Sonia Eiras

Adiponectin is synthesized and secreted by human and murine cardiomyocytes

Adiponectin is thought to play a decisive role in the relationships among obesity, insulin resistance and cardiovascular risk. This study investigated whether cardiomyocytes synthesize and secrete adiponectin, and the effects of this hormone on cardiac cells. RT‐PCR showed that mouse, rat and human cardiomyocytes produced mRNA for adiponectin and adiponectin receptors 1 and 2. Immunohistochemistry confirmed the presence of adiponectin in the cytoplasm of cultured cardiomyocytes, and radioimmunoassay showed that these cells secreted adiponectin into the culture medium. Exogenous adiponectin enhanced glucose and fatty acid uptake and induced AMPK phosphorylation in cultured cardiomyocytes. Our results demonstrate that adiponectin is synthesized and secreted by isolated murine and human cardiomyocytes, and suggest that the local production of this hormone by cardiomyocytes could be involved in the regulation of cardiac metabolism and function.

Elevated serum leptin concentrations induced by experimental acute inflammation

Leptin is a pleiotropic hormone that regulates body weight and energy expenditure. Recent findings suggest that leptin may be involved in acute and/or chronic inflammation, however only limited results are available describing the effects of in vivo models of acute inflammation on leptin secretion. The aim of this study was to evaluate serum leptin levels in response to two well-established models of acute inflammation in rats: carrageenan rat paw induced oedema and carrageenan induced pleurisy. Our results clearly show that leptin levels rise in rats in which both oedema and pleurisy were induced. Serum leptin levels in carrageenan induced paw oedema were 3.86+/-0.16 microg/L in comparison to 1.83+/-0.17 microg/L of control animals (p<0.001). A similar result was observed in carrageenan induced pleurisy animals in which leptin levels were 4.87+/-0.27 microg/L in comparison to 2.19+/-0.16 microg/L of control animals (p<0.001). The increase in leptin levels induced following carrageenan-induced pleurisy appears to be dependent on adrenal function and it is markedly blunted in adrenalectomized rats. Leptin levels in carrageenan induced pleurisy, carried out on adrenalectomized rats, were lower than intact inflamed animals, suggesting a possible involvement of endogenous glucocorticoids. In summary the results here presented show that: a) an elevated plasma leptin concentration was induced during experimental models of inflammation b) this increase is mediated to a large extent by glucocorticoids. In conclusion, acute experimental models of inflammation are associated with changes in circulating leptin suggesting a possible involvement of this hormone in the anorexia/cachexia that is frequently associated with inflammatory processes. Furthermore, our data indicate the existence of a feedback loop among glucocorticoids and leptin which might contribute to the immune response to lace the inflammatory process.

Extension of coronary artery disease is associated with increased IL-6 and decreased adiponectin gene expression in epicardial adipose tissue.

UNLABELLED Epicardial adipose tissue (EAT) expresses lower levels of adiponectin in patients with CAD and higher levels of inflammatory mediators such as IL-6 and leptin than subcutaneous adipose tissue. This showed one important role of EAT in coronary artery disease. However, the relationship of EAT adiponectin and IL-6 levels to the extension of coronary artery disease has not hitherto been determined. We sought to determine whether the levels of adiponectin and interleukin-6 (IL-6) mRNA in epicardial adipose tissue are associated with the extension of coronary artery disease (CAD). METHODS Angiographic and hormones expression were evaluated from epicardial and subcutaneous adipose tissue. 92 patients (58 CAD, 34 non-CAD) who underwent cardiac surgery. Adiponectin and IL-6 mRNA levels were measured by real time RT-PCR in epicardial and subcutaneous adipose tissue (SAT) following angiographic evaluation of their coronary arteries. RESULTS We found that epicardial adipose tissue of CAD expressed lower levels of adiponectin mRNA and higher levels of IL-6 mRNA than that of non-CAD patients. As the number of injured arteries rose, adiponectin mRNA levels decreased (r=-0.402, p<0.001) and IL-6 mRNA increased (r=0.514, p<0.001) in epicardial adipose tissue. CONCLUSIONS The extension of CAD is significantly associated with the expression of adiponectin and IL-6 mRNA in EAT. These findings suggest that low adiponectin and high IL-6 expression by EAT may contribute to CAD extension.

Proteomic analysis of epicardial and subcutaneous adipose tissue reveals differences in proteins involved in oxidative stress

Epicardial adipose tissue (EAT) is an endocrine organ adjacent to coronary arteries and myocardium without anatomy barriers. Locally produced adipokines may reflect or affect to cardiovascular physiology and pathology. Our aim was to study the protein expression profiles of EAT and subcutaneous adipose tissue (SAT) to identify local candidate molecules characterizing EAT in patients with cardiovascular disease. EAT and SAT samples were collected from 55 patients undergoing heart surgery. Proteins from these tissues were separated by two-dimensional (2D) gel electrophoresis, and differences between them were identified by MALDI-TOF/TOF spectra. Differences in protein levels were further investigated by real-time RT-PCR and Western blots, and production of reactive oxygen species (ROS) in EAT and SAT was evaluated by nitroblue tetrazolium chloride assays. ROS production was higher in EAT than SAT. We have found mRNA differences for catalase, glutathione S-transferase P, and protein disulfide isomerase, and 2D Western blots additionally showed post-translational differences for phosphoglycerate mutase 1; all four are related to oxidative stress pathways. EAT suffers greater oxidative stress than SAT in patients with cardiovascular diseases and exhibits associated proteomic differences that suggest the possibility of its association with myocardial stress in these patients.

Epicardial adipose tissue expression of adiponectin is lower in patients with hypertension

Low plasma adiponectin levels are related to a higher risk of development of metabolic and cardiovascular disorders, including hypertension (HT). To date, there have been no studies supporting the relationship between epicardial adipose tissue (EAT) expression of adiponectin and HT. We collected samples of EAT from 116 patients undergoing elective cardiac surgery, mostly for coronary artery bypass grafting (n=54), valve surgery (n=49) or both (n=12). Samples of subcutaneous adipose tissue (SAT) were harvested from 85 patients. After RNA isolation, the expression of adiponectin was analysed by real-time retrotranscriptase (RT)-PCR. Baseline clinical data were obtained from medical records. The diagnosis of HT was established mostly by the patients’ general physicians following current guidelines. We included 84 hypertensive and 32 non-hypertensive patients. Mean (±s.d.) age was 70.3±7.9 years. EAT expression levels of adiponectin were lower in hypertensives (14.0±3.6 vs 15.3±3.6 arbitrary units (a.u.), P=0.06). This difference was statistically significant (odds ratio (OR) 0.828 per a.u., P=0.020) after adjustment for age, gender, body mass index, diabetes mellitus, heart failure, coronary artery disease (CAD), total cholesterol and triglyceride levels. However, SAT adiponectin mRNA levels were similar in hypertensive and non-hypertensive patients (15.3±4.2 vs 15.3±5.0 a.u., P>0.99). Adjustment for potential confounding factors hardly altered this result. Our findings indicate that EAT expression of adiponectin may be associated with HT status independently of CAD or other comorbidities, whereas SAT expression does not. These results support the hypothesis that EAT is actively implicated in global cardiovascular risk, describing its association with HT.

Gender Differences in Adiponectin and Leptin Expression in Epicardial and Subcutaneous Adipose Tissue. Findings in Patients Undergoing Cardiac Surgery

INTRODUCTION AND OBJECTIVES The high level of metabolic activity present in visceral adipose tissue is associated with the development of atherothrombosis. Subcutaneous adipose tissue secretes larger quantities of the adipocytokines leptin and adiponectin than visceral adipose tissue. Epicardial adipose tissue secretes inflammatory mediators, and the resulting proinflammatory activity is greater than that associated with subcutaneous adipose tissue. The aim of this study was to compare adiponectin and leptin expression in human epicardial and subcutaneous adipose tissue. METHODS Samples of both epicardial and subcutaneous adipose tissue were taken from 46 patients who were undergoing heart surgery, coronary artery bypass surgery, or aortic or mitral valve replacement. Levels of protein and gene expression of leptin and adiponectin were assessed immunohistochemically and by RT-PCR, respectively. Levels of mRNA expression in the 2 adipose tissue types were compared by real-time quantitative PCR. RESULTS Significant differences were found between adiponectin and leptin mRNA expression in epicardial and subcutaneous adipose tissue, with epicardial adipose tissue exhibiting lower levels of adiponectin and leptin expression. Moreover, adiponectin and leptin mRNA expression in epicardial adipose tissue was higher in women than men. CONCLUSIONS Adiponectin and leptin expression is lower in epicardial than in subcutaneous adipose tissue. Moreover, there are differences in adiponectin and leptin expression in epicardial adipose tissue between women and men.

Relationship between epicardial adipose tissue adipocyte size and MCP-1 expression

Adipocyte size has been associated to increase in inflammatory cytokines expression that can be related to the cardiovascular risk of obesity. Epicardial adipose tissue (EAT) was discovered to play a key role in cardiovascular diseases by producing several inflammatory adipokines. We sought to study whether EAT and subcutaneous adipose tissue (SAT) mean adipocyte sizes are related to the expression of adipokines in patients with cardiovascular diseases. We collected EAT, SAT and blood samples from 22 patients aged 70.9 (s.d. 10.3) undergoing heart surgery. Monocyte chemoattractant protein (MCP)-1, interleukin (IL)-10 and tumor necrosis factor (TNF)-alpha were analyzed by real time RT-PCR, ELISA or immunohistochemistry. Hematoxylin-eosin staining was used for adipocyte area calculations. Adipocyte size is negatively correlated to MCP-1 expression (r=-0.475; p=0.034) in EAT and positively correlated in SAT (r=0.438; p=0.047). These trends persisted after stratification for sex and coronary artery disease (CAD), but only the relationship between EAT MCP-1 and adipocyte size reached statistical significance in the larger group of men with CAD. We have observed that SAT adipocyte size is correlated to BMI (r=0.601; p=0.003); whereas only a non-statistically significant trend was observed in EAT. IL-10 and TNF-alpha expression were not associated to adipocyte size in EAT nor SAT. Secondarily, we found that EAT IL-10 expression is higher in patients with CAD. These results suggest that adipocyte size is a negative determinant of MCP-1 expression in EAT and a positive determinant in SAT. These data might partly explain the different implications of EAT and SAT in cardiovascular diseases.

Leptin promotes the tyrosine phosphorylation of SHC proteins and SHC association with GRB2.

The identification and characterization of proteins that become tyrosine phosphorylated in response to growth factor stimulation is critical for furthering our understanding of the signal transduction pathways involved in the regulation of cell proliferation, differentiation as well as metabolic activities. In this report, we demonstrate for the first time, that leptin is able to induce the tyrosine phosphorylation of the SH(2) containing protein SHC. These studies have been carried out on a human embryonic cell line (HEK 293) transfected with the cDNA encoding for the long form of the leptin receptor and stably expressing the receptor itself. We also shown that upon tyrosine phosphorylation, SHC associated with the adaptor protein, Grb(2). The formation of this complex may directly link tyrosine phosphorylation events to Ras activation and may be a critical step in proliferation and/or differentiation of cells. In conclusion, these results indicate that leptin receptor, after binding the ligand, activates several pathways for signal transduction that might lead to mitogenic effect.

Coronary artery disease is associated with higher epicardial Retinol‐binding protein 4 (RBP4) and lower glucose transporter (GLUT) 4 levels in epicardial and subcutaneous adipose tissue

Objective  Retinol‐binding protein 4 (RBP4), produced by adipocytes and hepatocytes, contributes to an unfavourable lipid profile and insulin resistance, which can contribute to the development of coronary artery disease (CAD). Recently, several studies have shown that epicardial adipose tissue (EAT) differs from subcutaneous adipose tissue (SAT) and plays a role on the physiopathology of CAD because of its proximity to the coronary arteries. We aimed to study the expression and secretion levels of RBP4 in both fat tissues and explore its possible association with CAD.

Lower epicardial adipose tissue adiponectin in patients with metabolic syndrome

BACKGROUND Adiponectin is an anti-atherogenic insulin-sensitizer hormone whose plasma concentration is lower in patients with metabolic syndrome (MS). Visceral adiposity, including epicardial adipose tissue (EAT), is closely related to the development of MS and coronary artery disease (CAD). We sought to study whether EAT and subcutaneous adipose tissue (SAT) adiponectin mRNA levels are similar in patients with and without MS. METHODS EAT, SAT and blood samples were collected from patients undergoing elective cardiac surgery, for revascularization (n=19) or other procedures (n=27). Plasma adiponectin was measured using ELISA. mRNA was purified and adiponectin mRNA quantified by real time RT-PCR. RESULTS Mean (SD) age was 71.6 (9.6) years. Patients who met Adult Treatment Panel III MS criteria (n=29) presented lower plasma adiponectin concentrations (11.2 (7.4) vs. 19.6 (8.4) mg/l, P=0.004), lower EAT adiponectin mRNA (12.7 (3.0) vs. 15.1 (3.7) a.u., P=0.029) and similar SAT adiponectin mRNA levels (13.7 (4.2) vs. 15.6 (5.7) a.u., P=0.25) than those without MS. After adjusting for age, sex, CAD and heart failure, the association with MS remained statistically significant for plasma adiponectin (OR 0.862 (0.762-0.974)), was of borderline significance for EAT adiponectin mRNA (OR 0.796 (0.630-1.005)) and not significant for SAT adiponectin mRNA (OR 0.958 (0.818-1.122)). Patients in the lower quartiles of EAT adiponectin mRNA and plasma adiponectin presented a higher mean of components of the MS. CONCLUSIONS Subjects with MS present lower EAT adiponectin mRNA levels than those without MS, whereas SAT adiponectin mRNA levels do not seem to differ between both groups. EAT might be the link between MS and its atherothrombotic cardiac complications.