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Background/Aims: Accumulating evidences have demonstrated that long noncoding RNAs (lncRNA) play important roles in hepatic lipid metabolism in mammals. However, no systematic screening of the potential lncRNAs in the livers of laying hens has been performed, and few studies have been reported concerning the effects of the lncRNAs on lipid metabolism in the livers of chickens during egg-laying period. The purpose of this study was to compare the difference in lncRNA expression in the livers of pre-laying and peak-laying hens at the age of 20 and 30 weeks old by transcriptome sequencing and to investigate the interaction networks among lncRNAs, mRNAs and miRNAs. Moreover, the regulatory mechanism and biological function of lncLTR, a significantly differentially expressed lncRNA in the liver between pre- and peak-laying hens, was explored in vitro and in vivo. Methods: Bioinformatics analyses were conducted to identify the differentially expressed (DE) lncRNAs between the two groups of hens. The target genes of the DE lncRNA were predicated for further functional enrichment. An integrated analysis was performed among the DE lncRNA datasets, DE mRNAs and DE miRNA datasets obtained from the same samples to predict the interaction relationship. In addition, in vivo and in vitro trials were carried out to determine the expression regulation of lncLTR, and polymorphism association analysis was conducted to detect the biological role of lncLTR. Results: A total of 124 DE lncRNAs with a P-value ≤ 0.05 were identified. Among them, 44 lncRNAs including 30 known and 14 novel lncRNAs were significant differentially expressed (SDE) with FDR ≤ 0.05. Thirty-two lncRNAs were upregulated and 12 were downregulated in peak-laying group compared with pre-laying group. The functional enrichment results revealed that target genes of some lncRNAs are involved in the lipid metabolism process. Integrated analysis suggested that some of the genes involved in lipid metabolism might be regulated by both the lncRNA and the miRNA. In addition, an upregulated lncRNA, designated lncLTR, was demonstrated to be induced by estrogen via ERβ signaling. The c242. G>A SNP in lncLTR was significantly associated with chicken carcass weight, evisceration weight, semi-evisceration weight, head weight, double-wing weight, claw weight traits, and blood biochemical index, especially for the blood triglyceride content. Conclusion: A series of lncRNAs associated with lipid metabolism in the livers of chickens were identified by transcriptome sequencing and functional analysis, providing a valuable data resource for further studies on chicken hepatic metabolism activities. LncLTR was regulated by estrogen via ERβ signaling and associated with chicken carcass trait and blood triglyceride content. [ABSTRACT FROM AUTHOR]

Background/Aims: Malaria is the most deadly parasitic infection in the world, resulting in damage to various organs, including the liver, of the infected organism; however, the mechanism causing this damage in the liver remains unclear. Liver fibrosis, a major characteristic of liver diseases, occurs in response to liver injury and is regulated by a complex network of signaling pathways. Hedgehog (Hh) signaling orchestrates a number of hepatic responses including hepatic fibrogenesis. Therefore, we investigated whether Hh signaling influenced the liver's response to malarial infection. Methods: Eight-week-old male C57BL/6 mice inoculated with blood containing Plasmodium berghei ANKA (PbA)-infected erythrocytes were sacrificed when the level of parasitemia in the blood reached 10% or 30%, and the livers were collected for biochemical analysis. Liver responses to PbA infection were examined by hematoxylin and eosin staining, real-time polymerase chain reaction, immunohistochemistry and western blot. Results: Severe hepatic injury, such as ballooned hepatocytes, sinusoidal dilatation, and infiltrated leukocytes, was evident in the livers of the malaria-infected mice. Hypoxia was also induced in 30% parasitemia group. With the accumulation of Kupffer cells, inflammation markers, TNF-&$945;, interleukin-1β, and chemokine (C-X-C motif) ligand 1, were significantly upregulated in the infected group compared with the control group. Expression of fibrotic markers, including transforming growth factor-β, a-smooth muscle actin (a-SMA), collagen 1a1, thymosin β4, and vimentin, were significantly higher in the infected groups than in the control group. With increased collagen deposition, hepatic stellate cells expressing a-SMA accumulated in the liver of the PbA-infected mice, whereas those cells were rarely detected in the livers of the control mice. The levels of Hh signaling and Yes-associated protein (YAP), two key regulators for hepatic fibrogenesis, were significantly elevated in the infected groups compared with the control group. Treatment of mice with Hh inhibitor, GDC-0449, reduced hepatic inflammation and fibrogenesis with Hh suppression in PbA-infected mice. Conclusion: Our results demonstrate that HSCs are activated in and Hh and YAP signaling are associated with this process, contributing to increased hepatic fibrosis in malaria-infected livers. [ABSTRACT FROM AUTHOR]

Background/Aims: The Zucker diabetic fatty (ZDF, FA/FA) rat is a genetic model of type 2 diabetes, characterized by insulin resistance with progressive metabolic syndrome. We have previously demonstrated mitochondrial dysfunction and oxidative stress in the heart, kidneys and pancreas of ZDF rats. However, the precise molecular mechanism of disease progression is not clear. Our aim in the present study was to investigate oxidative stress and mitochondrial dysfunction in the liver and brain of ZDF rats. Methods: In this study, we have measured mitochondrial oxidative stress, bioenergetics and redox homeostasis in the liver and brain of ZDF rats. Results: Our results showed increased reactive oxygen species (ROS) production in the ZDF rat brain compared to the liver, while nitric oxide (NO) production was markedly increased both in the brain and liver. High levels of lipid and protein peroxidation were also observed in these tissues. Glutathione metabolism and mitochondrial respiratory functions were adversely affected in ZDF rats when compared to Zucker lean (ZL, +/FA) control rats. Reduced ATP synthesis was also observed in the liver and brain of ZDF rats. Western blot analysis confirmed altered expression of cytochrome P450 2E1, iNOS, p-JNK, and IκB-α confirming an increase in oxidative and metabolic stress in ZDF rat tissues. Conclusions: Our data shows that, like other tissues, ZDF rat liver and brain develop complications associated with redox homeostasis and mitochondrial dysfunction. These results, thus, might have implications in understanding the etiology and pathophysiology of diabesity which in turn, would help in managing the disease associated complications. © 2015 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

Background/Aims: Sterol regulatory element-binding protein (SREBP) 1, the master regulator of lipogenesis, was shown to be associated with non-alcoholic fatty liver disease, which is attributed to its major isoform SREBP1c. Based on studies in mice, the minor isoform SREBP1a is regarded as negligible for hepatic lipogenesis. This study aims to elucidate the expression and functional role of SREBP1a in human liver. Methods: mRNA expression of both isoforms was quantified in cohorts of human livers and primary human hepatocytes. Hepatocytes were treated with PF-429242 to inhibit the proteolytic activation of SREBP precursor protein. SREBP1a-specifc and pan-SREBP1 knock-down were performed by transfection of respective siRNAs. Lipogenic SREBP-target gene expression was analyzed by real-time RT-PCR. Results: In human liver, SREBP1a accounts for up to half of the total SREBP1 pool. Treatment with PF-429242 indicated SREBP-dependent auto-regulation of SREBP1a, which however was much weaker than of SREBP1c. SREBP1a-specifc knock-down also reduced significantly the expression of SREBP1c and of SREBP-target genes. Regarding most SREBP-target genes, simultaneous knock-down of both isoforms resulted in effects of only similar extent as SREBP1a-specific knock-down. Conclusion: We here showed that SREBP1a is significantly contributing to the human hepatic SREBP1 pool and has a share in human hepatic lipogenic gene expression. © 2015 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

Background: Pistacia lentiscus (Anacardiaceae) is a flowering plant traditionally used in the treatment of various skin, respiratory, and gastrointestinal disorders. The aim of this study was to assess whether Pistacia lentiscus oil has any short term toxic effects in vivo and in vitro. Methods: Pistacia lentiscus oil (100µl) was administered orally into mice for 5 days. Results: Measurements of body weight did not show any weight loss. Serum concentration of LDH did not show any significant statistical difference when compared to control mice. Similarly, blood, kidney or liver function tests showed no toxicity with Pistacia lentiscus oil when compared to the control group. Examination of gastrointestinal tissues sections revealed similar structural features with no difference in cell proliferation. In this context, pharmacological dilutions of Pistacia lentiscus oil (10-6 - 10-3) did not affect the viability (cell death and proliferation) of mouse gastric stem cells, human colorectal cancer cells HT29, human hepatoma cells HepG2. However, it appears that at the dose and time point studied, Pistacia lentiscus oil treatment has targeted various cytochrome P450s and has specifically inhibited the activities and the expression of CYP2E1, CYP3A4, CYP1A1 and CYP1A2 differentially in different tissues. Our results also demonstrate that there is no appreciable effect of Pistacia lentiscus oil on the GSH-dependent redox homoeostasis and detoxification mechanism in the tissues. Conclusion: These data suggest a good safety profile of short term oral use of Pistacia lentiscus oil as a monotherapy in the treatment of various skin, respiratory, and gastrointestinal disorders. However, due to its inhibitory effect of various cytochrome P450s and mainly CYP3A4, this might have implications on the bioavailability and metabolism of drugs taken in combination with Pistacia lentiscus oil. More attention is needed when Pistacia lentiscus oil is intended to be uses in combination with other pharmacological agents in order to avoid potential drug-drug interaction leading to toxicity. This study will help in safer use of Pistacia lentiscus oil for therapeutic purpose. © 2014 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

Background/Aims: Pendrin (SLC26A4), a transporter accomplishing anion exchange, is expressed in inner ear, thyroid gland, kidneys, lung, liver and heart. Loss or reduction of function mutations of SLC26A4 underlie Pendred syndrome, a disorder invariably leading to hearing loss with enlarged vestibular aqueducts and in some patients to hypothyroidism and goiter. Renal pendrin expression is up-regulated by mineralocorticoids such as aldosterone or deoxycorticosterone (DOCA). Little is known about the impact of mineralocorticoids on pendrin expression in extrarenal tissues. Methods: The present study utilized RT-qPCR and Western blotting to quantify the transcript levels and protein abundance of Slc26a4 in murine kidney, thyroid, heart and lung prior to and following subcutaneous administration of 100 mg/kg DOCA. Results: Slc26a4 transcript levels as compared to Gapdh transcript levels were significantly increased by DOCA treatment in kidney, heart, lung and thyroid. Accordingly pendrin protein expression was again significantly increased by DOCA treatment in kidney, heart, lung and thyroid. Conclusion: The observations reveal mineralocorticoid sensitivity of pendrin expression in kidney, heart, thyroid and lung. Copyright © 2012 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

Background/aims: Glitazones are synthetic insulin-sensitizing drugs which act as agonists of peroxisome proliferator-activated receptor gamma (PPARγ). However, TZDs action does not exclude independent PPARγ-activation effects. Remarkably, direct mitochondrial action of these agents has not been fully studied yet. Methods: Oxygen consumption rates (JO2) were measured using a Clark-type oxygen electrode in intact hepatocytes and isolated liver mitochondria. Mitochondrial reactive oxygen species (ROS) production was quantified by fluorescence assay. Moreover, activities of mitochondrial respiratory chain complex I, II and III were spectrometrically determined. Results: Pioglitazone and rosiglitazone inhibited JO2 in liver cells and mitochondria. This inhibition affected the state 3 of respiration (in the presence of ADP) and the uncoupled state (after addition of dinitrophenol). Moreover, these agents dramatically reduced mitochondrial ROS production in all situations tested. We also demonstrated that both glitazones specifically inhibited the activities of complex I and complex III, by 50% and 35% respectively. Additionally, they do not modify neither the oxidative phosphorylation yield nor the permeability transition pore opening. Conclusions: Pioglitazone and rosiglitazone reduce both respiration intensity and ROS production, acutely and by a probable PPARγ-independent way, through inhibition of complex I and III activities. This new finding could positively contribute to their anti-diabetic properties. Copyright © 2011 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

Background/Aims: GPR43 and GPR120 have recently been deorphanised as receptors for fatty acids. Fatty acids mediate a variety of metabolic processes in the body, however, the effect these receptors have on metabolism is not fully understood. Here, we characterise the effect of diet-induced obesity on the expression of GPR43 and GPR120 in tissues important in maintaining metabolic health. Methods: Six-week old male Sprague Dawley rats were fed either a high fat diet (HFD; 22% fat) or control diet (5% fat; n = 8-9/group) for 12 weeks. Rats were euthanized and the heart, liver, soleus and extensor digitorum longus (EDL) skeletal muscles were excised. GPR43 and GPR120 receptor abundance was quantified by 'real-time' PCR. Results: GPR43 mRNA abundance was significantly up-regulated by a HFD in liver and soleus and EDL skeletal muscles compared to control (p ≤ 0.05). Whilst a HFD significantly up-regulated GPR120 gene transcripts in cardiac tissue and EDL skeletal muscle when compare to control (p ≤ 0.05). Conclusion: We have shown for the first time that up-regulation of GPR43 and GPR120 in response to a HFD, is tissue specific. This suggests these receptors have different roles in mediating metabolic function in a number of tissues in the human body. Copyright © 2011 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

SLC26A4 encodes pendrin, a transporter exchanging anions such as chloride, bicarbonate, and iodide. Loss of function mutations of SLC26A4 cause Pendred syndrome characterized by hearing loss and enlarged vestibular aqueducts as well as variable hypothyroidism and goiter. In the kidney, pendrin is expressed in the distal nephron and accomplishes HCO3- secretion and Cl- reabsorption. Renal pendrin expression is regulated by acid-base balance. The liver contributes to acid-base regulation by producing or consuming glutamine, which is utilized by the kidney for generation and excretion of NH4+, paralleled by HCO3- formation. Little is known about the regulation of pendrin in liver. The present study thus examined the expression of Slc26a4 in liver and kidney of mice drinking tap water without or with NaHCO3 (150 mM), NH4Cl (280 mM) or acetazolamide (3.6 mM) for seven days. As compared to Gapdh transcript levels, Slc26a4 transcript levels were moderately lower in liver than in renal tissue. Slc26a4 transcript levels were not significantly affected by NaHCO3 in liver, but significantly increased by NaHCO3 in kidney. Pendrin protein expression was significantly enhanced in kidney and reduced in liver by NaHCO3. Slc26a4 transcript levels were significantly increased by NH4Cl and acetazolamide in liver, and significantly decreased by NH4Cl and by acetazolamide in kidney. NH4Cl and acetazolamide reduced pendrin protein expression significantly in kidney, but did not significantly modify pendrin protein expression in liver. The observations point to expression of pendrin in the liver and to opposite effects of acidosis on pendrin transcription in liver and kidney. Copyright © 2011 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

Aims: Mitochondrial dysfunction plays important roles in the development of diabetes. Elevated nitric oxide (NO) synthase inhibitor asymmetric dimethylarginine (ADMA) has been shown to be closely related to diabetes. But the relationship between them in diabetes has not been determined. This study was to explore the role of ADMA in hepatic mitochondrial dysfunction and its potential mechanisms in diabetic rats and hepatocytes. Methods: Respiratory enzymes activities, mitochondrial transmembrane potential and ATP content were measured to evaluate mitochondrial function. The copy number ratio of mitochondrial gene to nuclear gene was used to represent mitochondrial biogenesis. The activity of superoxide dismutase and malondialdehyde content were detected to reflect oxidative stress. Furthermore, changes in ADMA and NO contents, uncoupling protein 2 (UCP2) and peroxisome proliferator-activated receptor-γ coactivator 1α (PGC-1α) transcriptions were determined. Results: Elevated ADMA levels in serum of diabetic rats were found to be associated with hepatic mitochondrial dysfunction reflected by reductions of respiratory enzyme activities, mitochondrial membrane potential and ATP contents. Similar mitochondrial dysfunction also occurred in ADMA-treated hepatocytes. The mitochondrial dysfunction observed in diabetic rats or hepatocytes was accompanied with suppressions of mitochondrial biogenesis, PGC-1α transcription and NO synthesis as well as enhances of UCP 2 transcription and oxidative stress. These effects of ADMA could be attenuated by treatments with antioxidant or NO donor. Conclusions: These results indicate that elevated endogenous ADMA contributes to hepatic mitochondrial dysfunction in diabetic rats, and underlying mechanisms may be related to the suppression of mitochondrial biogenesis and mitochondrial uncoupling via inhibiting NO synthesis and enhancing oxidative stress. Copyright © 2011 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

Background/Aims: Mitochondrial biogenesis includes both mitochondrial proliferation and differentiation and its regulation under different physiological conditions is not clear. Given the sexual dimorphism previously found in mitochondrial function, the aim of this study was to investigate the gender-dependent effect of chronic high-fat-diet (HFD) feeding on rat liver mitochondrial function and biogenesis. Methods: Ten-week old male and female rats were fed a HFD (26% fat) or a control diet (2.9% fat) for 26 weeks. Mitochondrial morphology was studied. Mitochondrial DNA and protein content, hydrogen peroxide production, oxidative capacity, antioxidant defenses, as well as markers of oxidative damage and mitochondrial biogenesis were analyzed. Results: Female rats showed higher levels of mitochondrial protein and an enhanced oxidative capacity per mitochondrion than males. In both genders, HFD feeding increased mtDNA content and decreased mitochondrial differentiation markers. Conclusion: In comparison to male rats, females show higher oxidative capacity as a consequence of their greater mitochondrial differentiation under both control and obese status. In response to HFD feeding, the oxidative capacity of the whole mitochondrial population is maintained in both genders. This is obtained by means of an enhancement of mitochondrial proliferation, which counteracts the diet-induced impairment of the function of each mitochondrion. Copyright © 2010 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

Vitamin A, mainly as retinoic acid (RA), is known to affect the development and function of adipose tissues. Treatment with RA reduces body weight and adiposity independent of changes in food intake in mice. Lipid metabolism in liver can have a major impact on whole body adiposity. The aim of this work was to investigate the effects of an in vivo treatment with RA on hepatic lipid metabolism in mice. Adult, standard diet-fed mice were treated with different doses of all-trans RA or vehicle (subcutaneous injection) for 4 days before sacrifice. Food intake and body weight changes during treatment were determined, as well as adiposity, liver composition, levels of circulating metabolites and lipoproteins and expression levels of key mRNA species in liver following sacrifice. RA treatment resulted in reduced body weight and adiposity, as expected. In the liver, RA treatment triggered an increase in the mRNA expression levels of peroxisome proliferator-activated receptor alpha, retinoid X receptor alpha, uncoupling protein 2, liver-type carnitine palmitoyltransferase 1, and carnitine/acylcarnitine carrier, and a reduction in the mRNA expression levels of sterol regulatory element binding protein 1c and fatty acid synthase. Consistent with the changes in gene expression, hepatic triacylglycerol content and circulating VLDL fraction were reduced and levels of circulating ketone bodies increased after RA treatment. These results point to a capacity of active vitamin A forms to shift liver lipid metabolism in vivo towards increased catabolism and reduced lipogenesis. These effects might contribute to the reduction of adiposity brought about by RA treatment. Copyright © 2010 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

This study identifies a novel organic anion transporter Oat9 expressed in mouse liver and kidney. Two variants were detected by screening a mouse liver cDNA library; these varients consist of 1815 (designated Oat9S) and 2165 (Oat9L) base pairs which encode 443 and 551 amino acid proteins, respectively. Oat9S has a predicted structure containing eight transmembrane domains (TMD); whereas, Oat9L possesses twelve TMD. Oat9 mRNA expression was detected in kidney and liver. This transporter was located at the apical side of the late portion of proximal tubules and at the sinusoidal side of hepatocytes. When expressed in Xenopus oocytes, Oat9S mediated the transport of L-carnitine (Km = 2.9 μM), a representative zwitterion, as well as cimetidine (Km = 16.1 μM) and salicylic acid (Km = 175.5 μM), while Oat9L did not show any transport activity. Oat9S-mediated L-carnitine uptake was inhibited by D-carnitine, acetylcarnitine, octanoylcarnitine, betaine, and other organic compounds, suggesting that quaternary ammonium cation bulkiness and relative hydrophobicity are important factors for Oat9S-substrate interactions. Among OATs, Oat9S appears to be the first member to mediate the transport of carnitine and possesses eight TMD. Overall, these new results provide added insight into the structure-activity relationship comprising the organic ion-permeation pathway. Copyright © 2010 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

Background/Aims: Uncoupling protein-3 (UCP3) is expressed in liver only under conditions of high fatty acid catabolism. However, the specific role of UCP3 in liver mitochondria and overall hepatic function is still poorly known. Methods: A model of “in vivo” induction of UCP3 expression in mouse liver mitochondria via a tail-vein injection of a recombinant adenoviral vector was developed. The effects on liver mitochondrial bioenergetics and permeability transition, liver gene expression, and systemic metabolism were then determined. Results: UCP3 expression in liver did not cause basal, non-specific, uncoupling but led to a stimulation of palmitate-induced state 4 respiration. UCP3 expression in liver also caused an increase in the expression of certain genes involved in lipid catabolism and metabolic response to starvation (e.g. medium chain acyl-CoA-dehydrogenase or peroxisome proliferator-activated receptor-γ co-activator-1α). UCP3 also conferred to liver mitochondria an enhanced sensitivity to classical inducers of permeability transition, such as calcium and carboxyatractylate. Conclusion: UCP3 expression in liver exerts direct actions on mitochondrial activity, favoring fatty acid-induced uncoupling and sensitizing mitochondria to permeability transition, as well as causing retrograde signaling to nuclear gene expression consistent with favoring lipid catabolism and oxidative metabolism. Copyright © 2009 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

In this study, we demonstrate that a putative membrane unknown solute transporter 1 of the rat kidney (UST1r; Slc22a9) is a multispecific transporter of organic anions (OAs). When expressed in Xenopus oocytes, UST1r mediated uptake of ochratoxin A (OTA; Km = 1.0 μM) and sulfate conjugates of steroids, such as estrone-3-sulfate (ES; Km = 3.1 μM) and dehydroepiandrosterone sulfate (DHEAS; Km = 2.1 μM) in a sodium-independent manner. We herein propose that UST1r be renamed OA transporter 8 (rOat8). rOat8 interacted with chemically heterogenous anionic compounds, such as nonsteroidal anti-inflammatory drugs, diuretics, probenecid, taurocholate, and methotrexate, but not with the organic cation tetraethylammonium. The rOat8-mediated ES transport was: a) cis-inhibited by 4-methylumbelliferyl sulfate and β-estradiol sulfate, but not by glucuronide conjugates of these compounds, b) cis-inhibited by four- and five- carbon (C4/C5) dicarboxylates (succinate and glutarate (GA)), and c) trans-stimulated by GA, whereas the efflux of GA was significantly trans-stimulated by ES. By RT-PCR, rOat8 mRNA was expressed in proximal convoluted tubules and cortical and outer medullary collecting ducts, whereas in immunochemical studies, Oat8 was identified as the ñ58 kDa protein that in the collecting duct colocalized with the V-ATPase in plasma membranes and intracellular vesicles in various subtypes of intercalated cells. Molecular identification of Oat8 in these cells indicates a possible novel role of OAT family in the renal secretion/reabsorption of OA and acids and bases via affecting the V-ATPase-dependent functions. Copyright © 2008 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

Recently, we reported that the death of ouabain-treated C7-MDCK cells resembling principal cells from collecting ducts of the Madin-Darby canine kidney (MDCK) is caused by ouabain interaction with Na+,K+-ATPase but is not mediated by inversion of the [Na+]i/[K+]i ratio. The mechanism of this intriguing phenomenon remains unknown. We therefore examined the action of ouabain on serine/threonine phosphoproteins as possible intermediates of cell death signaling. The death of ouabain-treated C7-MDCK cells proceeded by altered phosphorylation of the RRXS*/T*-motif in 4 proteins with Mr from 80 to 25 kDa. Similarly to cell death, inversion of the [Na+]i/[K+]i ratio evoked by Na+,K+-ATPase inhibition in K+-free medium did not affect the phosphorylation of RRXS*/T*-proteins but increased their sensitivity to ouabain. The action of ouabain was preserved in the presence of activators of protein kinases A (forskolin), G (sodium nitroprusside) and C (PMA) as well as inhibitors of protein kinase C (Gö 6983, Gö 6976) and serine-threonine phosphatases (okadaic acid). Phosphorylation of RRXS*/T*-proteins was also noted in ouabain-sensitive C11-MDCK cells resembling intercalated cells from collecting ducts, but was absent in ouabain-resistant smooth muscle cells from the rat aorta. Our results show that altered phosphorylation of RRXS*/T*-proteins in ouabain-treated C7-MDCK cells is mediated by its interaction with Na+,K+-ATPase but is not caused by inversion of the [Na+]i/[K+]i ratio. The molecular origin of serine-threonine kinases and/or phosphatases involved in phosphorylation of ouabain-sensitive proteins and their role in cell death signaling should be examined further. Copyright © 2008 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

We were interested whether PKCα, δ, ε or ζ is the isoform actually employed in the activation of hypertonicity-induced cation channels (HICCs) in primary cultures of rat hepatocytes. Quantitative SDS-page and Western-blot experiments revealed that PKCα, δ and ε were stimulated by Indolactam V (as a DAG substitute for activation of c and nPKCs) but that only PKCδ and ε did respond to hypertonic stress. Furthermore, chelation of intracellular Ca++ by BAPTA-AM did not alter HICC activation in cable-analysis experiments whereas Indolactam V as well as V8 (an Indolactam derivative specific for PKCδ and ε) activated HICC currents under isotonic conditions. Finally, by use of Rottlerin (as an inhibitor exhibiting a slight preference for PKCδ over ε) PKCε could be identified as the most likely isoform responsible for the activation of the HICC. Copyright © 2007 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

Background/Aims: 3,4-Dimethoxy-2-phenylethylamine is catalyzed to its aldehyde derivative by monoamine oxidase B, but the subsequent oxidation into the corresponding acid has not yet been studied. Oxidation of aromatic aldehydes is catalyzed mainly by aldehyde dehydrogenase and aldehyde oxidase. Methods: The present study examines the metabolism of 3,4-dimethoxy-2-phenylethylamine in vitro and in freshly prepared and cryopreserved guinea pig liver slices and the relative contribution of different aldehyde-oxidizing enzymes was estimated by pharmacological means. Results: 3,4-Dimethoxy-2-phenylethylamine was converted into the corresponding aldehyde when incubated with monoamine oxidase and further oxidized into the acid when incubated with both, monoamine oxidase and aldehyde oxidase. In freshly prepared and cryopreserved liver slices, 3,4-dimethoxyphenylacetic acid was the main metabolite of 3,4-dimethoxy-2- phenylethylamine. 3,4-Dimethoxyphenylacetic acid formation was inhibited by 85% from disulfiram (aldehyde dehydrogenase inhibitor) and by 75-80% from isovanillin (aldehyde oxidase inhibitor), whereas allopurinol (xanthine oxidase inhibitor) inhibited acid formation by only 25-30%. Conclusions: 3,4-Dimethoxy-2-phenylethylamine is oxidized mainly to its acid, via 3,4-dimethoxyphenylacetaldehyde, by aldehyde dehydrogenase and aldehyde oxidase with a lower contribution from xanthine oxidase. [ABSTRACT FROM AUTHOR]

Background/Aims: Proteasome inhibitors such as MG-132 are considered as potential therapeutical tools in different clinical settings. The dual specificity MAP-kinase phosphatase MKP-1 plays a role in balancing signals mediating cell death or survival. Here the effect of cell hydration on MG-132-induced MKP-1 expression was investigated in H4IIE rat hepatoma cells. Results: Hyperosmolarity (405mosmol/l) increased MKP-1 expression by MG-132, which was accompanied by an induction of c-Fos, c-Jun, cJun Ser73 phosphorylation, and AP-1 DNA binding. MKP-1 induction by MG-132 plus hyperosmolarity was sensitive to inhibition of p38MAPK and c-Jun-N-terminal kinases (JNKs) but not extracellular signal-regulated kinases Erk-1/Erk-2, and was accompanied by a decline of MAP-kinase activities. Although hyperosmolarity increased overall protein ubiquitination in presence of MG-132, ubiquitination of MKP-1 was found under normo-, but not hyperosmotic conditions. Hyperosmolarity also enabled MG-132 to induce poly-ADP-ribose polymerase (PARP) cleavage which was sensitive to inhibition of p38MAPK and JNKs but not Erk-1/Erk-2. PARP cleavage and caspase-3 activation in H4IIE cells treated with hyperosmolarity plus MG-132 was further increased by vanadate, consistent with a contribution of MKP-1 to counterbalance proapoptotic MAP-kinase signals. Conclusion: The findings suggest that among other factors cell hydration critically determines the cellular response to proteasome inhibitors. Copyright © 2005 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]

Background / Aims: To investigate postulated differences related to FGF-2 in liver morphology and expression of apoptosis-related factors after partial hepatectomy (PH). Methods: Homogenous FGF-2-deficient mice (C57BL/6J) with their FGF-2-(+/+) littermates (control) were used to examine the structure of regenerating livers after PH with light and electron microscopy. The regenerative response and BrDu incorporation were monitored. The expression of BclX-l, Bax, Fas, TNF-α, and Caspase-3 were measured by reverse transcription PCR (RT-PCR) and Northern blot analysis. Results: In the FGF-2-(-/-) group, hepatocytes and endothelial cells contain mitochondria with atypical cristae and fragmented endoplasmic reticulum structures compared to control. Sinusoids show irregular basal laminae. These changes are in accordance with a differential expression of apoptosis-related factors: FasL was expressed throughout the entire observation span (days 0 to 10 post-PH). Following PH, tumor necrosis factor alpha (TNFα)-mRNA levels were higher in FGF-2-(+/+) animals, while Fas as well as Bax and BclXl were overexpressed in FGF-2-(-/-) mice. Caspase-3-mRNA was similarly expressed in both groups, but Caspase-3 activity was elevated for 4 days in FGF-2-(-/-) mice. Conclusion: Despite morphologic differences, differences in the time schedule of DNA synthesis and differences in apoptotic response, the dynamics of liver regeneration in FGF-2-(-/-) mice were not impaired. [ABSTRACT FROM AUTHOR]

Background/Aims: Cryopreserved precision-cut liver slices (PCLS) in culture exhibit a rapid decline in ATP level and anabolic processes by unknown mechanisms. The aim of this study is to elucidate the key events explaining the alterations occurring in cryopreserved PCLS. Methods: Glucose metabolism, mitochondrial activities, ionic homeostasis and caspase-3 activity were assessed in fresh or cryopreserved (rapid freezing-thawing conditions) rat PCLS within the first hour of incubation in Williams' medium E at 37°C. Results: Despite a similar glycolytic activity under both conditions, only fresh PCLS were able to gradually recover their ATP and potassium content. Glycogen content dropped more rapidly in cryopreserved than in control PCLS. Only cryopreserved PCLS exhibited a decline in O2 consumption and a lower ATP/ADP ratio from 30 min of incubation with a loss of coupling of oxygen consumption to ATP synthesis and mitochondrial calcium transport after 1 hour of incubation. Caspase-3 activation was already present in cryopreserved PCLS after a few minutes of incubation. Conclusions: The lack of restoration of potassium/sodium exchange might be a primary event in the metabolic alterations occurring in cryopreserved PCLS after thawing, that contributes to further mitochondrial alterations, caspase activation and compromises ATP-dependent anabolic pathways. [ABSTRACT FROM AUTHOR]