Hepcidin secretion was not directly proportional to intracellular iron-loading in recombinant-TfR1 HepG2 cells: short communication

Hepcidin is the master regulator of systemic iron homeostasis and its dysregulation is observed in several chronic liver diseases. Unlike the extracellular iron-sensing mechanisms, the intracellular iron-sensing mechanisms in the hepatocytes that lead to hepcidin induction and secretion are incompletely understood. Here, we aimed to understand the direct role of intracellular iron-loading on hepcidin mRNA and peptide secretion using our previously characterised recombinant HepG2 cells that over-express the cell-surface iron-importer protein transferrin receptor-1. Gene expression of hepcidin (HAMP) was determined by real-time PCR. Intracellular iron levels and secreted hepcidin peptide levels were measured by ferrozine assay and immunoassay, respectively. These measurements were compared in the recombinant and wild-type HepG2 cells under basal conditions at 30 min, 2 h, 4 h and 24 h. Data showed that in the recombinant cells, intracellular iron content was higher than wild-type cells at 30 min (3.1-fold, p<0.01), 2 h (4.6-fold, p<0.01), 4 h (4.6-fold, p<0.01) and 24 h (1.9-fold, p<0.01). Hepcidin (HAMP) mRNA expression was higher than wild-type cells at 30 min (5.9-fold; p=0.05) and 24 h (6.1-fold; p<0.03), but at 4 h, the expression was lower than that in wild-type cells (p<0.05). However, hepcidin secretion levels in the recombinant cells were similar to those in wild-type cells at all time-points, except at 4 h, when the level was lower than wild-type cells (p<0.01). High intracellular iron in recombinant HepG2 cells did not proportionally increase hepcidin peptide secretion. This suggests a limited role of elevated intracellular iron in hepcidin secretio

HFE mRNA expression is responsive to intracellular and extracellular iron loading:short communication

In liver hepatocytes, the HFE gene regulates cellular and systemic iron homeostasis by modulating cellular iron-uptake and producing the iron-hormone hepcidin in response to systemic iron elevation. However, the mechanism of iron-sensing in hepatocytes remain enigmatic. Therefore, to study the effect of iron on HFE and hepcidin (HAMP) expressions under distinct extracellular and intracellular iron-loading, we examined the effect of holotransferrin treatment (1, 2, 5 and 8 g/L for 6 h) on intracellular iron levels, and mRNA expressions of HFE and HAMP in wild-type HepG2 and previously characterized iron-loaded recombinant-TfR1 HepG2 cells. Gene expression was analyzed by real-time PCR and intracellular iron was measured by ferrozine assay. Data showed that in the wild-type cells, where intracellular iron content remained unchanged, HFE expression remained unaltered at low holotransferrin treatments but was upregulated upon 5 g/L (p < 0.04) and 8 g/L (p = 0.05) treatments. HAMP expression showed alternating elevations and increased upon 1 g/L (p < 0.05) and 5 g/L (p < 0.05). However, in the recombinant cells that showed higher intracellular iron levels than wild-type cells, HFE and HAMP expressions were elevated only at low 1 g/L treatment (p < 0.03) and were repressed at 2 g/L treatment (p < 0.03). Under holotransferrin-untreated conditions, the iron-loaded recombinant cells showed higher expressions of HFE (p < 0.03) and HAMP (p = 0.05) than wild-type cells. HFE mRNA was independently elevated by extracellular and intracellular iron-excess. Thus, it may be involved in sensing both, extracellular and intracellular iron. Repression of HAMP expression under simultaneous intracellular and extracellular iron-loading resembles non-hereditary iron-excess pathologies

High omega arachidonic acid/docosahexaenoic acid ratio induces mitochondrial dysfunction and altered lipid metabolism in human hepatoma cells

Background Non-alcoholic fatty liver disease (NAFLD) is a common cause of liver disease worldwide and is a growing epidemic. A high ratio of omega-6 fatty acids to omega-3 fatty acids in the diet has been implicated in the development of NAFLD. However, the inflicted cellular pathology remains unknown. A high ratio may promote lipogenic pathways and contribute to ROS-mediated damage, perhaps leading to mitochondrial dysfunction. Therefore, these parameters were investigated to understand their contribution to NAFLD development. Aim To examine the effect of increasing ratios of omega-6:3 fatty acids on mitochondrial function and lipid metabolism mediators. Methods HepG2-derived VL-17A cells were treated with normal (1:1, 4:1) and high (15:1, 25:1) ratios of omega-6: omega-3 fatty acids (arachidonic (AA): docosahexaenoic (DHA)) at various time points. Mitochondrial activity and function was examined via MTT assay and Seahorse XF24 analyzer, respectively. Triglyceride accumulation was determined by using EnzyChrom™ and levels of ROS were measured by fluorescence intensity. Protein expression of the mediators of lipogenic, lipolytic and endocannabinoid pathways was assessed by Western blotting. Results High AA:DHA ratio decreased mitochondrial activity (p<0.01; upto 80%) and promoted intracellular triglyceride accumulation (p<0.05; 40-70%). Mechanistically, it altered the mediators of lipid metabolism; increased the expression of stearoyl-CoA desaturase (p<0.05; 22-35%), decreased the expression of peroxisome proliferator-activated receptor-alpha (p<0.05; 30-40%) and increased the expression of cannabinoid receptor 1 (p<0.05; 31%). Furthermore, the high ratio increased ROS production (p<0.01; 74-115%) and reduced mitochondrial respiratory functions such as basal and maximal respiration, ATP production, spare respiratory capacity and proton leak (p<0.01; 35-68%). Conclusion High AA:DHA ratio induced triglyceride accumulation, increased oxidative stress and disrupted mitochondrial functions. Stimulation of lipogenic and steroidal transcription factors may partly mediate these effects and contribute to NAFLD development

On Nori's Fundamental Group Scheme

We determine the quotient category which is the representation category of the kernel of the homomorphism from Nori's fundamental group scheme to its \'etale and local parts. Pierre Deligne pointed out an error in the first version of this article. We profoundly thank him, in particular for sending us his enlightning example reproduced in Remark 2.4 2).Comment: 29 page

Iron enhances hepatic fibrogenesis and activates TGF-β signaling in murine hepatic stellate cells

Introduction Although excess iron induces oxidative stress in the liver, it is unclear whether it directly activates the hepatic stellate cells (HSC). Methods We evaluated effects of excess iron on fibrogenesis and TGF-β signaling in murine HSC. Cells were treated with holotransferrin (0.005–5 g/L) for 24 h, with or without the iron chelator deferoxamine (10 µM). Gene expressions (α-SMA, Col1-α1, Serpine-1, TGF-β, Hif1-α, Tfrc and Slc40a1) were analyzed by qRT-PCR, while TfR1, ferroportin, ferritin, vimentin, collagen, TGF-β RII and phospho-Smad2 proteins were evaluated by immunofluorescence, western blot and ELISA. Results HSC express the iron-uptake protein TfR1, and the iron-export protein ferroportin. Holotransferrin up-regulated TfR1 expression by 1.8-fold (p<0.03) and ferritin accumulation (iron storage) by 2-fold (p<0.01), and activated HSC with 2-fold elevations (p<0.03) in α-SMA mRNA and collagen secretion, and a 1.6-fold increase (p<0.01) in vimentin protein. Moreover, holotransferrin activated the TGF-β pathway with TGF-β mRNA elevated 1.6-fold (p=0.05), and protein levels of TGF-β RII and phospho-Smad2 increased by 1.8-fold (p<0.01) and 1.6-fold (p<0.01), respectively. By contrast, iron chelation decreased ferritin levels by 30% (p<0.03), inhibited collagen secretion by 60% (p<0.01), repressed fibrogenic genes α-SMA (0.2-fold; p<0.05) and TGF-β (0.4-fold; p<0.01), and reduced levels of TGF-β RII and phospho-Smad2 proteins. Conclusion HSC express iron transport proteins. Holotransferrin (iron) activates HSC fibrogenesis and the TGF-β pathway, while iron depletion by chelation reverses this, suggesting that this could be a useful adjunct therapy for patients with fibrosis. Further studies in primary human HSC and animal models are necessary to confirm this

Public support in England for a total ban on the sale of tobacco products

Background This study aimed to determine the level of support for a sales ban on tobacco in England to provide a benchmark against which any changes over time can be assessed.Methods 8735 people from England who participated in one of five monthly cross-sectional household surveys in 2008 were asked to indicate whether they would support the statement that 'the government should work towards banning the sale of tobacco completely within the next 10 years'. In addition, sociodemographic and smoking characteristics were assessed.Results A substantial proportion of the total sample (44.5%; 95% CI 43.5% to 45.6%) would support a move towards a complete ban. While never smokers (OR 2.02; 95% CI 1.82 to 2.25) and ex-smokers (OR 1.41; 95% CI 1.21 to 1.65) were more likely to support this idea, even among current smokers, a third would favour moving towards a sales ban of tobacco products. Adjusting for other background characteristics, younger, female participants, those living in London and those from lower socioeconomic groups were most likely to support a ban. Among smokers, a higher cigarette consumption, smoking enjoyment and contentment with being a smoker were associated with opposition to a ban, while feeling uncomfortable being a smoker, wanting to be a non-smoker and being worried about future health consequences of smoking were associated with support for a ban.Conclusion Support for movement towards a ban on the sale of tobacco is higher than might be imagined. It is conceivable that as smoking prevalence falls further and smoking becomes more socially unacceptable, support might grow to a point where such a policy could become feasible

Stability conditions and positivity of invariants of fibrations

We study three methods that prove the positivity of a natural numerical invariant associated to $1-$parameter families of polarized varieties. All these methods involve different stability conditions. In dimension 2 we prove that there is a natural connection between them, related to a yet another stability condition, the linear stability. Finally we make some speculations and prove new results in higher dimension.Comment: Final version, to appear in the Springer volume dedicated to Klaus Hulek on the occasion of his 60-th birthda

From Vicious Walkers to TASEP

We propose a model of semi-vicious walkers, which interpolates between the totally asymmetric simple exclusion process and the vicious walkers model, having the two as limiting cases. For this model we calculate the asymptotics of the survival probability for $m$ particles and obtain a scaling function, which describes the transition from one limiting case to another. Then, we use a fluctuation-dissipation relation allowing us to reinterpret the result as the particle current generating function in the totally asymmetric simple exclusion process. Thus we obtain the particle current distribution asymptotically in the large time limit as the number of particles is fixed. The results apply to the large deviation scale as well as to the diffusive scale. In the latter we obtain a new universal distribution, which has a skew non-Gaussian form. For $m$ particles its asymptotic behavior is shown to be $e^{-\frac{y^{2}}{2m^{2}}}$ as $y\to -\infty$ and $e^{-\frac{y^{2}}{2m}}y^{-\frac{m(m-1)}{2}}$ as $y\to \infty$.Comment: 37 pages, 4 figures, Corrected reference

Absorbing-state phase transitions in fixed-energy sandpiles

We study sandpile models as closed systems, with conserved energy density $\zeta$ playing the role of an external parameter. The critical energy density, $\zeta_c$, marks a nonequilibrium phase transition between active and absorbing states. Several fixed-energy sandpiles are studied in extensive simulations of stationary and transient properties, as well as the dynamics of roughening in an interface-height representation. Our primary goal is to identify the universality classes of such models, in hopes of assessing the validity of two recently proposed approaches to sandpiles: a phenomenological continuum Langevin description with absorbing states, and a mapping to driven interface dynamics in random media. Our results strongly suggest that there are at least three distinct universality classes for sandpiles.Comment: 41 pages, 23 figure

Search for direct production of charginos and neutralinos in events with three leptons and missing transverse momentum in √s = 7 TeV pp collisions with the ATLAS detector

A search for the direct production of charginos and neutralinos in final states with three electrons or muons and missing transverse momentum is presented. The analysis is based on 4.7 fb−1 of proton–proton collision data delivered by the Large Hadron Collider and recorded with the ATLAS detector. Observations are consistent with Standard Model expectations in three signal regions that are either depleted or enriched in Z-boson decays. Upper limits at 95% confidence level are set in R-parity conserving phenomenological minimal supersymmetric models and in simplified models, significantly extending previous results