Expression and physiological role of the novel adipokine nesfatin-1 in cardiomyocytes

Abstract

Purpose: Nesfatin-1 is a novel adipokine involved in the control of food intake and energy metabolism which shows anti-inflammatory properties. The role of this adipokine in cardiovascular physiology is unknown. In previous studies we determined that nesfatin-1 is expressed in human, rat and mouse heart. Our aim now is to study the effect of this adipokine in cardiomyocytes and the possible regulation of nesfatin-1 cardiac synthesis by diet and inflammatory mediators. Methods: Real-time PCR was used to determine nesfatin-1 mRNA levels in cultured neonatal cardiomyocytes of Sprague Dawley rats treated with TNF-a, dexamethasone and insulin. In heart tissue of rats fed with high fat diet for 16 weeks, we used real-time PCR to determine nesfatin-1 cardiac mRNA levels and an ELISA to determine nesfatin-1 plasma levels. Cardiomyocytes were treated with nesfatin-1 and confocal microscopy was used to study the glucose transporter Glut-4 movilization. Finally, western blot was used to identify possible transducing signalling molecules (Erk 1 2 , AMPK and AKT) after nesfatin-1 treatment in cardiomyocytes. Results: Cardiomyocytes treatment with 0.1-20 ng/ml TNF-a for 6-48 h induces an increase of nesfatin-1 mRNA levels with a maximum stimulatory effect at 20 ng/ml for 24 h (p=0.0159; Fold-Change (FC)=1.16, n=5). Treatment with 0.1-100 nM dexamethasone for 6-48 h also increases of nesfatin-1 mRNA levels with a maximum stimulatory effect at 100 nM for 24 h (p=0.0079; FC=2.457, n=5). On the other hand, 0.1-100 nM insulin treatment for 6-48 h decreases nesfatin-1 mRNA levels with a maximum stimulatory effect at 100 nM for 24 h (p=0.0159; FC= -0.6962, n=5). Treatment of cardiomyocytes with 10-1000 nM nesfatin-1 for 5-30 minutes induces Glut-4 movilization from the cytoplasm to the plasma membrane (p=0.0007; FC=1.125, n=216 cells) and an Erk 1 2 phosphorilation (p=0.0156; FC=3.133, n=7) with a maximum stimulatory effect at 1000 nM for 10 minutes. In rats fed with high fat diet for 16 weeks nesfatin-1 cardiac mRNA levels are higher compared with control (p=0.0111, FC =2.201, n=7) and there is a positive correlation with body weight (Spearman’s rho (Rho)= 0.715, Sig. =0.004, n=14) and the fat percentage (Rho= 0.692, Sig. =0.006, n=14). There is also a positive correlation between plasma nesfatin-1 levels and body weight (Rho= 0.582, Sig. =0.037, n=13). Conclusions: In cardiomyocytes nesfatin-1 regulates glucose homeostasis. Cardiac levels of this adipokine are modified by inflammatory and metabolic status. Our work provides the first evidences about a potential role of nesfatin-1 in a paracrine/autocrine system at cardiac level.