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Key Words: FABP4 Ⅲ heart failure Ⅲ adipocytes Ⅲ metabolic syndrome O besity is a major risk factor in the development of the metabolic syndrome and cardiovascular diseases and seems to be directly related to heart failure independently of other risk factors. 1 Indeed, a direct relationship between elevated body mass index and increased risk for heart failure has been demonstrated, without evidence of a threshold. 2 Several potential mechanisms are under discussion to explain this correlation, including hemodynamic changes with cardiac overload and left ventricular remodeling and lipid accumulation into the myocardium, leading to lipoapoptosis in cardiomyocytes. 3 These mechanisms, however, do not fully explain the development of heart dysfunction in obese subjects.Adipocytes are known to produce and release a wide variety of bioactive molecules into the bloodstream. 4 Based on these data, we have recently investigated whether secretory products from human adipocytes affect cardiac contractile function in an in vitro system of isolated rat cardiomyocytes. We have demonstrated that mature human adipocytes release substances that strongly and acutely suppress the contraction of cardiomyocytes by attenuating intracellular Ca 2ϩ levels. 5 Our previous findings have revealed a hitherto unknown acute depressant effect of adipocyte-derived factors on cardiac contraction, suggesting a new direct role of adipose tissue in the pathogenesis of myocardium dysfunction.Based on this initial work, we have further characterized cardiodepressant activity by fractionating adipocyte secretory products according to molecular weight and proteomic analysis, identifying adipocyte fatty acid-binding protein (FABP4) as the active agent. Fatty acid-binding proteins (FABPs) are members of a highly conserved family of cytosolic proteins with a molecular mass of 14 to 15 kDa found in different cell types, showing a high affinity for long-chain fatty acids and other hydrophobic ligands. 6 FABP4 is predominantly expressed in adipose tissue, and accounts for Ϸ1% of total cytosolic protein in human adipose tissue. 7 Cytoplasmic FABP4 is involved in trafficking intra-

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Lipopolysaccharides (LPS), the major components of the wall of gram-negative bacteria, trigger powerful defensive responses in the airways via mechanisms thought to rely solely on the Toll-like receptor 4 (TLR4) immune pathway. Here we show that airway epithelial cells display an increase in intracellular Ca2+ concentration within seconds of LPS application. This response occurs in a TLR4-independent manner, via activation of the transient receptor potential vanilloid 4 cation channel (TRPV4). We found that TRPV4 mediates immediate LPS-induced increases in ciliary beat frequency and the production of bactericidal nitric oxide. Upon LPS challenge TRPV4-deficient mice display exacerbated ventilatory changes and recruitment of polymorphonuclear leukocytes into the airways. We conclude that LPS-induced activation of TRPV4 triggers signaling mechanisms that operate faster and independently from the canonical TLR4 immune pathway, leading to immediate protective responses such as direct antimicrobial action, increase in airway clearance, and the regulation of the inflammatory innate immune reaction.

Long-term left coronary occlusion induces significant alterations in both Ca2+ and K+ currents that occur with similar amplitude in both ventricles. They include a marked reduction in Ito amplitude as well as a slowing of ICaL inactivation. Both factors could contribute to the disturbances in cellular electrical behaviour and the occurrence of arrhythmias in the post-myocardial infarcted heart.

Defective L-type Ca2+ channel (I(CaL)) regulation is one major cause for contractile dysfunction in the heart. The I(CaL) is enhanced by sympathetic nervous stimulation: via the activation of beta-adrenergic receptors, PKA phosphorylates the alpha1C(Ca(V)1.2)- and beta2-channel subunits and ahnak, an associated 5643-amino acid (aa) protein. In this study, we examined the role of a naturally occurring, genetic variant Ile5236Thr-ahnak on I(CaL). Binding experiments with ahnak fragments (wild-type, Ile5236Thr mutated) and patch clamp recordings revealed that Ile5236Thr-ahnak critically affected both beta2 subunit interaction and I(CaL) regulation. Binding affinity between ahnak-C1 (aa 4646-5288) and beta2 subunit decreased by approximately 50% after PKA phosphorylation or in the presence of Ile5236Thr-ahnak peptide. On native cardiomyocytes, intracellular application of this mutated ahnak peptide mimicked the PKA-effects on I(CaL) increasing the amplitude by approximately 60% and slowing its inactivation together with a leftward shift of its voltage dependency. Both mutated Ile5236Thr-peptide and Ile5236Thr-fragment (aa 5215-5288) prevented specifically the further up-regulation of I(CaL) by isoprenaline. Hence, we suggest the ahnak-C1 domain serves as physiological brake on I(CaL). Relief from this inhibition is proposed as common pathway used by sympathetic signaling and Ile5236Thr-ahnak fragments to increase I(CaL). This genetic ahnak variant might cause individual differences in I(CaL) regulation upon physiological challenges or therapeutic interventions.

The properties of the low threshold Ca current (/CAT) in bullfrog (Rana catesbeiana) isolated atrial cardiomyocytes were studied using the whole-cell recording patch-damp technique and compared with those of the high threshold Ca current (IC~L). In 91% of atrial cells we observed both lc~-r and IC~L when collagenase and trypsin were used to dissociate the cells. But when pronase was used, only 30% of the cells exhibited loT. Icax was never found in ventricular cells. /CaT could be investigated more easily when ICaL was inhibited by Cd ions (50 p.M). Its kinetics were unchanged by substituting Ba for Ca, or in the presence of high concentrations of Ba. Both lcax and IC~L exhibited reduced inactivation after high depolarizing prepulses, lcax was found to be sensitive to dihydropyridines: 1 ~,M nifedipine decreased this current while 1 ~M BAY K 8644 increased it; this occurred without significant variations in the steady-state inactivation curve. /CaT was more sensitive than IcaL to cq-adrenergic and P2-purinergic stimulations, while ICaL was more sensitive to [3-adrenergic stimulation. Isoproterenol was still able to increase Icax in the presence of high intracellular cAMP. Both currents were increased by 1 ~M ouabain (although ICaL only transiently) and decreased by 10 I~M ouabain. It is concluded that the two types of Ca channels can be observed in bullfrog atrial cells and that they are specifically altered by pharmacological agents and neuromediators. This may have implications for cardiac behavior.

BackgroundSilica nanoparticles (SiNPs) have numerous beneficial properties and are extensively used in cosmetics and food industries as anti-caking, densifying and hydrophobic agents. However, the increasing exposure levels experienced by the general population and the ability of SiNPs to penetrate cells and tissues have raised concerns about possible toxic effects of this material. Although SiNPs are known to affect the function of the airway epithelium, the molecular targets of these particles remain largely unknown. Given that SiNPs interact with the plasma membrane of epithelial cells we hypothesized that they may affect the function of Transient Receptor Potential Vanilloid 4 (TRPV4), a cation-permeable channel that regulates epithelial barrier function. The main aims of this study were to evaluate the effects of SiNPs on the activation of TRPV4 and to determine whether these alter the positive modulatory action of this channel on the ciliary beat frequency in airway epithelial cells.ResultsUsing fluorometric measurements of intracellular Ca2+ concentration ([Ca2+]i) we found that SiNPs inhibit activation of TRPV4 by the synthetic agonist GSK1016790A in cultured human airway epithelial cells 16HBE and in primary cultured mouse tracheobronchial epithelial cells. Inhibition of TRPV4 by SiNPs was confirmed in intracellular Ca2+ imaging and whole-cell patch-clamp experiments performed in HEK293T cells over-expressing this channel. In addition to these effects, SiNPs were found to induce a significant increase in basal [Ca2+]i, but in a TRPV4-independent manner. SiNPs enhanced the activation of the capsaicin receptor TRPV1, demonstrating that these particles have a specific inhibitory action on TRPV4 activation. Finally, we found that SiNPs abrogate the increase in ciliary beat frequency induced by TRPV4 activation in mouse airway epithelial cells.ConclusionsOur results show that SiNPs inhibit TRPV4 activation, and that this effect may impair the positive modulatory action of the stimulation of this channel on the ciliary function in airway epithelial cells. These findings unveil the cation channel TRPV4 as a primary molecular target of SiNPs.