, glucose metabolism, cardiac function and mitochondrial changes were evaluated in HFD-fed mice after 4 weeks of pressure overload. Cardiomyocytes from HFD-fed mice were isolated for analysis of metabolic responses. RESULTS: In HFD-fed mice, pressure overload-induced transition from hypertrophy to heart failure is associated with reduced FA utilization (P<0.05), accelerated glucose oxidation (P<0.05) and mitochondrial damage. Treatment of HFD-fed mice with apelin for 4 weeks prevented pressure overload-induced decline in FA metabolism (P<0.05) and mitochondrial defects. Furthermore, apelin treatment lowered fasting plasma glucose (P<0.01), improved glucose tolerance (P<0.05) and preserved cardiac function (P<0.05) in HFD-fed mice subjected to pressure overload. In apelin KO HFD-fed mice, spontaneous cardiac dysfunction is associated with reduced FA oxidation (P<0.001) and increased glucose oxidation (P<0.05). In isolated cardiomyocytes, apelin stimulated FA oxidation in a dose-dependent manner and this effect was prevented by small interfering RNA sirtuin 3 knockdown. CONCLUSIONS: These data suggest that obesity-related decline in cardiac function is associated with defective myocardial energy metabolism and mitochondrial abnormalities. Furthermore, our work points for therapeutic potential of apelin to prevent myocardial metabolic abnormalities in heart failure paired with obesity.

Effect of endurance training on skeletal muscle myokine expression in obese men: identification of apelin as a novel myokine.

BACKGROUND: It has been suggested that the metabolic benefits of physical exercise could be mediated by myokines. We examined here the effect of exercise training on skeletal muscle expression of a panel of myokines in humans. Pathways regulating myokine expression were investigated in human myotubes. METHODS: Eleven obese non-diabetic male subjects were enrolled in an 8-week endurance training program. Insulin sensitivity was assessed by an oral glucose tolerance test. Subcutaneous adipose tissue and Vastus lateralis muscle biopsy samples were collected before and after training. RNAs were prepared from adipose tissue and skeletal muscle. Primary culture of myoblasts was established. RESULTS: As expected, exercise training improved aerobic capacity and decreased fat mass. No significant change in interleukin 6, fibroblast growth factor 21, myostatin (MSTN) or irisin mRNA level was found in muscle after training. A twofold increase in apelin mRNA level was found in muscle but not in adipose tissue. No change in circulating myokine and adipokine plasma levels was observed in the resting state in response to training. Interestingly, apelin was significantly expressed and secreted in primary human myotubes. Apelin gene expression was upregulated by cyclic AMP and calcium, unlike the other myokines investigated. Importantly, changes in muscle apelin mRNA levels were positively related to whole-body insulin sensitivity improvement. CONCLUSION: Collectively, our data show that exercise training upregulates muscle apelin expression in obese subjects. Apelin expression is induced by exercise signaling pathways and secreted in vitro in human primary myotubes, and may behave as a novel exercise-regulated myokine with autocrine/paracrine action.

Impairment of adipose tissue in Prader-Willi syndrome rescued by growth hormone treatment.

BACKGROUND: Prader-Willi syndrome (PWS) results from abnormalities in the genomic imprinting process leading to hypothalamic dysfunction with an alteration of growth hormone (GH) secretion. PWS is associated with early morbid obesity and short stature which can be efficiently improved with GH treatment. OBJECTIVES: Our aims were to highlight adipose tissue structural and functional impairments in children with PWS and to study the modifications of those parameters on GH treatment. SUBJECTS AND METHODS: Plasma samples and adipose tissue biopsies were obtained from 23 research centers in France coordinated by the reference center for PWS in Toulouse, France. Lean controls (n=33), non-syndromic obese (n=53), untreated (n=26) and GH-treated PWS (n=43) children were enrolled in the study. Adipose tissue biopsies were obtained during scheduled surgeries from 15 lean control, 7 untreated and 8 GH-treated PWS children. RESULTS: Children with PWS displayed higher insulin sensitivity as shown by reduced glycemia, insulinemia and HOMA-IR compared with non-syndromic obese children. In contrast, plasma inflammatory cytokines such as TNF-α, MCP-1 and IL-8 were increased in PWS. Analysis of biopsies compared with control children revealed decreased progenitor cell content in the stromal vascular fraction of adipose tissue and an impairment of lipolytic response to ß-adrenergic agonist in PWS adipocytes. Interestingly, both of these alterations in PWS seem to be ameliorated on GH treatment. CONCLUSION: Herein, we report adipose tissue dysfunctions in children with PWS which may be partially restored by GH treatment.

Lysophosphatidic acid impairs glucose homeostasis and inhibits insulin secretion in high-fat diet obese mice.

AIMS/HYPOTHESIS: Lysophosphatidic acid (LPA) is a lipid mediator produced by adipocytes that acts via specific G-protein-coupled receptors; its synthesis is modulated in obesity. We previously reported that reducing adipocyte LPA production in high-fat diet (HFD)-fed obese mice is associated with improved glucose tolerance, suggesting a negative impact of LPA on glucose homeostasis. Here, our aim was to test this hypothesis. METHODS: First, glucose tolerance and plasma insulin were assessed after acute (30 min) injection of LPA (50 mg/kg) or of the LPA1/LPA3 receptor antagonist Ki16425 (5 mg kg(-1) day(-1), i.p.) in non-obese mice fed a normal diet (ND) and in obese/prediabetic (defined as glucose-intolerant) HFD mice. Glucose and insulin tolerance, pancreas morphology, glycogen storage, glucose oxidation and glucose transport were then studied after chronic treatment (3 weeks) of HFD mice with Ki16425. RESULTS: In ND and HFD mice, LPA acutely impaired glucose tolerance by inhibiting glucose-induced insulin secretion. These effects were blocked by pre-injection of Ki16425 (5 mg/kg, i.p.). Inhibition of glucose-induced insulin secretion by LPA also occurred in isolated mouse islets. Plasma LPA was higher in HFD mice than in ND mice and Ki16425 transiently improved glucose tolerance. The beneficial effect of Ki16425 became permanent after chronic treatment and was associated with increased pancreatic islet mass and higher fasting insulinaemia. Chronic treatment with Ki16425 also improved insulin tolerance and increased liver glycogen storage and basal glucose use in skeletal muscle. CONCLUSIONS/INTERPRETATION: Exogenous and endogenous LPA exerts a deleterious effect on glucose disposal through a reduction of plasma insulin; pharmacological blockade of LPA receptors improves glucose homeostasis in obese/prediabetic mice.

Hypothalamic actions of apelin on energy metabolism: new insight on glucose homeostasis and metabolic disorders.

Hypothalamus is key area implicated in control of glucose homeostasis. This structure integrates nervous and peripheral informations to adapt a response modifying peripheral glucose utilization and maintaining energetic balance. Among peripheral signals, adipokines such as adiponectin and leptin are of special importance since deregulations of their actions are closely associated to metabolic disorders such as obesity and type 2 diabetes. During the past ten years, we have identified a new adipokine named apelin which has emerging role in the control of metabolism. The originality of the apelinergic system is to be largely represented in peripheral tissues (adipose tissue, intestine, etc.) and in the brain. Then, apelin is released by adipose tissue as all adipokines, but also present another crucial role as neurotransmitter in hypothalamic neurons. By acting in the whole body, apelin exerts pleiotropic actions and is now considered as a major determinant of physiological functions. Besides its general beneficial effects on peripheral targets, central action of apelin remains still a matter of debate. In this review, we have made a parallel between peripheral vs. central actions of apelin in term of signalization and effects. Then, we have focused our attention on hypothalamic apelin and its potential role in glucose metabolism and associated pathologies.

Apelin controls emotional behavior in age- and metabolic state-dependent manner.

Apelin is a small peptide secreted by the adipose tissue notably in conditions of obesity-induced hyper-insulinemia. Apelin exerts a range of physiological functions at the periphery including the improvement of insulin sensitivity and the increase of muscle strength or cardiac contractibility. Interestingly, the brain is endowed with a high density of APJ, the single target of apelin, and growing evidence suggests various central actions of this adipokine. Recent studies reported that the intracerebroventricular infusion of apelin modulates emotional states in middle age stressed animals. However, results are so far been mixed and have not allowed for definitive conclusions about the impact of apelin on anxio-depressive-like phenotype. This study aims 1) to evaluate whether serum apelin levels are associated with mood in older adults and 2) to determine the impact of the genetic apelin inactivation in 12-month old mice fed a standard diet (STD) or in 6-month old mice fed a high fat diet (HFD). A higher plasma apelin level was associated with higher depressive symptoms in older adults. In line with these clinical findings, 12-month old apelin knock-out (Ap-/-) mice displayed a spontaneous antidepressant-like phenotype. In a marked contrast, 6-month old Ap-/- mice harbored a higher degree of peripheral insulin resistance than wild-types in response to HFD and were more prone to develop anxiety while the depressive-like state was not modified. We also provided evidence that such anxious behavior was associated with an impairment of central serotonergic and dopaminergic neuronal activities. Finally, although the insulin sensitizing drug metformin failed to reverse HFD-induced insulin resistance in 6-month old Ap-/- mice, it reversed their anxious phenotype. These results emphasize a complex contribution of apelin in the regulation of emotional state that might depend on the age and the metabolic status of the animals. Further investigations are warranted to highlight the therapeutic potential of manipulating the apelinergic system in mood-related disorders.

Circulating Levels of Apelin, GDF-15 and Sarcopenia: Lack of Association in the MAPT Study.

OBJECTIVES: Apelin and GDF-15 have been proposed as biomarkers of age-related sarcopenia but evidence in human models is scarce. This study aimed to explore the associations between blood apelin and GDF-15 with sarcopenia incidence and the evolution of sarcopenia components over two years in older adults >70 years. DESIGN: Secondary longitudinal analysis of the Multidomain Alzheimer Preventive Trial. PARTICIPANTS: Older adults (>70 years) attending primary care centers in France and Monaco. SETTING: Community. MEASUREMENTS: Serum Apelin (pg/mL) and plasma GDF-15 (pg/mL) were measured. Outcomes included sarcopenia defined by the European Working Group on Sarcopenia in Older People (EWGSOP) and its determinants (appendicular lean mass [ALM] evaluated through a Dual-energy X-ray Absorptiometry (DXA) scan, handgrip strength (HGS) and the 4-meter gait speed) measured over 2 years. Linear mixed models and logistic regression were used to explore the longitudinal associations. RESULTS: We included 168 subjects from MAPT (median age=76y, IQR=73-79; 78% women). Serum apelin was not significantly associated with sarcopenia incidence (OR=1.001;95%CI=1.000,1.001;p-value>0.05 in full-adjusted models) nor with ALM (ß=-5.8E-05;95%CI=-1.0E-04,2.12E-04;p>0.05), HGS (ß=-1.1E-04;95%CI=-5.0E-04,2.8E-04;p>0.05), and GS (ß=-5.1E-06;95%CI=-1.0E-05,2.0E-05;p>0.05) in fully adjusted models. Similarly, plasma GDF-15 was not associated with both the incidence of sarcopenia (OR=1.001,95%CI=1.000,1.002,p>0.05) and the evolution of its determinants ([ALM, ß=2.1E-05;95%CI=-2.6E-04,3.03E-04;p>0.05], HGS [ß=-5.9E-04;95%CI=-1.26E-03,8.1E-05; p>0.05] nor GS [ß=-2.6E-06;95%CI=-3.0E-05, 2.3E-05;p>0.05]) in fully adjusted models. CONCLUSIONS: Blood apelin and GDF-15 were not associated with sarcopenia incidence or with the evolution of sarcopenia components over a 2-year follow-up in community-dwelling older adults. Well-powered longitudinal studies are needed to confirm or refute our findings.

A gerophysiology perspective on healthy ageing.

Improvements in public health and health care have resulted in significant increases in lifespan globally, but also in a significant increase in chronic disease prevalence. This has led to a focus on healthy ageing bringing a shift from a pathology-centered to an intrinsic capacity and function-centered view. In parallel, the emerging field of geroscience has promoted the exploration of the biomolecular drivers of ageing towards a transverse vision by proposing an integrated set of molecular hallmarks. In this review, we propose to take a step further in this direction, highlighting a gerophysiological perspective that considers the notion of homeostasis/allostasis relating to robustness/fragility respectively. While robustness is associated with homeostasis achieved by an optimal structure/function relationship in all organs, successive repair processes occurring after daily injuries and infections result in accumulation of scar healing leading to progressive tissue degeneration, allostasis and frailty. Considering biological ageing as the accumulation of scarring at the level of the whole organism emphasizes three transverse and shared elements in the body - mesenchymal stroma cells/immunity/metabolism (SIM). This SIM tryptich drives tissue and organ fate to regulate the age-related evolution of body functions. It provides the basis of a gerophysiology perspective, possibly representing a better way to decipher healthy ageing, not only by defining a composite biomarker(s) but also by developing new preventive/curative strategies.

The therapeutic potentials of apelin in obesity-associated diseases.

Apelin, a peptide with several active isoforms ranging from 36 to 12 amino acids and its receptor APJ, a G-protein-coupled receptor, are widely distributed. However, apelin has emerged as an adipokine more than fifteen years ago, integrating the field of inter-organs interactions. The apelin/APJ system plays important roles in several physiological functions both in rodent and humans such as fluid homeostasis, cardiovascular physiology, angiogenesis, energy metabolism. Thus the apelin/APJ system has generated great interest as a potential therapeutic target in different pathologies. The present review will consider the effects of apelin in metabolic diseases such as obesity and diabetes with a focus on diabetic cardiomyopathy among the complications associated with diabetes and APJ agonists or antagonists of interest in these diseases.

The INSPIRE Research Initiative: A Program for GeroScience and Healthy Aging Research Going from Animal Models to Humans and the Healthcare System.

Aging is the most important risk factor for the onset of several chronic diseases and functional decline. Understanding the interplays between biological aging and the biology of diseases and functional loss as well as integrating a function-centered approach to the care pathway of older adults are crucial steps towards the elaboration of preventive strategies (both pharmacological and non-pharmacological) against the onset and severity of burdensome chronic conditions during aging. In order to tackle these two crucial challenges, ie, how both the manipulation of biological aging and the implementation of a function-centered care pathway (the Integrated Care for Older People (ICOPE) model of the World Health Organization) may contribute to the trajectories of healthy aging, a new initiative on Gerosciences was built: the INSPIRE research program. The present article describes the scientific background on which the foundations of the INSPIRE program have been constructed and provides the general lines of this initiative that involves researchers from basic and translational science, clinical gerontology, geriatrics and primary care, and public health.

The INSPIRE Bio-Resource Research Platform for Healthy Aging and Geroscience: Focus on the Human Translational Research Cohort (The INSPIRE-T Cohort).

BACKGROUND: The Geroscience field focuses on the core biological mechanisms of aging, which are involved in the onset of age-related diseases, as well as declines in intrinsic capacity (IC) (body functions) leading to dependency. A better understanding on how to measure the true age of an individual or biological aging is an essential step that may lead to the definition of putative markers capable of predicting healthy aging. OBJECTIVES: The main objective of the INStitute for Prevention healthy agIng and medicine Rejuvenative (INSPIRE) Platform initiative is to build a program for Geroscience and healthy aging research going from animal models to humans and the health care system. The specific aim of the INSPIRE human translational cohort (INSPIRE-T cohort) is to gather clinical, digital and imaging data, and perform relevant and extensive biobanking to allow basic and translational research on humans. METHODS: The INSPIRE-T cohort consists in a population study comprising 1000 individuals in Toulouse and surrounding areas (France) of different ages (20 years or over - no upper limit for age) and functional capacity levels (from robustness to frailty, and even dependency) with follow-up over 10 years. Diversified data are collected annually in research facilities or at home according to standardized procedures. Between two annual visits, IC domains are monitored every 4-month by using the ICOPE Monitor app developed in collaboration with WHO. Once IC decline is confirmed, participants will have a clinical assessment and blood sampling to investigate markers of aging at the time IC declines are detected. Biospecimens include blood, urine, saliva, and dental plaque that are collected from all subjects at baseline and then, annually. Nasopharyngeal swabs and cutaneous surface samples are collected in a large subgroup of subjects every two years. Feces, hair bulb and skin biopsy are collected optionally at the baseline visit and will be performed again during the longitudinal follow up. EXPECTED RESULTS: Recruitment started on October 2019 and is expected to last for two years. Bio-resources collected and explored in the INSPIRE-T cohort will be available for academic and industry partners aiming to identify robust (set of) markers of aging, age-related diseases and IC evolution that could be pharmacologically or non-pharmacologically targetable. The INSPIRE-T will also aim to develop an integrative approach to explore the use of innovative technologies and a new, function and person-centered health care pathway that will promote a healthy aging.

Healthy Aging Biomarkers: The INSPIRE's Contribution.

The find solutions for optimizing healthy aging and increase health span is one of the main challenges for our society. A novel healthcare model based on integration and a shift on research and care towards the maintenance of optimal functional levels are now seen as priorities by the WHO. To address this issue, an integrative global strategy mixing longitudinal and experimental cohorts with an innovative transverse understanding of physiological functioning is missing. While the current approach to the biology of aging is mainly focused on parenchymal cells, we propose that age-related loss of function is largely determined by three elements which constitute the general ground supporting the different specific parenchyma: i.e. the stroma, the immune system and metabolism. Such strategy that is implemented in INSPIRE projects can strongly help to find a composite biomarker capable of predicting changes in capacity across the life course with thresholds signalling frailty and care dependence.

Towards a Large-Scale Assessment of the Relationship between Biological and Chronological Aging: The INSPIRE Mouse Cohort.

Aging is the major risk factor for the development of chronic diseases. After decades of research focused on extending lifespan, current efforts seek primarily to promote healthy aging. Recent advances suggest that biological processes linked to aging are more reliable than chronological age to account for an individual's functional status, i.e. frail or robust. It is becoming increasingly apparent that biological aging may be detectable as a progressive loss of resilience much earlier than the appearance of clinical signs of frailty. In this context, the INSPIRE program was built to identify the mechanisms of accelerated aging and the early biological signs predicting frailty and pathological aging. To address this issue, we designed a cohort of outbred Swiss mice (1576 male and female mice) in which we will continuously monitor spontaneous and voluntary physical activity from 6 to 24 months of age under either normal or high fat/high sucrose diet. At different age points (6, 12, 18, 24 months), multiorgan functional phenotyping will be carried out to identify early signs of organ dysfunction and generate a large biological fluids/feces/organs biobank (100,000 samples). A comprehensive correlation between functional and biological phenotypes will be assessed to determine: 1) the early signs of biological aging and their relationship with chronological age; 2) the role of dietary and exercise interventions on accelerating or decelerating the rate of biological aging; and 3) novel targets for the promotion of healthy aging. All the functional and omics data, as well as the biobank generated in the framework of the INSPIRE cohort will be available to the aging scientific community. The present article describes the scientific background and the strategies employed for the design of the INSPIRE Mouse cohort.

Revisiting the Hallmarks of Aging to Identify Markers of Biological Age.

The Geroscience aims at a better understanding of the biological processes of aging, to prevent and/or delay the onset of chronic diseases and disability as well as to reduce the severity of these adverse clinical outcomes. Geroscience thus open up new perspectives of care to live a healthy aging, that is to say without dependency. To date, life expectancy in healthy aging is not increasing as fast as lifespan. The identification of biomarkers of aging is critical to predict adverse outcomes during aging, to implement interventions to reduce them, and to monitor the response to these interventions. In this narrative review, we gathered information about biomarkers of aging under the perspective of Geroscience. Based on the current literature, for each hallmark of biological aging, we proposed a putative biomarker of healthy aging, chosen for their association with mortality, age-related chronic diseases, frailty and/or functional loss. We also discussed how they could be validated as useful predictive biomarkers.

Apelin/APJ signaling system: a potential link between adipose tissue and endothelial angiogenic processes.

Adipose tissue is an active endocrine organ that produces a variety of secretory factors involved in the initiation of angiogenic processes. The bioactive peptide apelin is the endogenous ligand of the G protein-coupled receptor, APJ. Here we investigated the potential role of apelin and its receptor, APJ, in the angiogenic responses of human endothelial cells and the development of a functional vascular network in a model of adipose tissue development in mice. Treatment of human umbilical vein endothelial cells with apelin dose-dependently increased angiogenic responses, including endothelial cell migration, proliferation, and Matrigel(R) capillary tubelike structure formation. These endothelial effects of apelin were due to activation of APJ, because siRNA directed against APJ, which led to long-lasting down-regulation of APJ mRNA, abolished cell migration induced by apelin in contrast to control nonsilencing siRNA. Hypoxia up-regulated the expression of apelin in 3T3F442A adipocytes, and we therefore determined whether apelin could play a role in adipose tissue angiogenesis in vivo. Epididymal white adipose tissue (EWAT) transplantation was performed as a model of adipose tissue angiogenesis. Transplantation led to increased apelin mRNA levels 2 and 5 days after transplantation associated with tissue hypoxia, as evidenced by hydroxyprobe staining on tissue sections. Graft revascularization evolved in parallel, as the first functional vessels in EWAT grafts were observed 2 days after transplantation and a strong angiogenic response was apparent on day 14. This was confirmed by determination of graft hemoglobin levels, which are indicative of functional vascularization and were strongly increased 5 and 14 days after transplantation. The role of apelin in the graft neovascularization was then assessed by local delivery of stable complex apelin-targeting siRNA leading to dramatically reduced apelin mRNA levels and vascularization (quantified by hemogloblin content) in grafted EWAT on day 5 when compared with control siRNA. Taken together, our data provide the first evidence that apelin/APJ signaling pathways play a critical role in the development of the functional vascular network in adipose tissue. In addition, we have shown that adipocyte-derived apelin can be up-regulated by hypoxia. These findings provide novel insights into the complex relationship between adipose tissue and endothelial vascular function and may lead to new therapeutic strategies to modulate angiogenesis.

Native fructose extracted from apple improves glucose tolerance in mice.

Fructose is one of the most abundant monosaccharide in nature. It is also the sweetest naturally occurring carbohydrate. Since decades, fructose used for food preparations is not provided by fruit or vegetable but by a chemical process of starch or inulin conversion. We processed a new method of fructose extraction from apple and investigated the acute and long term effect of this carbohydrate on glucose metabolism in C57Bl6/j mice. By using the glycemic index (GI), we have shown that one of the sugars obtained from apple, FructiLight, has a very low impact on glycemic and insulin response during acute treatment compared to other sugars. This carbohydrate, essentially constituted by fructose, has also beneficial properties when administrated for long term treatment. Indeed, as two other sugars extracted from apple (FructiSweetApple and FructiSweet67), FructiLight exposure during 21 weeks in beverage has promoted an enhancement of glucose tolerance compared to glucose treatment without affecting food intake and weight. All these results indicate that apple-extracted sugars and more precisely fructose from these fruits could be a promising way to produce new food and sweet beverages.

Neuropeptide Y and peptide YY inhibit lipolysis in human and dog fat cells through a pertussis toxin-sensitive G protein.

Neuropeptide Y (NPY) and peptide YY (PYY) are regulatory peptides that have considerable sequence homology with pancreatic polypeptide. Because (a) NPY has been shown to be colocalized with noradrenaline in peripheral as well as central catecholaminergic neurons, and (b) alpha 2-adrenergic receptors of adipocytes play a major role in the regulation of lipolysis, we investigated the effect of NPY and PYY on isolated fat cells. In human fat cells NPY and PYY promoted a dose-dependent inhibition of lipolysis elicited by 2 micrograms/ml adenosine deaminase (removal of adenosine) whatever the lipolytic index used (glycerol or nonesterified fatty acids). In dog fat cells NPY and PYY inhibited adenosine deaminase-, isoproterenol- and forskolin-induced lipolysis. In humans and dogs the effects of NPY or PYY were abolished by treatment of cells with Bordetella pertussis toxin, clearly indicating the involvement of a Gi protein in the antilipolytic effects. This study indicates that, in addition to alpha 2-adrenergic agonists, NPY and PYY are also involved in the regulation of lipolysis in human and dog adipose tissue as powerful antilipolytic agents. Further studies are needed to characterize the pharmacological nature of the receptor mediating the inhibitory effect of NPY and PYY in fat cells.

Alpha2-adrenergic receptor-mediated release of lysophosphatidic acid by adipocytes. A paracrine signal for preadipocyte growth.

In the search for the existence of adrenergic regulation of the autocrine/paracrine function of the white adipose tissue, it was observed that conditioned media from isolated adipocytes or dialysates obtained by in situ microdialysis of human subcutaneous adipose tissue increased spreading and proliferation of 3T3F442A preadipocytes. These effects were amplified when an alpha2-adrenergic agonist was present during the obtention of conditioned media and microdialysates. This alpha2-adrenergic-dependent trophic activity was completely abolished by pretreatment of the conditioned media or microdialysates with the lysophospholipase, phospholipase B. Among the different lysophospholipids tested only lysophosphatidic acid (LPA) was able to induce spreading and proliferation of 3T3F442A preadipocytes. Moreover, previous chronic treatment of 3T3F442A preadipocytes with LPA which led to a specific desensitization of LPA responsiveness, abolished the alpha2-adrenergic-dependent trophic activities of the conditioned media and microdialysates. Finally, alpha2-adrenergic stimulation led to a rapid, sustained, and pertussis toxin-dependent release of [32P]LPA from [32P]-labeled adipocytes. Based upon these results it was proposed that in vitro and in situ stimulation of adipocyte alpha2-adrenergic receptors provokes the extracellular release of LPA leading, in turn, to regulation of preadipocyte growth.

Characterization and distribution of alpha 2-adrenergic receptors in the human intestinal mucosa.

The subtype and the expression of the alpha 2-adrenergic receptor were investigated in the normal mucosa from human intestine by means of radioligand binding, RNase mapping, and measurement of adenylate cyclase activity. The study of the binding of the alpha 2-adrenergic antagonist, [3H]RX821002, to epithelial cell membranes indicated the existence of a single class of noninteracting sites displaying a high affinity for the radioligand (Kd = 1.1 +/- 0.5 nM). The rank order of potency of antagonists to inhibit [3H]RX821002 binding (RX821002 > yohimbine = rauwolscine > phentolamine approximately idazoxan >> chlorpromazine > prazosin) suggested that the receptor is of the alpha 2A subtype. A conclusion which is confirmed by the fact that only alpha 2C10 transcripts were found in the human intestine mucosa. Competition curves with (-)-norepinephrine demonstrated that 60% of the receptor population exhibited high affinity for agonists. This high-affinity state was abolished by the addition of GTP plus Na+ or by prior treatment of the membranes with pertussis toxin indicating it corresponded to G protein-coupled receptors. [32P]ADP-ribosylation and immunoblotting experiments identified two pertussis toxin-sensitive G proteins corresponding to Gi2 and Gi3. The study of the distribution of the receptor indicated that (a) the proximal colon is the intestine segment exhibiting the highest receptor density and (b) the receptor is predominantly expressed in crypts and is preferentially located in the basolateral membrane of the polarized cell. The distribution of the receptor along the crypt-surface axis of the colon mucosa can be correlated with a higher level of alpha 2C10-specific mRNA and a higher efficiency of UK14304 to inhibit adenylate cyclase in crypt cells.

Caffeine reduces TNFalpha up-regulation in human adipose tissue primary culture.

Adipose tissue secretions play an important role in the development of obesity-related pathologies such as diabetes. Through inflammatory cytokines production, adipose tissue stromavascular fraction cells (SVF), and essentially macrophages, promote adipocyte insulin resistance by a paracrine way. Since xanthine family compounds such as caffeine were shown to decrease inflammatory production by human blood cells, we investigated the possible effect of caffeine on Tumor Necrosis Factor alpha (TNFalpha) and Interleukin-6 (IL-6) expression by human adipose tissue primary culture. For that purpose, human subcutaneous adipose tissue obtained from healthy non-obese women (BMI: 26.7 +/- 2.2 kg/m2) after abdominal dermolipectomy, was split into explants and cultured for 6 hours with or without caffeine. Three different concentrations of caffeine were tested (0.5 microg/mL, 5 microg/mL and 50 microg/mL). After 6 hours of treatment, explants were subjected to collagenase digestion in order to isolate adipocytes and SVF cells. Then, TNFalpha and IL-6 mRNA were analysed by real-time PCR alternatively in adipocytes and SVF cells. In parallel, we checked gene expression of markers involved in adipocyte differenciation and in SVF cells inflammation and proliferation. Our findings show a strong and dose dependent down-regulation of TNF-alpha gene expression in both adipocyte and SVF cells whereas IL-6 was only down regulated in SVF cells. No effect of caffeine was noticed on the other genes studied. Thus, caffeine, by decreasing TNFalpha expression, could improve adipose tissue inflammation during obesity.