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

Faecal Calprotectin and a Twenty-Four-Parameter Questionnaire in Autistic Children with Gastrointestinal Symptoms

This study investigated potential correlation between the inflammatory marker, Calprotectin, and a S.O.S questionnaire from forty-nine Autistic children. Symptom and behavioral questionnaires were completed contemporaneously with stool sample collection. Mixed Model data analysis showed strong correlation between some questionnaire parameters and Calprotectin. ‘Need for a fixed routine’ was highly significantly correlated with Calprotectin (\u1d45d<0.00009) with Multivariate Coefficient of 3.227, whilst paradoxically ‘constipation’ indicated significant change (\u1d45d<0.02) with negative Multivariate Coefficient (-1.584). The negative ‘constipation’ appears to associate with the positive ‘need for a fixed routine’ indicating possibility of reciprocal, independent prediction of gastrointestinal inflammation. Results suggest that ‘need for a fixed routine’ and ‘constipation’ be included in a screening questionnaire as independent predictors of bowel dysfunction in these children

Acute Alcohol Tissue Damage: Protective Properties of Betaine

Teenage binge drinking is a major health issue; however, there is a paucity of data on liver injury. Herein, we investigated how acute ethanol affects juvenile hepatic cells through changes in oxidative stress, apoptosis, and liver function, as well as the ability of betaine, which can replen-ish the antioxidant glutathione and mitigate oxidative injury. Juvenile male Wistar rats were given either water or betaine (2% w/v) for 6 days and treated with either saline 0.15 mol/L NaCl or ethanol (75 mmol/kg bodyweight). After 24 h, liver enzymes, oxidative damage, apoptosis, and parameters of antioxidant enzyme activity were examined. Acute ethanol increased hepatic enzymes (99%, P < 0.05). Total protein and albumin levels were reduced by 14 and 18% (P < 0.001), respectively, which was prevented by betaine treatment. Cytosolic cytochrome c increased by 59% (P < 0.05), corresponding to a decrease in mitochondrial cytochrome c content, which was ameliorated with betaine. Cytosolic glutathione peroxidase was reduced with alcohol (P < 0.05) and was prevented with betaine. Subtle changes were observed in catalase, superoxide dismutase, glutathione reductase, and complex I activity after ethanol treatment. In summary, whilst juvenile animals appear to have higher basal levels of antioxidant enzymes, betaine conferred some protection against alcohol-induced oxidative stress

933-93 The Regression of Left Ventricular Myofibrillary Proteins by ACE-inhibition (Lisinopril) is Associated with an Increase in Protein Synthetic Rates in Vivo

One of the most serious complications of systemic hypertension is myocardial tissue damage, including left ventricular hypertrophy, as a consequence of increased protein synthesis. However, the modulating role of translational events in the hypertrophy-regression transitionis poorly understood, especially where therapeutic regimes have been employed. These events were investigated in vivoin a genetic model of hypertension, namely in the spontaneously hypertensive rat (SHR); comparative responses were investigated in normotensive Wistar Kyoto rats (WKY). Rats were used at 4 months of age and treated with either the ACE-inhibitor lisinopril (5mg/kg/day) in tap water or plain tap water (controls). The groups were assigned as follows: WKY-CON, normotensive controls; WKY-LIS, normotensives plus lisinopril; SHR-CON, hypertensive controls; SHR-LIS, hypertensives plus lisinopril. Fractional rates of protein synthesis (ksdefined as the percentage of the myofibrillary protein pool renewed each day; %/day) were measured in vivowith the flooding dose technique using L-[4-3H]phenylalanine. Left ventricular myofibrillary proteins were extracted by differential solubility and high-speed centrifugation techniques; purity was assessed with SDS-PAGE. After 8 weeks treatment the myofibrillary protein contents (mg per region) in normotensive rats were as follows (all data as mean±SEMS, n=6–9): WKY-CON, 45±1 mg; WKY–LIS, 36±1 (p&lt;0.001). In the hypertensive group regression of contractile protein content occurred; i.e., SHR-CON, 52±3mg; SHR-L1S, 38±1mg (p&lt;0.001). Corresponding ksvalues were: WKY-CON, 8.3±0.2 %/d; WKY–LIS, 8.2±0.3 %/d (NS). In the SHR-group ksvalues were: SHR-CON, 8.2±0.3%/d; SHR-L1S, 9.2±0.3mg (p&lt;0.025)ConclusionACE-induced regression of contractile protein composition in hypertension is associated with an increase in rates of translatio

Baseline and recurrent exposure to the standard dose of artemisinin-based combination therapies (ACTs) induces oxidative stress and liver damage in mice (BALB/c)

Background In malaria-endemic countries, repeated intake of artemisinin-based combination therapies (ACTs) is rampant and driven by drug resistance, improper usage, and easy accessibility. Stress effects and potential liver tox- icity due to the frequent therapeutic use of ACTs have not been extensively studied. Here, we investigated the effects of repeated treatment with standard doses of the commonly used ACTs artemether/lumefantrine (A/L) and artesu- nate-amodiaquine (A/A) on oxidative stress and liver function markers in male mice (BALB/c). Methods Forty Five mice were divided into three groups: control, A/L, and A/A. The drugs were administered three days in a row per week, and the regimen was repeated every two weeks for a total of six cycles. The levels of oxidative stress and liver function markers were measured in both plasma and liver tissue after initial (baseline) and repeated exposures for the second, third, and sixth cycles. Results Exposure to A/L or A/A caused a significant (p < 0.001) increase in plasma malondialdehyde (MDA) lev- els after the first and repeated exposure periods. However, Hepatic MDA levels increased significantly (p < 0.01) only after the sixth exposure to A/A. Following either single or repeated exposure to A/L or A/A, plasma and liver glutathione peroxidase (GPx) and catalase (CAT) activities, plasma aspartate and alanine transaminase, alkaline phos- phatase activity, and bilirubin levels increased, whereas total plasma protein levels decreased significantly (p < 0.001). Varying degrees of hepatocyte degeneration and blood vessel congestion were observed in liver tissues after a single or repeated treatment period. Conclusion Irrespective of single or repeated exposure to therapeutic doses of A/L or A/A, plasma oxidative stress and liver damage were observed. However, long-term repeated A/A exposure can led to hepatic stress. Compensa- tory processes involving GPx and CAT activities may help reduce the observed stress

Laser scribing optimization of RF magnetron sputtered molybdenum thin films

The optimization process of laser scribing of back contacts is carried out by varying different parameters of laser and thickness of Molybdenum (Mo) thin-films. Mo thin films were deposited by RF magnetron sputtering on the organically cleaned soda lime glass substrate. The thickness of Mo was in the range of 60 nm to 800 nm. For the scribing process the laser power and the laser pulse frequency were varied. Different thickness of Mo shows the different scribe behavior. The optimized process provides a successful isolative laser scribing, having a minimum scribe line width, of Mo layer on glass substrate without any presence of walls, ridges, or collars in scribed areas. When you are citing the document, use the following link http://essuir.sumdu.edu.ua/handle/123456789/929

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

Relationship of postpartum interval to estrus, body condition score, milk yield and blood biochemical parameters in Surti buffaloes (Bubalus bubalis)

The aim of the present investigation was to find out the relationship among postpartum interval to estrus, body condition score, milk yield and blood biochemical parameters of Surti buffaloes (Bubalus bubalis). The study was conducted on sixteen clinically healthy Surti buffaloes (parity 1-7) with normal parturition. These animals were divided into two groups on the basis of their postpartum interval to estrus (PPIE). Group 1 animals had PPIE ? 50 days whereas group 2 had PPIE &gt; 50 days. Body condition score (BCS), milk yield and Blood samples were collected by jugular venipuncture on days starting from 6th day after calving thereafter at fortnight interval till 90th day postpartum. Blood serum parameters such as glucose, total protein, blood urea, creatinine, cholesterol, triglyceride, progesterone and estrogen were measured. Perusal of data revealed that animals having higher BCS on the day of estrus had significantly (P?0.05) shorter PPIE. There was non-significant effect of daily and cumulative 100 days milk yield on PPIE. Serum concentration of glucose and creatinine was significantly (P?0.05) higher for group 1 animals at most of the stages. There was non-significant difference between serum concentration of total protein, blood urea nitrogen and cholesterol between both the groups. Progesterone and Estradiol-17 ? concentrations were significantly (P?0.05) higher in group 1 animals than group 2 animals at different stages of this study