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Find video protocols related to scientific articles indexed in Pubmed.
Macrophage migration inhibitory factor is required for recruitment of scar-associated macrophages during liver fibrosis.
J. Leukoc. Biol.
PUBLISHED: 11-16-2014
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Recruitment of peripheral monocytes to the liver is a key contributor to the response to injury. MIF can act as a chemokine and cytokine, regulating innate immune responses in many tissues and cell types. We hypothesized that MIF contributes to the progression of CCl4-induced hepatic fibrosis by regulating recruitment of SAM. SAMs dynamically regulate HSC activation and ECM degradation. To gain insight into the role of MIF in progression of liver fibrosis, we investigated markers of fibrosis and immune responses after chronic CCl4 administration to female C57BL/6 and MIF(-/-) mice. Chronic CCl4 exposure increased activation of HSC in WT mice, indicated by increased expression of ?SMA mRNA and protein, as well as mRNA for collagen 1?1; these responses were blunted in female MIF(-/-) mice. Despite lower activation of HSC in MIF(-/-) mice, accumulation of ECM was similar in WT and MIF(-/-)mice, suggesting a decreased rate of ECM degradation. Recruitment of SAMs was lower in MIF(-/-) mice compared with WT mice, both in their initial inflammatory phenotype, as well as in the later phase as proresolution macrophages. The decreased presence of resolution macrophages was associated with lower expression of MMP13 in MIF(-/-) mice. Taken together, these data indicate that MIF-dependent recruitment of SAMs contributes to degradation of ECM via MMP13, highlighting the importance of appropriate recruitment and phenotypic profile of macrophages in the resolution of fibrosis.
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Moderate, chronic ethanol feeding exacerbates carbon-tetrachloride-induced hepatic fibrosis via hepatocyte-specific hypoxia inducible factor 1?
Pharmacol Res Perspect
PUBLISHED: 08-05-2014
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The hypoxia-sensing transcriptional factor HIF1? is implicated in a variety of hepato-pathological conditions; however, the contribution of hepatocyte-derived HIF1? during progression of alcoholic liver injury is still controversial. HIF1? induces a variety of genes including those involved in apoptosis via p53 activation. Increased hepatocyte apoptosis is critical for progression of liver inflammation, stellate cell activation and fibrosis. Using hepatocyte-specific HIF1?-deficient mice (?HepHIF1?-/-), here we investigated the contribution of HIF1? to ethanol-induced hepatocyte apoptosis and its role in amplification of fibrosis after carbon tetrachloride (CCl4) exposure. Moderate ethanol feeding (11% of Kcal) induced accumulation of hypoxia-sensitive pimonidazole adducts and HIF1? expression in the liver within 4 days of ethanol feeding. Chronic CCl4 treatment increased M30-positive cells, a marker of hepatocyte apoptosis in pair-fed control mice. Concomitant ethanol feeding (11% of Kcal) amplified CCl4-induced hepatocyte apoptosis in livers of wild-type mice, associated with elevated p53(K386)acetylation, PUMA expression and Ly6c+ cell infiltration. Subsequent to increased apoptosis, ethanol enhanced induction of pro-fibrotic markers including stellate cell activation, collagen 1 expression and extracellular matrix deposition, following CCl4 exposure. Ethanol-induced exacerbation of hepatocyte apoptosis, p53(K386) acetylation and PUMA expression following CCl4 exposure was attenuated in livers of ?HepHIF1?-/- mice. This protection was also associated with a reduction in Ly6c(+) cell infiltration and decreased fibrosis in livers of ?HepHIF1?-/- mice. In summary, these results indicate that moderate ethanol exposure leads to hypoxia/HIF1?-mediated signaling in hepatocytes and induction of p53-dependent apoptosis of hepatocytes, resulting in increased hepatic fibrosis during chronic CCl4 exposure.
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The natural history of glycogen storage disease types VI and IX: Long-term outcome from the largest metabolic center in Canada.
Mol. Genet. Metab.
PUBLISHED: 08-01-2014
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Glycogen storage disease (GSD) types VI and IX are caused by phosphorylase system deficiencies. To evaluate the natural history and long-term treatment outcome of the patients with GSD-VI and -IX, we performed an observational retrospective case study of 21 patients with confirmed diagnosis of GSD-VI or -IX.
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Innate immunity and cell death in alcoholic liver disease: role of cytochrome P4502E1.
Redox Biol
PUBLISHED: 08-01-2014
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Ethanol-induced liver injury is a complex process dependent upon the interaction of multiple cell types in the liver, as well as activation of the innate immune response. Increased expression of CYP2E1 in response to high concentrations of ethanol leads to greater production of cytotoxic ethanol metabolites, which in turn contribute to production of reactive oxygen species, oxidative stress, and ultimately, cell death. Necroptotic hepatocyte cell death in response to ethanol is mediated via a CYP2E1-dependent expression of receptor-interacting protein kinase 3 (RIP3), a key component of the necroptosome. In response to alarmins released during ethanol-induced necroptosis, the innate immune response is activated. Macrophage migration inhibitory factor (MIF), a pro-inflammatory multikine involved in many disease processes, is an essential component to this response to injury. MIF expression is increased during ethanol exposure via a CYP2E1-dependent pathway, likely contributing to an exacerbated innate immune response and chronic inflammation after chronic ethanol. This review will discuss the complex interactions between CYP2E1-dependent expression of RIP3 and MIF in the pathophysiology of chronic ethanol-induced liver injury.
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Globular adiponectin inhibits ethanol-induced reactive oxygen species production through modulation of NADPH oxidase in macrophages: involvement of liver kinase B1/AMP-activated protein kinase pathway.
Mol. Pharmacol.
PUBLISHED: 05-21-2014
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Adiponectin, an adipokine predominantly secreted from adipocytes, has been shown to play protective roles against chronic alcohol consumption. Although excessive reactive oxygen species (ROS) production in macrophages is considered one of the critical events for ethanol-induced damage in various target tissues, the effect of adiponectin on ethanol-induced ROS production is not clearly understood. In the present study, we investigated the effect of globular adiponectin (gAcrp) on ethanol-induced ROS production and the potential mechanisms underlying these effects of gAcrp in macrophages. Here we demonstrated that gAcrp prevented ethanol-induced ROS production in both RAW 264.7 macrophages and primary murine peritoneal macrophages. Globular adiponectin also inhibited ethanol-induced activation of NADPH oxidase. In addition, gAcrp suppressed ethanol-induced increase in the expression of NADPH oxidase subunits, including Nox2 and p22(phox), via modulation of nuclear factor-?B pathway. Furthermore, pretreatment with compound C, a selective inhibitor of AMPK, or knockdown of AMPK by small interfering RNA restored suppression of ethanol-induced ROS production and Nox2 expression by gAcrp. Finally, we found that gAcrp treatment induced phosphorylation of liver kinase B1 (LKB1), an upstream signaling molecule mediating AMPK activation. Knockdown of LKB1 restored gAcrp-suppressed Nox2 expression, suggesting that LKB1/AMPK pathway plays a critical role in the suppression of ethanol-induced ROS production and activation of NADPH oxidase by gAcrp. Taken together, these results demonstrate that globular adiponectin prevents ethanol-induced ROS production, at least in part, via modulation of NADPH oxidase in macrophages. Further, LKB1/AMPK axis plays an important role in the suppression of ethanol-induced NADPH oxidase activation by gAcrp in macrophages.
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MyD88-dependent interplay between myeloid and endothelial cells in the initiation and progression of obesity-associated inflammatory diseases.
J. Exp. Med.
PUBLISHED: 04-21-2014
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Low-grade systemic inflammation is often associated with metabolic syndrome, which plays a critical role in the development of the obesity-associated inflammatory diseases, including insulin resistance and atherosclerosis. Here, we investigate how Toll-like receptor-MyD88 signaling in myeloid and endothelial cells coordinately participates in the initiation and progression of high fat diet-induced systemic inflammation and metabolic inflammatory diseases. MyD88 deficiency in myeloid cells inhibits macrophage recruitment to adipose tissue and their switch to an M1-like phenotype. This is accompanied by substantially reduced diet-induced systemic inflammation, insulin resistance, and atherosclerosis. MyD88 deficiency in endothelial cells results in a moderate reduction in diet-induced adipose macrophage infiltration and M1 polarization, selective insulin sensitivity in adipose tissue, and amelioration of spontaneous atherosclerosis. Both in vivo and ex vivo studies suggest that MyD88-dependent GM-CSF production from the endothelial cells might play a critical role in the initiation of obesity-associated inflammation and development of atherosclerosis by priming the monocytes in the adipose and arterial tissues to differentiate into M1-like inflammatory macrophages. Collectively, these results implicate a critical MyD88-dependent interplay between myeloid and endothelial cells in the initiation and progression of obesity-associated inflammatory diseases.
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Protective role of HO-1 and carbon monoxide in ethanol-induced hepatocyte cell death and liver injury in mice.
J. Hepatol.
PUBLISHED: 04-04-2014
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Alcoholic liver disease is associated with inflammation and cell death. Heme oxygenase-1 (HO-1) is a stress-inducible enzyme with anti-apoptotic and anti-inflammatory properties. Here we tested the hypothesis that induction of HO-1 or treatment with a carbon monoxide releasing molecule (CORM) during chronic ethanol exposure protects and/or reverses ethanol-induced liver injury.
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Cot/tpl2 participates in the activation of macrophages by adiponectin.
J. Leukoc. Biol.
PUBLISHED: 02-14-2014
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Whereas the main function of APN is to enhance insulin activity, it is also involved in modulating the macrophage phenotype. Here, we demonstrate that at physiological concentrations, APN activates Erk1/2 via the IKK?-p105/NF-??1-Cot/tpl2 intracellular signal transduction cassette in macrophages. In peritoneal macrophages stimulated with APN, Cot/tpl2 influences the ability to phagocytose beads. However, Cot/tpl2 did not modulate the known capacity of APN to decrease lipid content in peritoneal macrophages in response to treatment with oxLDL or acLDL. A microarray analysis of gene-expression profiles in BMDMs exposed to APN revealed that APN modulated the expression of ?3300 genes; the most significantly affected biological functions were the inflammatory and the infectious disease responses. qRT-PCR analysis of WT and Cot/tpl2 KO macrophages stimulated with APN for 0, 3, and 18 h revealed that Cot/tpl2 participated in the up-regulation of APN target inflammatory mediators included in the cytokine-cytokine receptor interaction pathway (KEGG ID 4060). In accordance with these data, macrophages stimulated with APN increased secretion of cytokines and chemokines, including IL-1?, IL-1?, TNF-?, IL-10, IL-12, IL-6, and CCL2. Moreover, Cot/tpl2 also played an important role in the production of these inflammatory mediators upon stimulation of macrophages with APN. It has been reported that different types of signals that stimulate TLRs, IL-1R, TNFR, Fc?R, and proteinase-activated receptor-1 activate Cot/tpl2. Here, we demonstrate that APN is a new signal that activates the IKK?-p105/NF-??1-Cot/tpl2-MKK1/2-Erk1/2 axis in macrophages. Furthermore, this signaling cassette modulates the biological functions triggered by APN in macrophages.
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Alcohol-induced autophagy contributes to loss in skeletal muscle mass.
Autophagy
PUBLISHED: 01-31-2014
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Patients with alcoholic cirrhosis and hepatitis have severe muscle loss. Since ethanol impairs skeletal muscle protein synthesis but does not increase ubiquitin proteasome-mediated proteolysis, we investigated whether alcohol-induced autophagy contributes to muscle loss. Autophagy induction was studied in: A) Human skeletal muscle biopsies from alcoholic cirrhotics and controls, B) Gastrocnemius muscle from ethanol and pair-fed mice, and C) Ethanol-exposed murine C2C12 myotubes, by examining the expression of autophagy markers assessed by immunoblotting and real-time PCR. Expression of autophagy genes and markers were increased in skeletal muscle from humans and ethanol-fed mice, and in myotubes following ethanol exposure. Importantly, pulse-chase experiments showed suppression of myotube proteolysis upon ethanol-treatment with the autophagy inhibitor, 3-methyladenine (3MA) and not by MG132, a proteasome inhibitor. Correspondingly, ethanol-treated C2C12 myotubes stably expressing GFP-LC3B showed increased autophagy flux as measured by accumulation of GFP-LC3B vesicles with confocal microscopy. The ethanol-induced increase in LC3B lipidation was reversed upon knockdown of Atg7, a critical autophagy gene and was associated with reversal of the ethanol-induced decrease in myotube diameter. Consistently, CT image analysis of muscle area in alcoholic cirrhotics was significantly reduced compared with control subjects. In order to determine whether ethanol per se or its metabolic product, acetaldehyde, stimulates autophagy, C2C12 myotubes were treated with ethanol in the presence of the alcohol dehydrogenase inhibitor (4-methylpyrazole) or the acetaldehyde dehydrogenase inhibitor (cyanamide). LC3B lipidation increased with acetaldehyde treatment and increased further with the addition of cyanamide. We conclude that muscle autophagy is increased by ethanol exposure and contributes to sarcopenia.
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Tributyrin supplementation protects mice from acute ethanol-induced gut injury.
Alcohol. Clin. Exp. Res.
PUBLISHED: 01-27-2014
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Excessive alcohol consumption leads to liver disease. Interorgan crosstalk contributes to ethanol (EtOH)-induced liver injury. EtOH exposure causes gut dysbiosis resulting in negative alterations in intestinal fermentation byproducts, particularly decreased luminal butyrate concentrations. Therefore, in the present work, we investigated the effect of butyrate supplementation, in the form of trybutyrin, as a prophylactic treatment against EtOH-induced gut injury.
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Chronic ethanol ingestion induces oxidative kidney injury through taurine-inhibitable inflammation.
Free Radic. Biol. Med.
PUBLISHED: 01-02-2014
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Chronic ethanol ingestion mildly damages liver through oxidative stress and lipid oxidation, which is ameliorated by dietary supplementation with the anti-inflammatory ?-amino acid taurine. Kidney, like liver, expresses cytochrome P450 2E1 that catabolizes ethanol with free radical formation, and so also may be damaged by ethanol catabolism. Sudden loss of kidney function, and not liver disease itself, foreshadows mortality in patients with alcoholic hepatitis [J. Altamirano, Clin. Gastroenterol. Hepatol. 2012, 10:65]. We found that ethanol ingestion in the Lieber-deCarli rat model increased kidney lipid oxidation, 4-hydroxynonenal protein adduction, and oxidatively truncated phospholipids that attract and activate leukocytes. Chronic ethanol ingestion increased myeloperoxidase-expressing cells in kidney and induced an inflammatory cell infiltrate. Apoptotic terminal deoxynucleotidyl transferase nick-end labeling-positive cells and active caspase-3 increased in kidney after ethanol ingestion, with reduced filtration with increased circulating blood urea nitrogen (BUN) and creatinine. These events were accompanied by release of albumin, myeloperoxidase, and the acute kidney injury biomarkers kidney injury molecule-1 (KIM-1), neutrophil gelatinase-associated lipocalin, and cystatin c into urine. Taurine sequesters HOCl from myeloperoxidase of activated leukocytes, and taurine supplementation reduced renal lipid oxidation, reduced leukocyte infiltration, and reduced the increase in myeloperoxidase-positive cells during ethanol feeding. Taurine supplementation also normalized circulating BUN and creatinine levels and suppressed enhanced myeloperoxidase, albumin, KIM-1, and cystatin c in urine. Thus, chronic ethanol ingestion oxidatively damages kidney lipids and proteins, damages renal function, and induces acute kidney injury through an inflammatory cell infiltrate. The anti-inflammatory nutraceutical taurine effectively interrupts this ethanol-induced inflammatory cycle in kidney.
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Anaphylatoxin C5a modulates hepatic stellate cell migration.
Fibrogenesis Tissue Repair
PUBLISHED: 01-01-2014
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C5a and its cognate receptor, C5a receptor (C5aR), key elements of complement, are critical modulators of liver immunity and fibrosis. However, the molecular mechanism for the cross talk between complement and liver fibrosis is not well understood. C5a is a potent chemokine regulating migration of cells in the innate immune system. Since activation and migration of hepatic stellate cells (HSC) are hallmarks of liver fibrosis, we hypothesized that C5a contributes to fibrosis by regulating HSC activation and/or migration.
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Mice lacking C1q are protected from high fat diet-induced hepatic insulin resistance and impaired glucose homeostasis.
J. Biol. Chem.
PUBLISHED: 06-20-2013
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Complement activation is implicated in the development of obesity and insulin resistance, and loss of signaling by the anaphylatoxin C3a prevents obesity-induced insulin resistance in mice. Here we have identified C1q in the classical pathway as required for activation of complement in response to high fat diets. After 8 weeks of high fat diet, wild-type mice became obese and developed glucose intolerance. This was associated with increased apoptotic cell death and accumulation of complement activation products (C3b/iC3b/C3c) in liver and adipose tissue. Previous studies have shown that high fat diet-induced apoptosis is dependent on Bid; here we report that Bid-mediated apoptosis was required for complement activation in adipose and liver. Although C1qa deficiency had no effect on high fat diet-induced apoptosis, accumulation of complement activation products and the metabolic complications of high fat diet-induced obesity were dependent on C1q. When wild-type mice were fed a high fat diet for only 3 days, hepatic insulin resistance was associated with the accumulation of C3b/iC3b/C3c in the liver. Mice deficient in C3a receptor were protected against this early high fat diet-induced hepatic insulin resistance, whereas mice deficient in the negative complement regulator CD55/DAF were more sensitive to the high fat diet. C1qa(-/-) mice were also protected from high fat diet-induced hepatic insulin resistance and complement activation. Evidence of complement activation was also detected in adipose tissue of obese women compared with lean women. Together, these studies reveal an important role for C1q in the classical pathway of complement activation in the development of high fat diet-induced insulin resistance.
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Kupffer cells in the liver.
Compr Physiol
PUBLISHED: 05-31-2013
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Kupffer cells are a critical component of the mononuclear phagocytic system and are central to both the hepatic and systemic response to pathogens. Kupffer cells are reemerging as critical mediators of both liver injury and repair. Kupffer cells exhibit a tremendous plasticity; depending on the local metabolic and immune environment, then can express a range of polarized phenotypes, from the proinflammatory M1 phenotype to the alternative/M2 phenotype. Multiple M2 phenotypes can be distinguished, each involved in the resolution of inflammation and wound healing. Here, we have provided an update on recent research that has contributed to the developing delineation of the contribution of Kupffer cells to different types of liver injury, with an emphasis on alcoholic and nonalcoholic liver diseases. These recent advances in our understanding of Kupffer cell function and regulation will likely provide new insights into the potential for therapeutic manipulation of Kupffer cells to promote the resolution of inflammation and enhance wound healing in liver disease.
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Lactobacillus GG and tributyrin supplementation reduce antibiotic-induced intestinal injury.
JPEN J Parenter Enteral Nutr
PUBLISHED: 04-29-2013
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Antibiotic therapy negatively alters the gut microbiota. Lactobacillus GG (LGG) decreases antibiotic-associated diarrhea (AAD) symptoms, but the mechanisms are unknown. Butyrate has beneficial effects on gut health. Altered intestinal gene expression occurs in the absence of gut microbiota. We hypothesized that antibiotic-induced changes in gut microbiota reduce butyrate production, varying genes involved with gut barrier integrity and water and electrolyte absorption, lending to AAD, and that simultaneous supplementation with LGG and/or tributyrin would prevent these changes.
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Absence of receptor interacting protein kinase 3 prevents ethanol-induced liver injury.
Hepatology
PUBLISHED: 03-14-2013
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Hepatocyte cell death via apoptosis and necrosis are major hallmarks of ethanol-induced liver injury. However, inhibition of apoptosis is not sufficient to prevent ethanol-induced hepatocyte injury or inflammation. Because receptor-interacting protein kinase (RIP) 3-mediated necroptosis, a nonapoptotic cell death pathway, is implicated in a variety of pathological conditions, we tested the hypothesis that ethanol-induced liver injury is RIP3-dependent and RIP1-independent. Increased expression of RIP3 was detected in livers of mice after chronic ethanol feeding, as well as in liver biopsies from patients with alcoholic liver disease. Chronic ethanol feeding failed to induce RIP3 in the livers of cytochrome P450 2E1 (CYP2E1)-deficient mice, indicating CYP2E1-mediated ethanol metabolism is critical for RIP3 expression in response to ethanol feeding. Mice lacking RIP3 were protected from ethanol-induced steatosis, hepatocyte injury, and expression of proinflammatory cytokines. In contrast, RIP1 expression in mouse liver remained unchanged following ethanol feeding, and inhibition of RIP1 kinase by necrostatin-1 did not attenuate ethanol-induced hepatocyte injury. Ethanol-induced apoptosis, assessed by terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling-positive nuclei and accumulation of cytokeratin-18 fragments in the liver, was independent of RIP3. Conclusion: CYP2E1-dependent RIP3 expression induces hepatocyte necroptosis during ethanol feeding. Ethanol-induced hepatocyte injury is RIP3-dependent, but independent of RIP1 kinase activity; intervention of this pathway could be targeted as a potential therapeutic strategy.
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Genetic resistance to liver fibrosis on A/J mouse chromosome 17.
Alcohol. Clin. Exp. Res.
PUBLISHED: 03-03-2013
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Because the histological and biochemical progression of liver disease is similar in alcoholic steatohepatitis (ASH) and nonalcoholic steatohepatitis (NASH), we hypothesized that the genetic susceptibility to these liver diseases would be similar. To identify potential candidate genes that regulate the development of liver fibrosis, we studied a chromosome substitution strain (CSS-17) that contains chromosome 17 from the A/J inbred strain substituted for the corresponding chromosome on the C57BL/6J (B6) genetic background. Previously, we identified quantitative trait loci (QTLs) in CSS-17, namely obesity-resistant QTL 13 and QTL 15 (Obrq13 and Obrq15, respectively), that were associated with protection from diet-induced obesity and hepatic steatosis on a high-fat diet.
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Macrophage migration inhibitory factor contributes to ethanol-induced liver injury by mediating cell injury, steatohepatitis, and steatosis.
Hepatology
PUBLISHED: 01-18-2013
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Macrophage migration inhibitory factor (MIF), a multipotent protein that exhibits both cytokine and chemotactic properties, is expressed by many cell types, including hepatocytes and nonparenchymal cells. We hypothesized that MIF is a key contributor to liver injury after ethanol exposure. Female C57BL/6 or MIF-/- mice were fed an ethanol-containing liquid diet or pair-fed control diet for 4 (11% total kcal;early response) or 25 (32% kcal; chronic response) days. Expression of MIF messenger RNA (mRNA) was induced at both 4 days and 25 days of ethanol feeding. After chronic ethanol, hepatic triglycerides and plasma alanine aminotransferase (ALT) and aspartate aminotransferase (AST) were increased in wildtype, but not MIF-/-, mice. In order to understand the role of MIF in chronic ethanol-induced liver injury, we investigated the early response of wildtype and MIF-/- to ethanol. Ethanol feeding for 4 days increased apoptosis of hepatic macrophages and activated complement in both wildtype and MIF-/- mice. However, tumor necrosis factor alpha (TNF-?) expression was increased only in wildtype mice. This attenuation of TNF-? expression was associated with fewer F4/80+ macrophages in liver of MIF-/- mice. After 25 days of ethanol feeding, chemokine expression was increased in wildtype mice, but not MIF-/- mice. Again, this protection was associated with decreased F4/80+ cells in MIF-/- mice after ethanol feeding. Chronic ethanol feeding also sensitized wildtype, but not MIF-/-, mice to lipopolysaccharide, increasing chemokine expression and monocyte recruitment into the liver. Conclusion: Taken together, these data indicate that MIF is an important mediator in the regulation of chemokine production and immune cell infiltration in the liver during ethanol feeding and promotes ethanol-induced steatosis and hepatocyte damage.
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4-Hydroxy-2(E)-nonenal (HNE) catabolism and formation of HNE adducts are modulated by ? oxidation of fatty acids in the isolated rat heart.
Free Radic. Biol. Med.
PUBLISHED: 01-06-2013
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We previously reported that a novel metabolic pathway functionally catabolizes 4-hydroxy-2(E)-nonenal (HNE) via two parallel pathways, which rely heavily on ?-oxidation pathways. The hypothesis driving this report is that perturbations of ? oxidation will alter the catabolic disposal of HNE, favoring an increase in the concentrations of HNE and HNE-modified proteins that may further exacerbate pathology. This study employed Langendorff perfused hearts to investigate the impact of cardiac injury modeled by ischemia/reperfusion and, in a separate set of perfusions, the effects of elevated lipid (typically observed in obesity and type II diabetes) by perfusing with increased fatty acid concentrations (1mM octanoate). During ischemia, HNE concentrations doubled and the glutathione-HNE adduct and 4-hydroxynonanoyl-CoA were increased by 7- and 10-fold, respectively. Under conditions of increased fatty acid, oxidation to 4-hydroxynonenoic acid was sustained; however, further catabolism through ? oxidation was nearly abolished. The inhibition of HNE catabolism was not compensated for by other disposal pathways of HNE, rather an increase in HNE-modified proteins was observed. Taken together, this study presents a mechanistic rationale for the accumulation of HNE and HNE-modified proteins in pathological conditions that involve alterations to ? oxidation, such as myocardial ischemia, obesity, and high-fat diet-induced diseases.
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Caspase-1 as a central regulator of high fat diet-induced non-alcoholic steatohepatitis.
PLoS ONE
PUBLISHED: 01-04-2013
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Nonalcoholic steatohepatitis (NASH) is associated with caspase activation. However, a role for pro-inflammatory caspases or inflammasomes has not been explored in diet-induced liver injury. Our aims were to examine the role of caspase-1 in high fat-induced NASH. C57BL/6 wild-type and caspase 1-knockout (Casp1(-/-)) mice were placed on a 12-week high fat diet. Wild-type mice on the high fat diet increased hepatic expression of pro-caspase-1 and IL-1?. Both wild-type and Casp1(-/-) mice on the high fat diet gained more weight than mice on a control diet. Hepatic steatosis and TG levels were increased in wild-type mice on high fat diet, but were attenuated in the absence of caspase-1. Plasma cholesterol and free fatty acids were elevated in wild-type, but not Casp1(-/-) mice, on high fat diet. ALT levels were elevated in both wild-type and Casp1(-/-) mice on high fat diet compared to control. Hepatic mRNA expression for genes associated with lipogenesis was lower in Casp1(-/-) mice on high fat diet compared to wild-type mice on high fat diet, while genes associated with fatty acid oxidation were not affected by diet or genotype. Hepatic Tnf? and Mcp-1 mRNA expression was increased in wild-type mice on high fat diet, but not in Casp1(-/-) mice on high fat diet. ?SMA positive cells, Sirius red staining, and Col1?1 mRNA were increased in wild-type mice on high fat diet compared to control. Deficiency of caspase-1 prevented those increases. In summary, the absence of caspase-1 ameliorates the injurious effects of high fat diet-induced obesity on the liver. Specifically, mice deficient in caspase-1 are protected from high fat-induced hepatic steatosis, inflammation and early fibrogenesis. These data point to the inflammasome as an important therapeutic target for NASH.
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Adenosine 2A receptor antagonist prevented and reversed liver fibrosis in a mouse model of ethanol-exacerbated liver fibrosis.
PLoS ONE
PUBLISHED: 01-01-2013
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The effect of moderate alcohol consumption on liver fibrosis is not well understood, but evidence suggests that adenosine may play a role in mediating the effects of moderate ethanol on tissue injury. Ethanol increases the concentration of adenosine in the liver. Adenosine 2A receptor (A2AR) activation is known to enhance hepatic stellate cell (HSC) activation and A2AR deficient mice are protected from fibrosis in mice. Making use of a novel mouse model of moderate ethanol consumption in which female C57BL/6J mice were allowed continued access to 2% (vol/vol) ethanol (11% calories) or pair-fed control diets for 2 days, 2 weeks or 5 weeks and superimposed with exposure to CCl4, we tested the hypothesis that moderate ethanol consumption increases fibrosis in response to carbon tetrachloride (CCl4) and that treatment of mice with an A2AR antagonist prevents and/or reverses this ethanol-induced increase in liver fibrosis. Neither the expression or activity of CYP2E1, required for bio-activation of CCl4, nor AST and ALT activity in the plasma were affected by ethanol, indicating that moderate ethanol did not increase the direct hepatotoxicity of CCl4. However, ethanol feeding enhanced HSC activation and exacerbated liver fibrosis upon exposure to CCl4. This was associated with an increased sinusoidal angiogenic response in the liver. Treatment with A2AR antagonist both prevented and reversed the ability of ethanol to exacerbate liver fibrosis.
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Ethanol-induced oxidative stress via the CYP2E1 pathway disrupts adiponectin secretion from adipocytes.
Alcohol. Clin. Exp. Res.
PUBLISHED: 09-06-2011
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Adipose tissue is an important target for ethanol action. One important effect of ethanol is to reduce the secretion of adiponectin from adipocytes; this decrease is associated with lowered circulating adiponectin in rodent models of chronic ethanol feeding. Adiponectin is an insulin-sensitizing, anti-inflammatory adipokine; decreased adiponectin activity may contribute to tissue injury in response to chronic ethanol. Here, we investigated the role of cytochrome P450 2E1 (CYP2E1) and oxidative stress in the mechanism for impaired adiponectin secretion from adipocytes in response to ethanol.
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Identification of a cytochrome P4502E1/Bid/C1q-dependent axis mediating inflammation in adipose tissue after chronic ethanol feeding to mice.
J. Biol. Chem.
PUBLISHED: 08-19-2011
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Chronic, heavy alcohol exposure results in inflammation in adipose tissue, insulin resistance, and liver injury. Here we have identified a CYP2E1/Bid/C1q-dependent pathway that is activated in response to chronic ethanol and is required for the development of inflammation in adipose tissue. Ethanol feeding for 25 days to wild-type (C57BL/6J) mice increased expression of multiple markers of adipose tissue inflammation relative to pair-fed controls independent of increased body weight or adipocyte size. Ethanol feeding increased the expression of CYP2E1 in adipocytes, but not stromal vascular cells, in adipose tissue and Cyp2e1(-/-) mice were protected from adipose tissue inflammation in response to ethanol. Ethanol feeding also increased the number of TUNEL-positive nuclei in adipose tissue of wild-type mice but not in Cyp2e1(-/-) or Bid (-/-) mice. Apoptosis contributed to adipose inflammation, as the expression of multiple inflammatory markers was decreased in mice lacking the Bid-dependent apoptotic pathway. The complement protein C1q binds to apoptotic cells, facilitating their clearance and activating complement. Making use of C1q-deficient mice, we found that activation of complement via C1q provided the critical link between CYP2E1/Bid-dependent apoptosis and onset of adipose tissue inflammation in response to chronic ethanol. In summary, chronic ethanol increases CYP2E1 activity in adipose, leading to Bid-mediated apoptosis and activation of complement via C1q, finally resulting in adipose tissue inflammation. Taken together, these data identify a novel mechanism for the development of adipose tissue inflammation that likely contributes to the pathophysiological effects of ethanol.
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Redox signaling and the innate immune system in alcoholic liver disease.
Antioxid. Redox Signal.
PUBLISHED: 05-25-2011
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The development of alcoholic liver disease (ALD) is a complex process involving both parenchymal and nonparenchymal cells resident in the liver. Although the mechanisms for ALD are not completely understood, it is clear that increased oxidative stress, and activation of the innate immune system are essential elements in the pathophysiology of ALD. Oxidative stress from ethanol exposure results from increased generation of reactive oxygen species and decreased hepatocellular antioxidant activity, including changes in the thioredoxin/peroxiredoxin family of proteins. Both cellular and circulating components of the innate immune system are activated by exposure to ethanol. For example, ethanol exposure enhances toll-like receptor-4 (TLR-4)-dependent cytokine expression by Kupffer cells, likely due, at least in part, to dysregulation of redox signaling. Similarly, complement activation in response to ethanol leads to increased production of the anaphylatoxins, C3a and C5a, and activation C3a receptor and C5a receptor. Complement activation thus contributes to increased inflammatory cytokine production and can influence redox signaling. Here we will review recent progress in understanding the interactions between oxidative stress and innate immunity in ALD. These data illustrate that ethanol-induced oxidative stress and activation of the innate immune system interact dynamically during ethanol exposure, exacerbating ethanol-induced liver injury.
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Pathogenesis of alcoholic liver disease: interactions between parenchymal and non-parenchymal cells.
J Dig Dis
PUBLISHED: 05-25-2011
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The development of alcoholic liver disease (ALD) is a complex process involving both the parenchymal and non-parenchymal cells in the liver. The impact of ethanol on hepatocytes can be characterized as a condition of organelle stress with multifactorial changes in hepatocellular function accumulating during ethanol exposure. These changes include oxidative stress, mitochondrial dysfunction, decreased methylation capacity, endoplasmic reticulum stress, impaired vesicular trafficking and altered proteasome function. Injury to hepatocytes is attributed, in part, to ethanol metabolism by the hepatocytes. Changes in the structural integrity of hepatic sinusoidal endothelial cells, as well as enhanced inflammation in the liver during ethanol exposure are also important contributors to injury. Activation of hepatic stellate cells initiates the deposition of extracellular matrix proteins characteristic of fibrosis. Kupffer cells, the resident macrophages in the liver, are particularly critical to the onset of ethanol-induced liver injury. Chronic ethanol exposure sensitizes Kupffer cells to activation by lipopolysaccharides via toll-like receptor 4. This sensitization enhances the production of inflammatory mediators, such as tumor necrosis factor-? and reactive oxygen species that contribute to hepatocyte dysfunction, necrosis and apoptosis of hepatocytes and the generation of extracellular matrix proteins leading to fibrosis. In this review we provide an overview of the complex interactions between parenchymal and non-parenchymal cells in the liver during the progression of ethanol-induced liver injury.
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Early growth response (EGR)-1 is required for timely cell-cycle entry and progression in hepatocytes after acute carbon tetrachloride exposure in mice.
Am. J. Physiol. Gastrointest. Liver Physiol.
PUBLISHED: 03-17-2011
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Cell-cycle induction in hepatocytes protects from prolonged tissue damage after toxic liver injury. Early growth response (Egr)-1(-/-) mice exhibit increased liver injury after carbon tetrachloride (CCl(4)) exposure and reduced TNF-? production. Because TNF-? is required for prompt cell-cycle induction after liver injury, here, we tested the hypothesis that Egr-1 is required for timely hepatocyte entry into the cell cycle after CCl(4)-induced liver injury. Acute liver injury was induced by a single injection of CCl(4). Assays were employed to assess indices of the cell cycle in liver after CCl(4) exposure. Bromodeoxyuridine incorporation peaked in wild-type mice at 48 h after CCl(4) but was reduced by 80% in Egr-1(-/-) mice. Proliferating-cell nuclear-antigen immunohistochemistry revealed blocks in cell-cycle entry and progression to DNA synthesis in Egr-1-deficient mice 48 h after CCl(4). Cyclin D, important for G0/G1 progression, was reduced at baseline and 36 h after CCl(4). Cyclin E1, required for G1/S-phase transition, was reduced in Egr-1(-/-) mice 24 and 48 h after CCl(4) exposure and was associated with reduced phosphorylation of the retinoblastoma protein. Proliferation in Egr-1(-/-) mice was delayed, rather than blocked, because indices of cell-cycle progression were restored 72 h after CCl(4) exposure. We concluded that Egr-1 was required for prompt cell-cycle entry (G0- to G1-phase) and G1/S-phase transition after toxic liver injury. These data support the hypothesis that Egr-1 provides hepatoprotection in the CCl(4)-injured liver, attributable, in part, to timely cell-cycle induction and progression.
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Molecular mechanism for adiponectin-dependent M2 macrophage polarization: link between the metabolic and innate immune activity of full-length adiponectin.
J. Biol. Chem.
PUBLISHED: 02-25-2011
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The anti-inflammatory effects of globular adiponectin (gAcrp) are mediated by IL-10/heme oxygenase 1 (HO-1)-dependent pathways. Although full-length (flAcrp) adiponectin also suppresses LPS-induced pro-inflammatory signaling, its signaling mechanisms are not yet understood. The aim of this study was to examine the differential mechanisms by which gAcrp and flAcrp suppress pro-inflammatory signaling in macrophages. Chronic ethanol feeding increased LPS-stimulated TNF-? expression by Kupffer cells, associated with a shift to an M1 macrophage polarization. Both gAcrp and flAcrp suppressed TNF-? expression in Kupffer cells; however, only the effect of gAcrp was dependent on IL-10. Similarly, inhibition of HO-1 activity or siRNA knockdown of HO-1 in RAW264.7 macrophages only partially attenuated the suppressive effects of flAcrp on MyD88-dependent and -independent cytokine signatures. Instead, flAcrp, acting via the adiponectin R2 receptor, potently shifted the polarization of Kupffer cells and RAW264.7 macrophages to an M2 phenotype. gAcrp, acting via the adiponectin R1 receptor, was much less effective at eliciting an M2 pattern of gene expression. M2 polarization was also partially dependent on AMP-activated kinase. flAcrp polarized RAW264.7 macrophages to an M2 phenotype in an IL-4/STAT6-dependent mechanism. flAcrp also increased the expression of genes involved in oxidative phosphorylation in RAW264.7 macrophages, similar to the effect of flAcrp on hepatocytes. In summary, these data demonstrate that gAcrp and flAcrp utilize differential signaling strategies to decrease the sensitivity of macrophages to activation by TLR4 ligands, with flAcrp utilizing an IL-4/STAT6-dependent mechanism to shift macrophage polarization to the M2/anti-inflammatory phenotype.
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Obesity, diabetes mellitus, and liver fibrosis.
Am. J. Physiol. Gastrointest. Liver Physiol.
PUBLISHED: 02-24-2011
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Obesity is a global epidemic with more than 1 billion overweight adults and at least 300 million obese patients worldwide. Diabetes is characterized by a defect in insulin secretion or a decrease in sensitivity to insulin, which results in elevated fasting blood glucose. Both obesity and elevated fasting glucose are risk factors for nonalcoholic fatty liver disease, a disease spectrum that includes hepatic steatosis (nonalcoholic fatty liver), nonalcoholic steatohepatitis (NASH), fibrosis, and cirrhosis. Increased adiposity and insulin resistance contribute to the progression from NASH to fibrosis through the development of a profibrotic mileau in the liver, including increased hepatocellular death, increased reactive oxygen species generation, and an altered adipokine/cytokine balance. This review will summarize recent advances in our understanding of the pathological interactions among excessive fat accumulation, insulin resistance, and hepatic fibrogenesis and discuss specific molecular pathways that may be of interest in the development of therapeutic interventions to prevent and/or reverse hepatic fibrosis.
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Innate immunity in alcoholic liver disease.
Am. J. Physiol. Gastrointest. Liver Physiol.
PUBLISHED: 01-20-2011
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Excessive alcohol consumption is a leading cause of chronic liver disease in the Western world. Alcohol-induced hepatotoxicity and oxidative stress are important mechanisms contributing to the pathogenesis of alcoholic liver disease. However, emerging evidence suggests that activation of innate immunity involving TLR4 and complement also plays an important role in initiating alcoholic steatohepatitis and fibrosis, but the role of adaptive immunity in the pathogenesis of alcoholic liver disease remains obscure. Activation of a TLR4-mediated MyD88-independent (TRIF/IRF-3) signaling pathway in Kupffer cells contributes to alcoholic steatohepatitis, whereas activation of TLR4 signaling in hepatic stellate cells promotes liver fibrosis. Alcohol consumption activates the complement system in the liver by yet unidentified mechanisms, leading to alcoholic steatohepatitis. In contrast to activation of TLR4 and complement, alcohol consumption can inhibit natural killer cells, another important innate immunity component, contributing to alcohol-mediated acceleration of viral infection and liver fibrosis in patients with chronic viral hepatitis. Understanding of the role of innate immunity in the pathogenesis of alcoholic liver disease may help us identify novel therapeutic targets to treat this disease.
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Adiponectin and heme oxygenase-1 suppress TLR4/MyD88-independent signaling in rat Kupffer cells and in mice after chronic ethanol exposure.
J. Immunol.
PUBLISHED: 09-22-2010
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Alcoholic liver disease is mediated via activation of TLR4 signaling; MyD88-dependent and -independent signals are important contributors to injury in mouse models. Adiponectin, an anti-inflammatory adipokine, suppresses TLR4/MyD88-dependent responses via induction of heme oxygenase-1 (HO-1). Here we investigated the interactions between chronic ethanol, adiponectin, and HO-1 in regulation of TLR4/MyD88-independent signaling in macrophages and an in vivo mouse model. After chronic ethanol feeding, LPS-stimulated expression of IFN-? and CXCL10 mRNA was increased in primary cultures of Kupffer cells compared with pair-fed control mice. Treatment of Kupffer cells with globular adiponectin (gAcrp) normalized this response. LPS-stimulated IFN-?/CXCL10 mRNA and CXCL10 protein was also reduced in RAW 264.7 macrophages treated with gAcrp or full-length adiponectin. gAcrp and full-length adiponectin acted via adiponectin receptors 1 and 2, respectively. gAcrp decreased TLR4 expression in both Kupffer cells and RAW 264.7 macrophages. Small interfering RNA knockdown of HO-1 or inhibition of HO-1 activity with zinc protoporphyrin blocked these effects of gAcrp. C57BL/6 mice were exposed to chronic ethanol feeding, with or without treatment with cobalt protoporphyrin, to induce HO-1. After chronic ethanol feeding, mice were sensitized to in vivo challenge with LPS, expressing increased IFN-?/CXCL10 mRNA and CXCL10 protein in liver compared with control mice. Pretreatment with cobalt protoporphyrin 24 h before LPS challenge normalized this effect of ethanol. Adiponectin and induction of HO-1 potently suppressed TLR4-dependent/MyD88-independent cytokine expression in primary Kupffer cells from rats and in mouse liver after chronic ethanol exposure. These data suggest that induction of HO-1 may be a useful therapeutic strategy in alcoholic liver disease.
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Chronic alcohol exposure increases circulating bioactive oxidized phospholipids.
J. Biol. Chem.
PUBLISHED: 05-11-2010
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Ethanol metabolism by liver generates short lived reactive oxygen species that damage liver but also affects distal organs through unknown mechanisms. We hypothesized that dissemination of liver oxidative stress proceeds through release of biologically active oxidized lipids to the circulation. We searched for these by tandem mass spectrometry in plasma of rats fed a Lieber-DeCarli ethanol diet or in patients with established alcoholic liver inflammation, steatohepatitis. We found a severalfold increase in plasma peroxidized phosphatidylcholines, inflammatory and pro-apoptotic oxidatively truncated phospholipids, and platelet-activating factor, a remarkably potent and pleiotropic inflammatory mediator, in rats chronically ingesting ethanol. Circulating peroxidized phospholipids also increased in humans with established steatohepatitis. However, reactive oxygen species generated by liver ethanol catabolism were not directly responsible for circulating oxidized phospholipids because the delayed appearance of these lipids did not correlate with ethanol exposure, hepatic oxidative insult, nor plasma alanine transaminase marking hepatocyte damage. Rather, circulating oxidized lipids correlated with steatohepatitis and tumor necrosis factor-alpha deposition in liver. The organic osmolyte 2-aminoethylsulfonic acid (taurine), which reduces liver endoplasmic reticulum stress and inflammation, even though it is not an antioxidant, abolished liver damage and the increase in circulating oxidized phospholipids. Thus, circulating oxidized phospholipids are markers of developing steatohepatitis temporally distinct from oxidant stress associated with hepatic ethanol catabolism. Previously, circulating markers of the critical transition to pathologic steatohepatitis were unknown. Circulating oxidatively truncated phospholipids are pro-inflammatory and pro-apoptotic mediators with the potential to systemically distribute the effect of chronic ethanol exposure. Suppressing hepatic inflammation, not ethanol catabolism, reduces circulating inflammatory and apoptotic agonists.
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Early growth response-1 attenuates liver injury and promotes hepatoprotection after carbon tetrachloride exposure in mice.
J. Hepatol.
PUBLISHED: 04-23-2010
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Inflammatory gene expression plays a pathological role in acute and chronic hepatic inflammation, yet, inflammation also promotes liver repair by inducing protective mechanisms to limit collateral tissue damage by priming hepatocytes for proliferation. Early growth response (Egr)-1, a transcription factor that regulates inflammatory gene expression, plays a pathological role in many animal models of acute and chronic inflammatory disease. Here, we tested the hypothesis that Egr-1 is beneficial after toxic liver injury.
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Hepatic fibrosis is enhanced and accompanied by robust oval cell activation after chronic carbon tetrachloride administration to Egr-1-deficient mice.
Am. J. Pathol.
PUBLISHED: 04-15-2010
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The transcription factor early growth response (Egr)-1 regulates the expression of genes required for execution of the wound healing response. Multiple cycles of injury, coupled to incomplete wound healing, lead to fibrosis. Therefore, we hypothesized that Egr-1 is required for the development of hepatic fibrosis. To test this hypothesis, we exposed wild-type and egr-1(-/-) mice to acute or chronic carbon tetrachloride (CCl(4)). Acute CCl(4) exposure established a profibrotic milieu in the liver, including activation of hepatic stellate cells as well as expression of type 1 collagen genes and tissue inhibitor of matrix metalloproteinase 1 in both wild-type and egr-1(-/-) mice. This response was exacerbated in egr-1(-/-) mice. After chronic CCl(4) exposure, hepatic fibrosis was established in both genotypes; however, the fibrotic response was profoundly worsened in Egr-1-deficient mice. Importantly, enhanced fibrosis in egr-1(-/-) mice was accompanied by a robust activation of the oval cell response, suggesting more severe liver injury and/or reduced hepatocyte proliferation when compared with wild-type mice. Hepatic expression of genes indicative of oval cell activation, as well as the number of cells expressing A6, a mouse oval cell marker, was greater in egr-1(-/-) mice. Taken together, these data reveal novel roles for Egr-1 as a negative regulator of both CCl(4)-induced hepatic fibrosis and the oval cell response.
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Anti-inflammatory pathways and alcoholic liver disease: role of an adiponectin/interleukin-10/heme oxygenase-1 pathway.
World J. Gastroenterol.
PUBLISHED: 03-19-2010
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The development of alcoholic liver disease (ALD) is a complex process involving both the parenchymal and non-parenchymal cells in the liver. Enhanced inflammation in the liver during ethanol exposure is an important contributor to injury. Kupffer cells, the resident macrophages in liver, are particularly critical to the onset of ethanol-induced liver injury. Chronic ethanol exposure sensitizes Kupffer cells to activation by lipopolysaccharide via Toll-like receptor 4. This sensitization enhances production of inflammatory mediators, such as tumor necrosis factor-alpha and reactive oxygen species, that contribute to hepatocyte dysfunction, necrosis, apoptosis, and fibrosis. Impaired resolution of the inflammatory process probably also contributes to ALD. The resolution of inflammation is an active, highly coordinated response that can potentially be manipulated via therapeutic interventions to treat chronic inflammatory diseases. Recent studies have identified an adiponectin/interleukin-10/heme oxygenase-1 (HO-1) pathway that is profoundly effective in dampening the enhanced activation of innate immune responses in primary cultures of Kupffer cells, as well as in an in vivo mouse model of chronic ethanol feeding. Importantly, induction of HO-1 also reduces ethanol-induced hepatocellular apoptosis in this in vivo model. Based on these data, we hypothesize that the development of therapeutic agents to regulate HO-1 and its downstream targets could be useful in enhancing the resolution of inflammation during ALD and preventing progression of early stages of liver injury.
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Complement and alcoholic liver disease: role of C1q in the pathogenesis of ethanol-induced liver injury in mice.
Gastroenterology
PUBLISHED: 03-08-2010
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Complement is involved in the development of alcoholic liver disease in mice; however, the mechanisms for complement activation during ethanol exposure have not been identified. C1q, the recognition subunit of the first complement component, binds to apoptotic cells, thereby activating the classical complement pathway. Because ethanol exposure increases hepatocellular apoptosis, we hypothesized that ethanol-induced apoptosis would lead to activation of complement via the classical pathway.
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The anti-inflammatory effects of adiponectin are mediated via a heme oxygenase-1-dependent pathway in rat Kupffer cells.
Hepatology
PUBLISHED: 01-07-2010
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Altered expression and activity of immunomodulatory cytokines plays a major role in the pathogenesis of alcoholic liver disease. Chronic ethanol feeding increases the sensitivity of Kupffer cells, the resident hepatic macrophage, to lipopolysaccharide (LPS), leading to increased tumor necrosis factor alpha (TNF-alpha) expression. This sensitization is normalized by treatment of primary cultures of Kupffer cells with adiponectin, an anti-inflammatory adipokine. Here we tested the hypothesis that adiponectin-mediated suppression of LPS signaling in Kupffer cells is mediated via an interleukin-10 (IL-10)/heme oxygenase-1 (HO-1) pathway after chronic ethanol feeding. Knockdown of IL-10 expression in primary cultures of Kupffer cells with small interfering RNA (siRNA) prevented the inhibitory effect of globular adiponectin (gAcrp) on LPS-stimulated TNF-alpha expression. gAcrp increased IL-10 mRNA and protein expression, as well as expression of the IL-10 inducible gene, HO-1; expression was higher in Kupffer cells from ethanol-fed rats compared with pair-fed controls. Although IL-10 receptor surface expression on Kupffer cells was not affected by ethanol feeding, IL-10-mediated phosphorylation of STAT3 and expression of HO-1 was higher in Kupffer cells after ethanol feeding. Inhibition of HO-1 activity, either by treatment with the HO-1 inhibitor zinc protoporphyrin or by siRNA knockdown of HO-1, prevented the inhibitory effect of gAcrp on LPS-stimulated TNF-alpha expression in Kupffer cells. LPS-stimulated TNF-alpha expression in liver was increased in mice after chronic ethanol exposure. When mice were treated with cobalt protoporphyrin to induce HO-1 expression, ethanol-induced sensitivity to LPS was ameliorated. Conclusion: gAcrp prevents LPS-stimulated TNF-alpha expression in Kupffer cells through the activation of the IL-10/STAT3/HO-1 pathway. Kupffer cells from ethanol-fed rats are highly sensitive to the anti-inflammatory effects of gAcrp; this sensitivity is associated with both increased expression and sensitivity to IL-10.
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Current experimental perspectives on the clinical progression of alcoholic liver disease.
Alcohol. Clin. Exp. Res.
PUBLISHED: 07-23-2009
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Chronic alcohol abuse is an important cause of morbidity and mortality throughout the world. Liver damage due to chronic alcohol intoxication initially leads to accumulation of lipids within the liver and with ongoing exposure this condition of steatosis may first progress to an inflammatory stage which leads the way for fibrogenesis and finally cirrhosis of the liver. While the earlier stages of the disease are considered reversible, cirrhotic destruction of the liver architecture beyond certain limits causes irreversible damage of the organ and often represents the basis for cancer development. This review will summarize current knowledge about the molecular mechanisms underlying the different stages of alcoholic liver disease (ALD). Recent observations have led to the identification of new molecular mechanisms and mediators of ALD. For example, plasminogen activator inhibitor 1 was shown to play a central role for steatosis, the anti-inflammatory adipokine, adiponectin profoundly regulates liver macrophage function and excessive hepatic deposition of iron is caused by chronic ethanol intoxication and increases the risk of hepatocellular carcinoma development.
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Taurine supplementation prevents ethanol-induced decrease in serum adiponectin and reduces hepatic steatosis in rats.
Hepatology
PUBLISHED: 03-20-2009
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Chronic ethanol feeding decreases expression of adiponectin by adipocytes and circulating adiponectin. Adiponectin treatment during chronic ethanol feeding prevents liver injury in mice. Chronic ethanol feeding also increases oxidative and endoplasmic reticulum (ER) stress in adipose tissue. Here we tested the hypothesis that supplemental taurine, an amino acid that functions as a chemical chaperone/osmolyte and enhances cellular antioxidant activity, would prevent ethanol-induced decreases in adiponectin expression and attenuate liver injury. Serum adiponectin concentrations decreased as early as 4 to 7 days after feeding rats a 36% ethanol diet. This rapid decrease was associated with increased oxidative, but not ER, stress in subcutaneous adipose tissue. Taurine prevented ethanol-induced oxidative stress and increased inflammatory cytokine expression in adipose tissue. Ethanol feeding also rapidly decreased expression of transcription factors regulating adiponectin expression (CCAAT/enhancer binding protein alpha; peroxisome proliferator-activated receptor alpha/gamma) in subcutaneous adipose tissue. Taurine prevented the ethanol-induced decrease in CCAAT/enhancer binding protein alpha and peroxisome proliferator-activated receptor alpha, normalizing adiponectin messenger (m)RNA and serum adiponectin concentrations. In the liver, taurine prevented ethanol-induced oxidative stress and attenuated tumor necrosis factor alpha expression and steatosis, at least in part, by increasing expression of genes involved in fatty acid oxidation. Conclusion: In subcutaneous adipose tissue, taurine decreased ethanol-induced oxidative stress and cytokine expression, as well as normalized expression of adiponectin mRNA. Taurine prevented ethanol-induced decreases in serum adiponectin; normalized adiponectin was associated with a reduction in hepatic oxidative stress, tumor necrosis factor alpha expression, and steatosis. Taken together, these data demonstrate that taurine has important protective effects against ethanol-induced tissue injury in both adipose and liver tissue.
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Isolevuglandins covalently modify phosphatidylethanolamines in vivo: detection and quantitative analysis of hydroxylactam adducts.
Free Radic. Biol. Med.
PUBLISHED: 03-06-2009
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Levuglandins (LGs) and isolevuglandins (isoLGs, also called "isoketals" or "isoKs") are extraordinarily reactive products of cyclooxygenase- and free radical-induced oxidation of arachidonates. We now report the detection in vivo and quantitative analysis of LG/isoLG adducts that incorporate the amino group of phosphatidylethanolamines (PEs) into LG/isoLG-hydroxylactams. Notably, LC-MS/MS detection of these hydroxylactams is achieved with samples that are an order of magnitude smaller and sample processing is much simpler and less time consuming than required for measuring protein-derived LG/isoLG-lysyl lactams. A key feature of our protocol is treatment of biological phospholipid extracts with phospholipase A(2) to generate mainly 1-palmitoyl-2-lysoPE-hydroxylactams from heterogeneous mixtures of phospholipids with a variety of acyl groups on the 2 position. Over 160% higher mean levels of LG/isoLG-PE-hydroxylactam (P<0.001) were detected in liver from chronic ethanol-fed mice (32.4+/-6.3 ng/g, n=6) compared to controls (12.1+/-1.5 ng/g, n=4), and mean levels in plasma from patients with age-related macular degeneration (5.2+/-0.4 ng/ml, n=15) were elevated approximately 53% (P<0.0001) compared to those of healthy volunteers (3.4+/-0.1 ng/ml, n=15). Just as LG/isoLG-protein adducts provide a dosimeter of oxidative injury, this study suggests that LG/isoLG-PE-hydroxylactams are potential biomarkers for assessing risk for oxidative stress-stimulated diseases.
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Formation of gamma-ketoaldehyde-protein adducts during ethanol-induced liver injury in mice.
Free Radic. Biol. Med.
PUBLISHED: 03-05-2009
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Ethanol metabolism promotes the formation of a variety of reactive aldehydes in the liver. These aldehydes can rapidly form covalent protein adducts. Accumulating evidence indicates that these protein adducts may contribute to ethanol-mediated liver injury. Overproduction of gamma-ketoaldehydes, levuglandins (LGs) and isolevuglandins, is implicated in the pathogenesis of several chronic inflammatory diseases. gamma-Ketoaldehydes can form protein adducts orders of magnitude more quickly than 4-hydroxynonenal (4-HNE) or malondialdehyde. We hypothesized that ethanol-induced oxidative stress in vivo results in overproduction of LGE(2)- and iso[4]LGE(2)-protein adducts in mouse liver. Female C57BL/6 mice were allowed free access to an ethanol-containing diet for up to 39 days or pair-fed control diets. Pathological markers of ethanol-induced hepatic injury including serum alanine aminotransferase, hepatic triglyceride, and CYP2E1 were elevated in response to ethanol feeding. Ethanol-induced formation of iso[4]LGE(2)-, LGE(2)-, and 4-HNE-protein adducts in mouse liver was dependent on both dose and duration of ethanol feeding. Deficiency of cyclooxygenase 1 or 2 did not prevent ethanol-induced iso[4]LGE(2) or LGE(2) adducts in the liver, but adduct formation was reduced in both TNFR1- and CYP2E1-deficient mice. In summary, ethanol feeding enhanced gamma-ketoaldehyde-protein adduct production via a TNFR1/CYP2E1-dependent, but cyclooxygenase-independent, mechanism in mouse liver.
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Exogenous thioredoxin prevents ethanol-induced oxidative damage and apoptosis in mouse liver.
Hepatology
PUBLISHED: 02-11-2009
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Ethanol-induced liver injury is characterized by increased formation of reactive oxygen species (ROS) and inflammatory cytokines, resulting in the development of hepatic steatosis, injury, and cell death by necrosis and apoptosis. Thioredoxin (Trx), a potent antioxidant and antiinflammatory molecule with antiapoptotic properties, protects animals from a number of inflammatory diseases. However, the effects of ethanol on Trx or its role in ethanol-induced liver injury are not known. Female C57BL/6 mice were allowed ad libitum access to a Lieber-deCarli ethanol diet with 5.4% of calories as ethanol for 2 days to acclimate them to the diet, followed by 2 days with 32.4% of calories as ethanol or pair-fed control diet. Hepatic Trx-1 was decreased by ethanol feeding; daily supplementation with recombinant human Trx (rhTrx) prevented this ethanol-induced decrease. Therefore, we tested the hypothesis that administration of rhTrx during ethanol exposure would attenuate ethanol-induced oxidative stress, inflammatory cytokine production, and apoptosis. Mice were treated with a daily intraperitoneal injection of either 5 g/kg of rhTrx or phosphate-buffered saline (PBS). Conclusion: Ethanol feeding increased accumulation of hepatic 4-hydroxynonenal protein adducts, expression of hepatic tumor necrosis factor alpha, and resulted in hepatic steatosis and increased plasma aspartate aminotransferase and alanine aminotransferase. In ethanol-fed mice, treatment with rhTrx reduced 4-hydroxynonenal adduct accumulation, inflammatory cytokine expression, decreased hepatic triglyceride, and improved liver enzyme profiles. Ethanol feeding also increased transferase-mediated dUTP-biotin nick-end labeling-positive cells, caspase-3 activity, and cytokeratin-18 staining in the liver. rhTrx treatment prevented these increases. In summary, rhTrx attenuated ethanol-induced increases in markers of oxidative stress, inflammatory cytokine expression, and apoptosis.
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Hepatic sinusoidal cells in health and disease: update from the 14th International Symposium.
Liver Int.
PUBLISHED: 01-31-2009
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This review aims to give an update of the field of the hepatic sinusoid, supported by references to presentations given at the 14th International Symposium on Cells of the Hepatic Sinusoid (ISCHS2008), which was held in Tromsø, Norway, August 31-September 4, 2008. The subtitle of the symposium, Integrating basic and clinical hepatology, signified the inclusion of both basal and applied clinical results of importance in the field of liver sinusoidal physiology and pathophysiology. Of nearly 50 oral presentations, nine were invited tutorial lectures. The authors of the review have avoided writing a flat summary of the presentations given at ISCHS2008, and instead focused on important novel information. The tutorial presentations have served as a particularly important basis in the preparation of this update. In this review, we have also included references to recent literature that may not have been covered by the ISCHS2008 programme. The sections of this review reflect the scientific programme of the symposium (http://www.ub.uit.no/munin/bitstream/10037/1654/1/book.pdf): 1. Liver sinusoidal endothelial cells. 2. Kupffer cells. 3. Hepatic stellate cells. 4. Immunology. 5. Tumor/metastasis. Symposium abstracts are referred to by a number preceded by the letter A.
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An early complement-dependent and TLR-4-independent phase in the pathogenesis of ethanol-induced liver injury in mice.
Hepatology
PUBLISHED: 01-13-2009
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The innate immune system has been implicated in the pathogenesis of alcoholic liver disease. Although innate immunity is usually considered an early response to injury, previous work implicating innate immunity in ethanol-induced liver injury focuses primarily on long-term ethanol exposure. We investigated the early period of ethanol exposure to determine whether there were temporal associations between activation of innate immune responses and known correlates of liver injury. Female C57BL/6 mice were allowed free access to an ethanol-containing Lieber-DeCarli diet or were pair-fed a control diet. Within 4 days of ethanol exposure, we observed a striking spike in expression of hepatic proinflammatory cytokines-including tumor necrosis factor alpha (TNF-alpha), interleukin-6, and interferon-gamma-prior to hepatic triglyceride accumulation or increased plasma alanine aminotransferase activities, as well as before the induction of cytochrome P450 2E1 or oxidative stress. This early spike in inflammatory cytokines coincided with deposition of C3b-iC3b/C3c (C3b) in the liver. This deposition, resulting from the cleavage of the third component of the complement system (C3), is evidence for activation of complement in response to ethanol. C3(-/-) mice were protected from the early, ethanol-induced increase in hepatic TNF-alpha expression. Ethanol increased C3b deposition in mice deficient in C3a receptor or C5a receptor, as well as in wild-type mice depleted of hepatic macrophages; however, there was no increase in hepatic TNF-alpha in the absence of C3a receptor, C5a receptor, or hepatic macrophages. In contrast, the absence of Toll-like receptor 4 (TLR-4) had no effect on the early, ethanol-induced increase in either C3b or TNF-alpha.
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Inflammation in alcoholic liver disease.
Annu. Rev. Nutr.
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Frank Burr Mallorys landmark observation in 1911 on the histopathology of alcoholic liver disease (ALD) was the first identification of a link between inflammation and ALD. In this review, we summarize recent advances regarding the origins and roles of various inflammatory components in ALD. Metabolism of ethanol generates a number of metabolites, including acetate, reactive oxygen species, acetaldehyde, and epigenetic changes, that can induce inflammatory responses. Alcohol and its metabolites can also initiate and aggravate inflammatory conditions by promoting gut leakiness of microbial products, by sensitizing immune cells to stimulation, and by activating innate immune pathways, such as complement. Chronic alcohol consumption also sensitizes nonimmune cells, e.g., hepatocytes, to inflammatory signals and impairs their ability to respond to protective signals. Based on these advances, a number of inflammatory targets have been identified with potential for therapeutic intervention in ALD, presenting new opportunities and challenges for translational research.
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Inhibition of apoptosis protects mice from ethanol-mediated acceleration of early markers of CCl4 -induced fibrosis but not steatosis or inflammation.
Alcohol. Clin. Exp. Res.
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Correlative evidence indicates that apoptosis is associated with the progression of alcoholic liver disease. If apoptosis contributes to ethanol (EtOH)-induced steatohepatitis and/or fibrosis, then mice deficient in Bid, a key pro-apoptotic Bcl-2 family member, or mice treated with a pan-caspase inhibitor (VX166) should be resistant to EtOH-induced liver injury.
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In developing our video relationships, we compare around 5 million PubMed articles to our library of over 4,500 methods videos. In some cases the language used in the PubMed abstracts makes matching that content to a JoVE video difficult. In other cases, there happens not to be any content in our video library that is relevant to the topic of a given abstract. In these cases, our algorithms are trying their best to display videos with relevant content, which can sometimes result in matched videos with only a slight relation.