GDFI5, un nou factor secretable regulador del metabolisme. Paper com a "batoquina" en models experimentals, i estudis en humans

[cat] L’obesitat s’ha convertit en les últimes dècades en un greu problema per la salut pública arribant a ser una de les principals causes de morbiditat, incapacitació i mortalitat prematura. Un teixit que pot tenir un paper important en la protecció contra l’obesitat és el teixit adipós marró (TAM). El TAM és un dels principals teixits consumidors d’energia en els mamífers. El TAM, descrit primerament en rosegadors i en nounats ha anat guanyant rellevància la última dècada amb el descobriment de la presència d’aquest i la seva activació front a estímuls de fred en humans adults. Recentment, s’ha descrit un paper secretor del TAM. En resposta a l’activació termogènica el TAM allibera factors que actuen de manera autocrina, paracrina o endocrina reforçant aquesta activació. A aquests factors secretats pel TAM se’ls anomena adipoquines marrons o batoquines. Ara fa aproximadament uns 20 anys sis laboratoris van publicar en poc temps la descripció d’un nou factor secretable de la família del TGFβ, anomenat GDF15. GDF15 és coneguda principalment com una citocina de resposta a estrès. La seva expressió augmenta en resposta tant a situacions d’estrès patològic, com gran varietat de malalties, com en situacions d’estrès fisiològiques com ara durant la gestació. Recentment, s’ha descobert un receptor de GDF15 al cervell, GFRAL. S’ha vist que GDF15 actua a nivell central com a factor anorexigènic. Tot i això, altres estudis postulen que GDF15 té altres accions en teixits perifèrics. En aquesta tesi, s’ha identificat el factor GDF15 com una adipoquina marró o batoquina, alliberada pels adipòcits marrons i beix en resposta a l’activació termogènica d’aquestes cèl·lules. S’ha determinat la via adrenèrgica β3 com la responsable de l’augment de l’expressió i secreció de GDF15 en resposta a l’estímul termogènic, i s’han trobat implicades proteïnes d’aquesta via com la PKA i adipoquines marrons prèviament identificades, com FGF21. A més a més, s’ha observat que el GDF15 produït pels adipòcits té afectes en els macròfags residents dins del teixit, reprimint-ne la inflamació. Aquests resultants suggereixen que la secreció de GDF15 per part dels adipòcits marrons pot ser un mecanisme per regular localment l’activitat pro-inflamatòria dels macròfags associada amb una millora de l’activitat del teixit adipós marró. Paral·lelament, s’han realitzat diferents estudis sobre GDF15 en poblacions humanes. S’ha vist com GDF15 augmenta en patologia mitocondrial en nens i s’han determinat els miotubs com les cèl·lules responsables d’aquest augment. També s’han analitzat els nivells circulants de GDF15 durant el desenvolupament postnatal, observant una possible associació entre aquests nivells i el pes dels nounats en el moment del naixement. Per últim, s’ha avaluat la resposta de GDF15 a un exercici intens; s’ha vist un dràstic augment de GDF15 circulant en atletes no professionals en acabar la marató de Barcelona del 2017, que era totalment revertit dos dies després de la realització de la cursa. En més d’un dels estudis s’ha observat un paral·lelisme entre els nivells de GDF15 i els nivells d’FGF21, tot i això sembla ser que aquestes dues proteïnes que sovint augmenten en paral·lel actuen a través de vies totalment independents. En resum, en aquesta tesi hem estudiat la proteïna GDF15 en situacions molt diverses; patològiques i fisiològiques, sistèmiques i locals i en models humans i de ratolí. GDF15 sembla ser una important proteïna reguladora del metabolisme, amb efectes molt diversos en funció de teixit i condició. Regula la termogènesi localment en el teixit adipós, esta implicada en el desenvolupament post-natal actuant a nivell central i respon a situacions d’estrès tant patològiques (patologia mitocondrial) com fisiològiques (exercici). És necessari, però, seguir investigant per tal d’entendre els mecanismes i la importància biològica de GDF15 en cadascuna d’aquestes situacions.[eng] Brown adipose tissue (BAT) is the main site of energy expenditure in mammals, via adaptive thermogenesis. The thermogenic capacity of BAT occurs through the unique expression of the uncoupling protein UCP1 and high oxidative capacity of brown adipocytes. High BAT activity favors a negative energy balance, protecting against obesity. GDF15 (growth and differentiation factor-15) is a member of the TGFβ superfamily and is secreted by the liver, placenta and other organs. Its biological function is unclear, but it has been proposed as a biomarker of various pathological states, such as inflammatory diseases, cardiovascular disease, or obesity. Recently, several studies reported the presence of a specific receptor for GDF15 in the brain, known as GFRAL, which mediates the anorexigenic response to this molecule. In this thesis we were able to analyze this protein in different mouse and human models and in different physiological and pathological conditions. In mice, cold exposure strongly induced GDF15 expression in BAT, norepinephrine and cAMP induced GDF15 expression and release by cells through protein kinase A-mediated mechanisms and the GDF15 released by brown adipocytes targeted macrophages, and down-regulated the expression of pro-inflammatory genes. These results indicated that GDF15 is a novel brown adipokine released by brown adipocytes in response to thermogenic activation. In humans, children with mitochondrial diseases have increased GDF15 levels compared to healthy controls, proposing GDF15 as a biomarker for these diseases. GDF15, as well as FGF21, mRNA expression and protein secretion, were significantly induced after treatment of myotubes with drugs that block the respiratory chain, indicating that myotubes are the responsible cells for GDF15 production in these conditions. In another study, GDF15 levels appeared significantly high at birth and dropped at age 4 months, particularly in infants born with lower weight, and continued to decline progressively in both subgroups reaching adult concentrations by age 24 months. Finally, we assessed the value of GDF15 and FGF21 as biomarkers in relation to the biological response to strenuous exercise. A marked increase in the mean levels of GDF15 and FGF21 was found immediately after a marathon race. These levels returned to basal values 48h post-race

Altered GDF15 and FGF21 Levels in Response to Strenuous Exercise: A Study in Marathon Runners

Background: Recreational marathon runners face strong physiological challenges. Assessment of potential biomarkers for the biological responses of runners will help to discriminate individual race responsiveness and their physiological consequences. This study sought to analyze the changes in the plasma levels of GDF15 and FGF21, novel endocrine factors related to metabolic stress, in runners following the strenuous exercise of a marathon race. Methods: Blood samples were obtained from eighteen male runners (mean ±SD, age: 41.7 ±5.0 years, BMI: 23.6 ± 1.8) 48 h before, immediately after, and 48 h after a marathon race, and from age-matched sedentary individuals. The level of GDF15, FGF21, and 38 additional biochemical and hematological parameters were determined. Results: The basal levels of GDF15 and FGF21 did not differ between runners before the race and sedentary individuals. Significant increases in the mean levels of GDF15 (4.2-fold) and FGF21 (20-fold) were found in runners immediately after the race. The magnitudes of these increases differed markedly among individuals and did not correlate with each other. The GDF15 and FGF21 levels had returned to the basal level 48 h post-race. The post-race value of GDF15 (but not FGF21) correlated positively with increased total white cell count (r = 0.50, P = 0.01) and neutrophilia (r = 0.10, P = 0.01). Conclusion: GDF15 and FGF21 are transiently increased in runners following a marathon race. The induction of GDF15 levels is associated with alterations in circulating immune cells levels

GPR120 controls neonatal brown adipose tissue thermogenic induction

Adaptive induction of thermogenesis in brown adipose tissue (BAT) is essential for the survival of mammals after birth. We herein show that G-coupled receptor protein-120 (GPR120) expression is dramatically induced after birth in mouse BAT. GPR120 expression in neonatal BAT is the highest among GPR120-expressing tissues in mouse at any developmental stage tested. The induction of GPR120 in neonatal BAT is caused by the postnatal thermal stress rather than by the initiation of suckling. GPR120-null neonates were found to be relatively intolerant to cold: close to one-third did not survive at 21ºC, but all such pups survived at 25ºC. Heat production in BAT was significantly impaired in GPR120-null pups. Deficiency in GPR120 did not modify brown adipocyte morphology or the anatomical architecture of BAT, as assessed by electron microscopy, but instead impaired the expression of UCP1 and the fatty acid oxidation capacity of neonatal BAT. Moreover, GPR120 deficiency impaired FGF21 gene expression in BAT and reduced plasma FGF21 levels. These results indicate that GPR120 is essential for neonatal adaptive thermogenesis through the control of the FGF21 system

The kallikrein-kinin pathway as a mechanism for auto-control of brown adipose tissue activity

Brown adipose tissue (BAT) is known to secrete regulatory factors in response to thermogenic stimuli. Components of the BAT secretome may exert local effects that contribute to BAT recruitment and activation. Here, we found that a thermogenic stimulus leads to enhanced secretion of kininogen (Kng) by BAT, owing to induction of kininogen 2 (Kng2) gene expression. Noradrenergic, cAMP-mediated signals induce KNG2 expression and release in brown adipocytes. Conversely, the expression of kinin receptors, that are activated by the Kng products bradykinin and [Des-Arg9]-bradykinin, are repressed by thermogenic activation of BAT in vivo and of brown adipocytes in vitro. Loss-of-function models for Kng (the circulating-Kng-deficient BN/Ka rat) and bradykinin (pharmacological inhibition of kinin receptors, kinin receptor-null mice) signaling were coincident in showing abnormal overactivation of BAT. Studies in vitro indicated that Kng and bradykinin exert repressive effects on brown adipocyte thermogenic activity by interfering the PKA/p38 MAPK pathway of control of Ucp1 gene transcription, whereas impaired kinin receptor expression enhances it. Our findings identify the kallikrein-kinin system as a relevant component of BAT thermogenic regulation that provides auto-regulatory inhibitory signaling to BAT

Differential association between S100A4 levels and insulin resistance in prepubertal children and adult subjects with clinically severe obesity

Objectives: S100A4 has been recently identified as an adipokine associated with insulin resistance (IR) in adult subjects with obesity. However, no data about its levels in children with obesity and only a few approaches regarding its potential mechanism of action have been reported. To obtain a deeper understanding of the role of S100A4 in obesity, (a) S100A4 levels were measured in prepubertal children and adult subjects with and without obesity and studied the relationship with IR and (b) the effects of S100A4 in cultured human adipocytes and vascular smooth muscle cells (VSMCs) were determined. Methods: Sixty-five children (50 with obesity, age 9.0 ±1.1 years and 15 normal weight, age 8.4 ±0.8 years) and fifty-nine adults (43 with severe obesity, age 46 ±11 years and 16 normal weight, age 45 ±9 years) were included. Blood from children and adults and adipose tissue samples from adults were obtained and analysed. Human adipocytes and VSMC were incubated with S100A4 to evaluate their response to this adipokine. Results: Circulating S100A4 levels were increased in both children (P = .002) and adults (P < .001) with obesity compared with their normal-weight controls. In subjects with obesity, S100A4 levels were associated with homeostatic model assessment-insulin resistance (HOMA-IR) in adults (βstd = .42, P = .008) but not in children (βstd = .12, P = .356). Human adipocytes were not sensitive to S100A4, while incubation with this adipokine significantly reduced inflammatory markers in VSMC. Conclusions: Our human data demonstrate that higher S100A4 levels are a marker of IR in adults with obesity but not in prepubertal children. Furthermore, the in vitro results suggest that S100A4 might exert an anti-inflammatory effect. Further studies will be necessary to determine whether S100A4 can be a therapeutic target for obesity

A role for Oncostatin M in the impairment of glucose homeostasis in obesity

CONTEXT: Oncostatin M (OSM) plays a key role in inflammation, but its regulation and function during obesity is not fully understood. OBJECTIVE: The aim of this study was to evaluate the relationship of OSM with the inflammatory state that leads to impaired glucose homeostasis in obesity. We also assessed whether OSM immunoneutralization could revert metabolic disturbances caused by a high-fat diet (HFD) in mice. DESIGN: 28 patients with severe obesity were included and stratified into two groups: (1) glucose levels 100 mg/dL. White adipose tissue was obtained to examine OSM gene expression. Human adipocytes were used to evaluate the effect of OSM in the inflammatory response, and HFD-fed C57BL/6J mice were injected with anti-OSM antibody to evaluate its effects. RESULTS: OSM expression was elevated in subcutaneous and visceral fat from patients with obesity and hyperglycemia, and correlated with Glut4 mRNA levels, serum insulin, homeostatic model assessment of insulin resistance, and inflammatory markers. OSM inhibited adipogenesis and induced inflammation in human adipocytes. Finally, OSM receptor knockout mice had increased Glut4 mRNA levels in adipose tissue, and OSM immunoneutralization resulted in a reduction of glucose levels and Ccl2 expression in adipose tissue from HFD-fed mice. CONCLUSIONS: OSM contributes to the inflammatory state during obesity and may be involved in the development of insulin resistance

GDF-15 is elevated in children with mitochondrial diseases and is induced by mitochondrial dysfunction

Background We previously described increased levels of growth and differentiation factor 15 (GDF-15) in skeletal muscle and serum of patients with mitochondrial diseases. Here we evaluated GDF-15 as a biomarker for mitochondrial diseases affecting children and compared it to fibroblast-growth factor 21 (FGF-21). To investigate the mechanism of GDF-15 induction in these pathologies we measured its expression and secretion in response to mitochondrial dysfunction. Methods We analysed 59 serum samples from 48 children with mitochondrial disease, 19 samples from children with other neuromuscular diseases and 33 samples from aged-matched healthy children. GDF-15 and FGF-21 circulating levels were determined by ELISA. Results Our results showed that in children with mitochondrial diseases GDF-15 levels were on average increased by 11-fold (mean 4046pg/ml, 1492 SEM) relative to healthy (350, 21) and myopathic (350, 32) controls. The area under the curve for the receiver-operating-characteristic curve for GDF-15 was 0.82 indicating that it has a good discriminatory power. The overall sensitivity and specificity of GDF-15 for a cut-off value of 550pg/mL was 67.8% (54.4%-79.4%) and 92.3% (81.5%-97.9%), respectively. We found that elevated levels of GDF-15 and or FGF-21 correctly identified a larger proportion of patients than elevated lev- els of GDF-15 or FGF-21 alone. GDF-15, as well as FGF-21, mRNA expression and protein secretion, were significantly induced after treatment of myotubes with oligomycin and that levels of expression of both factors significantly correlated. Conclusions Our data indicate that GDF-15 is a valuable serum quantitative biomarker for the diagnosis of mitochondrial diseases in children and that measurement of both GDF-15 and FGF-21 improves the disease detection ability of either factor separately. Finally, we demonstrate for the first time that GDF-15 is produced by skeletal muscle cells in response to mitochon- drial dysfunction and that its levels correlate in vitro with FGF-21 level