The various impacts of the hepatitis C virus core protein upon hepatic oxidative stress after common bile duct ligation and partial hepatectomy

Abstract

Introduction: Although it is uncertain how the hepatitis C virus (HCV) core protein influences hepatic oxidative stress after partial hepatectomy and common bile duct ligation (CBDL) this may be crucial for the prognosis of patients with HCV infection who have undergone hepatic resection, or who have complications due to a biliary tract obstruction.

Materials and methods: A group of double transgenic mice (DTM) that express both the tetracycline transactivator (tTA) and the HCV core, with conditional, acute expression of the HCV core in the context of the mature liver were subjected to 43% partial hepatectomy and CBDL. The levels of thioredoxin-1, thiobarbituric acid reactive substances (TBARS), and 4-hydroxynonenal (4-HNE) were evaluated in liver samples taken 3 days after the operations.

Results: The DTM had significantly higher TBARS levels than mice that were transgenic for only tTA (i.e. single transgenic mice; STM) and non-transgenic mice (NTM) after a sham laparotomy, CBDL and partial hepatectomy. Of the DTM, the TBARS levels were higher in female mice than in males after a sham laparotomy (P = 0.02) and CBDL (P = 0.0001). 4-HNE staining data were compatible with these results. Furthermore, male DTM exhibited higher levels of thioredoxin-1 than female DTM after sham laparotomy (P = 0.012) and CBDL (P = 0.008).

Conclusions: The HCV core increases hepatic oxidative stress in vivo and female DTM are more vulnerable to the oxidative stress caused by acute core expression with, or without, CBDL. The fact that the female DTM had lower thioredoxin-1 levels may account for this observation.     

Overexpression of human adiponectin in transgenic mice results in suppression of fat accumulation and prevention of premature death by high-calorie diet

Adiponectin, a physiologically active polypeptide secreted by adipocytes, shows insulin-sensitizing, anti-inflammatory, and antiatherogenic properties in rodents and humans. To assess the effects of chronic hyperadiponectinemia on metabolic phenotypes, we established three lines of transgenic mice expressing human adiponectin in the liver. When maintained on a high-fat/high-sucrose diet, mice of two lines that had persistent hyperadiponectinemia exhibited significantly decreased weight gain associated with less fat accumulation and smaller adipocytes in both visceral and subcutaneous adipose tissues. Macrophage infiltration in adipose tissue was markedly suppressed in the transgenic mice. Expression levels of adiponectin receptors were not altered in skeletal muscle or liver. Circulating levels of endogenous adiponectin were elevated, whereas fasting glucose, insulin, and leptin levels were reduced compared with control mice. In the hyperadiponectinemic mice daily food intake was not altered, but oxygen consumption was significantly greater, suggesting increased energy expenditure. Moreover, high-calorie diet-induced premature death was almost completely prevented in the hyperadiponectinemic mice in association with attenuated oxidative DNA damage. The transgenic mice also showed longer life span on a conventional low-fat chow. In conclusion, transgenic expression of human adiponectin blocked the excessive fat accumulation and reduced the morbidity and mortality in mice fed a high-calorie diet. These observations may provide new insights into the prevention and therapy of metabolic syndrome in humans.     

Conjugated linoleic acid fails to worsen insulin resistance but induces hepatic steatosis in the presence of leptin in ob/ob mice

Conjugated linoleic acid (CLA) induces insulin resistance preceded by rapid depletion of the adipokines leptin and adiponectin, increased inflammation, and hepatic steatosis in mice. To determine the role of leptin in CLA-mediated insulin resistance and hepatic steatosis, recombinant leptin was coadministered with dietary CLA in ob/ob mice to control leptin levels and to, in effect, negate the leptin depletion effect of CLA. In a 2 x 2 factorial design, 6 week old male ob/ob mice were fed either a control diet or a diet supplemented with CLA and received daily intraperitoneal injections of either leptin or vehicle for 4 weeks. In the absence of leptin, CLA significantly depleted adiponectin and induced insulin resistance, but it did not increase hepatic triglyceride concentrations or adipose inflammation, marked by interleukin-6 and tumor necrosis factor-alpha mRNA expression. Insulin resistance, however, was accompanied by increased macrophage infiltration (F4/80 mRNA) in adipose tissue. In the presence of leptin, CLA depleted adiponectin but did not induce insulin resistance or macrophage infiltration. Despite this, CLA induced hepatic steatosis. In summary, CLA worsened insulin resistance without evidence of inflammation or hepatic steatosis in mice after 4 weeks. In the presence of leptin, CLA failed to worsen insulin resistance but induced hepatic steatosis in ob/ob mice.     

Aldehyde oxidase 1 is highly abundant in hepatic steatosis and is downregulated by adiponectin and fenofibric acid in hepatocytes in vitro

Adiponectin protects the liver from steatosis caused by obesity or alcohol and therefore the influence of adiponectin on human hepatocytes was analyzed. GeneChip experiments indicated that recombinant adiponectin downregulates aldehyde oxidase 1 (AOX1) expression and this was confirmed by real-time RT-PCR and immunoblot. AOX1 is a xenobiotic metabolizing protein and produces reactive oxygen species (ROS), that promote cell damage and fibrogenesis. Adiponectin and fenofibric acid activate peroxisome proliferator-activated receptor-alpha (PPAR-alpha) and both suppress AOX1 protein and this is blocked by the PPAR-alpha antagonist RU486. Obesity is associated with low adiponectin, reduced hepatic PPAR-alpha activity and fatty liver, and AOX1 was found induced in the liver of rats on a high-fat diet when compared to controls. Free fatty acids and leptin, that are elevated in obesity, failed to upregulate AOX1 in vitro. The current data indicate that adiponectin reduces AOX1 by activating PPAR-alpha whereas fatty liver disease is associated with elevated hepatic AOX1. High AOX1 may be associated with higher ROS well described to induce fibrogenesis in liver tissue but may also influence drug metabolism and activity.     

Maintenance of adiponectin attenuates insulin resistance induced by dietary conjugated linoleic acid in mice

Conjugated linoleic acid (CLA) causes insulin resistance and hepatic steatosis in conjunction with depletion of adipokines in some rodent models. Our objective was to determine whether the maintenance of adipokines, mainly leptin and adiponectin, by either removing CLA from diets or using an adiponectin enhancer, rosiglitazone (ROSI), could attenuate CLA-induced insulin resistance. Male C57BL/6 mice were consecutively fed two experimental diets containing 1.5% CLA mixed isomer for 4 weeks followed by a diet without CLA for 4 weeks. CLA significantly depleted adiponectin but not leptin and was accompanied by hepatic steatosis and insulin resistance. These effects were attenuated after switching mice to the diet without CLA along with restoration of adiponectin. To further elucidate the role of adiponectin in CLA-mediated insulin resistance, ROSI was used in a subsequent study in male ob/ob mice fed either control (CON) or CLA diet. ROSI maintained significantly higher adiponectin levels in CON- and CLA-fed mice and prevented the depletion of epididymal adipose tissue and the development of insulin resistance. In conclusion, we show that insulin resistance induced by CLA may be related more to adiponectin depletion than to leptin and that maintaining adiponectin levels alone either by removing CLA or using ROSI can attenuate these effects.     

Adiponectin activation of AMPK disrupts leptin‐mediated hepatic fibrosis via suppressors of cytokine signaling (SOCS‐3)

Adiponectin is an adipocytokine that was recently shown to be anti‐fibrogenic in hepatic fibrosis. Leptin, on the other hand, promotes hepatic fibrosis. The purpose of the present study was to elucidate a mechanism (or mechanisms) whereby adiponectin dampens leptin signaling in activated hepatic stellate cells (HSCs), and prevents excess extracellular matrix production. Activated HSCs, between passages 2 and 5, were cultured and exposed to recombinant human adiponectin and recombinant leptin. Immunoblot analysis for SOCS‐3, TIMP‐1, and the phosphorylated species of Stat3 and adenosine monophosphate‐activated protein kinase (AMPK) were conducted. We also examined MMP‐1 activity by immunosorbant fluorimetric analysis. In HSCs, adiponectin‐induced phosphorylation of AMPK, and subsequently suppressed leptin‐mediated Stat3 phosphorylation and SOCS‐3 induction. Adiponectin also blocked leptin‐stimulated secretion of TIMP‐1, and significantly increased MMP‐1 activity, in vitro. To extend this study, we treated adiponectin knockout mice (Ad?/?) daily with 5 mg/kg recombinant leptin and/or carbon tetrachloride (2 ml/kg) for 6 weeks. Post‐necropsy analysis was performed to examine for inflammation, and histological changes in the Ad?/? and wild‐type mice. There was no significant difference in inflammation, or aminotransferases, between mice receiving carbon tetrachloride and leptin versus carbon tetrachloride alone. As anticipated, the combination of leptin and CCl₄ enhanced hepatic fibrosis in both wild‐type and Ad?/? mice, as estimated by amount of collagen in injured livers, but wild‐type mice had significantly higher levels of SOCS‐3 and significantly lower levels of TIMP‐1 mRNA and protein than did adiponectin KO mice exposed to both CCl₄ and leptin. We therefore conclude that the protective effects of adiponectin against liver fibrosis require AMPK activation, and may occur through inhibition of the Jak‐Stat signal transduction pathway. J. Cell. Biochem. 110: 1195–1207, 2010. Published 2010 Wiley‐Liss, Inc.     

Polychlorinated biphenyl 153 is a diet-dependent obesogen that worsens nonalcoholic fatty liver disease in male C57BL6/J mice

BackgroundPolychlorinated biphenyls (PCBs) are persistent environmental pollutants that are detectable in the serum of all American adults. Amongst PCB congeners, PCB 153 has the highest serum level. PCBs have been dose-dependently associated with obesity, metabolic syndrome and nonalcoholic fatty liver disease (NAFLD) in epidemiological studies.ObjectiveThe purpose of this study is to determine mechanisms by which PCB 153 worsens diet-induced obesity and NAFLD in male mice fed a high-fat diet (HFD).MethodsMale C57BL6/J mice were fed either control or 42% milk fat diet for 12 weeks with or without PCB 153 coexposure (50 mg/kg ip ×4). Glucose tolerance test was performed, and plasma and tissues were obtained at necropsy for measurements of adipocytokine levels, histology and gene expression.ResultsIn control diet-fed mice, addition of PCB 153 had minimal effects on any of the measured parameters. However, PCB 153 treatment in high-fat-fed mice was associated with increased visceral adiposity, hepatic steatosis and plasma adipokines including adiponectin, leptin, resistin and plasminogen activator inhibitor-1 levels. Likewise, coexposure reduced expression of hepatic genes implicated in β-oxidation while increasing the expression of genes associated with lipid biosynthesis. Regardless of diet, PCB 153 had no effect on insulin resistance or tumor necrosis factor alpha levels.ConclusionPCB 153 is an obesogen that exacerbates hepatic steatosis, alters adipocytokines and disrupts normal hepatic lipid metabolism when administered with HFD but not control diet. Because all US adults have been exposed to PCB 153, this particular nutrient–toxicant interaction potentially impacts human obesity/NAFLD.     

Downregulation of ADIPOQ and PPARγ2 Gene Expression in Subcutaneous Adipose Tissue of Obese Adolescents With Hepatic Steatosis

Hepatic steatosis is associated with hypoadiponectinemia. The mechanism(s) resulting in lower serum adiponectin levels in obese adolescents with fatty liver is unknown. In two groups of equally obese adolescents, but discordant for hepatic fat content, we measured adiponectin, leptin, peroxisome proliferator–activated receptor γ 2 (PPARγ2) and tumor necrosis factor‐α (TNFα) gene expression in the abdominal subcutaneous adipose tissue (SAT). Twenty six adolescents with similar degrees of obesity underwent a subcutaneous periumbilical adipose tissue biopsy, in addition to metabolic (oral glucose tolerance test, and hyperinsulinemic—euglycemic clamp), and imaging studies (magnetic resonance imaging (MRI), DEXA). Using quantitative real‐time‐PCR; adiponectin, PPARγ2, TNFα, and leptin mRNA were measured. Based on a hepatic fat content (hepatic fat fraction, HFF) >5.5%, measured by fast MRI, the subjects were divided into low and high HFF group. In addition to the hypoadiponectinemia in the high HFF group, we found that the expression of adiponectin as well as PPARγ2 in the SAT was significantly decreased in this group. No differences were noted for TNFα and leptin plasma or mRNA levels between the groups. An inverse relationship was observed between adiponectin or PPARγ2 expression and hepatic fat content, whereas, adiponectin expression was positively related to PPARγ2 expression. Independent of overall obesity, a reduced expression of adiponectin and PPARγ2 in the abdominal SAT is associated with high liver fat content, as well as with insulin resistance in obese adolescents.     

LXR-alpha/SREBP-1c通路参与HCV NS5A诱导的肝细胞脂质代谢紊乱

肝细胞脂肪变性是丙型肝炎患者的突出病理特征,但丙肝病毒（HCV）诱导脂肪变性的分子机制尚不清楚.为探究HCV非结构蛋白5A（NS5A）参与诱导脂肪变性的可能分子机制,本研究以HCV NS5A转基因小鼠为研究对象,采集3～16月龄NS5A转基因小鼠和同窝非转基因小鼠的肝组织,进行病理学检测,并用气相色谱 质谱（GC-MS）法分析脂质主要成分胆固醇酯的含量.采用RT-PCR法检测肝细胞中与脂质代谢密切相关基因肝X受体（LXR-alpha）、固醇调节元件结合蛋白（SREBP-1c）、脂肪酸合成酶（FAS）、硬脂酰辅酶A去饱和酶1（SCD1）、过氧化物酶体增殖物受体alpha（PPAR-alpha）的mRNA表达水平.结果表明,与同窝非转基因对照小鼠相比,3～5月龄NS5A转基因小鼠的肝组织没有发生显著的病理性变化,但6～16月龄的NS5A转基因小鼠的肝脏发生了显著的脂肪变性（47.1% vs 130%;P=0.003）.与此相一致,胆固醇酯的含量在NS5A转基因小鼠的肝脏中显著升高（P < 0.01）.RT PCR检测结果表明,与对照小鼠相比,14月龄NS5A转基因小鼠肝组织中与脂质代谢相关的基因LXR.alpha、SREBP.1c、FAS、SCD1的mRNA表达水平显著增高（P < 0.05）,而PPAR alpha的表达则没有显著变化（P > 0.05）. 以上结果提示,NS5A在小鼠肝细胞中可能通过调高LXR.alpha/SREBP.1c信号通路相关基因的表达,进而促进脂质重新合成,诱导脂肪变性.     

Subcutaneous obesity is not associated with sympathetic neural activation

We tested the hypothesis that muscle sympathetic nerve activity (MSNA) would not differ in subcutaneously obese (SUBOB) and nonobese (NO) men with similar levels of abdominal visceral fat despite higher plasma leptin concentrations in the former. We further hypothesized that abdominal visceral fat would be the strongest body composition- or regional fat distribution-related correlate of MSNA among these individuals. To accomplish this, we measured MSNA (via microneurography), body composition (via dual-energy X-ray absorptiometry), and abdominal fat distribution (via computed tomography) in 15 NO (body mass index 0.05, respectively) despite approximately 2.6-fold higher (P < 0.05) plasma leptin concentration in the SUBOB men. Furthermore, abdominal visceral fat was the only body composition- or regional fat distribution-related correlate (r = 0.45; P < 0.05) of MSNA in the pooled sample. In addition, abdominal visceral fat was related to MSNA in NO (r = 0.58; P = 0.0239) but not SUBOB (r = 0.39; P = 0.3027) men. Taken together with our previous observations, our findings suggest that the relation between obesity and MSNA is phenotype dependent. The relation between abdominal visceral fat and MSNA was evident in NO but not in SUBOB men and at levels of abdominal visceral fat below the level typically associated with elevated cardiovascular and metabolic disease risk. Our observations do not support an obvious role for leptin in contributing to sympathetic neural activation in human obesity and, in turn, are inconsistent with the concept of selective leptin resistance.     

Adiponectin is inversely associated with intramyocellular and intrahepatic lipids in obese premenopausal women

Adiponectin, an adipokine secreted by adipocytes, exerts beneficial effects on glucose and lipid metabolism and has been found to improve insulin resistance by decreasing triglyceride content in muscle and liver in obese mice. Adiponectin is found in several isoforms and the high-molecular weight (HMW) form has been linked most strongly to the insulin-sensitizing effects. Fat content in skeletal muscle (intramyocellular lipids, IMCL) and liver (intrahepatic lipids, IHL) can be quantified noninvasively using proton magnetic resonance spectroscopy ((1)H-MRS). The purpose of our study was to assess the relationship between HMW adiponectin and measures of glucose homeostasis, IMCL and IHL, and to determine predictors of adiponectin levels. We studied 66 premenopausal women (mean BMI 31.0 ± 6.6 kg/m(2)) who underwent (1)H-MRS of calf muscles and liver for IMCL and IHL, computed tomography (CT) of the abdomen for abdominal fat depots, dual-energy X-ray absorptiometry (DXA) for fat and lean mass assessments, HMW and total adiponectin, fasting lipid profile and an oral glucose tolerance test (homeostasis model assessment of insulin resistance (HOMA(IR)), glucose and insulin area under the curve). There were strong inverse associations between HMW adiponectin and measures of insulin resistance, IMCL and IHL, independent of visceral adipose tissue (VAT) and total body fat. IHL was the strongest predictor of adiponectin and adiponectin was a predictor of HOMA(IR). Our study showed that in premenopausal obese women HMW adiponectin is inversely associated with IMCL and IHL content. This suggests that adiponectin exerts positive effects on insulin sensitivity in obesity by decreasing intracellular triglyceride content in skeletal muscle and liver; it is also possible that our results reflect effects of insulin on adiponectin.     

WKYMVm ameliorates obesity by improving lipid metabolism and leptin signalling

Obesity is a metabolic disorder that results from an imbalance of energy intake and consumption. As low-grade chronic inflammation caused by obesity can lead to various complications, it is important to develop effective treatments against obesity. In this study, we investigate the effects of WKYMVm, a strong anti-inflammatory agent, against obesity. Administration of WKYMVm into high fat diet (HFD)-induced obese mice significantly attenuated body weight gain, food intake and increased insulin sensitivity. HFD-induced hepatic steatosis and adipose tissue hypertrophy were also markedly ameliorated by WKYMVm. During the maturation of adipocytes, WKYMVm improves lipid metabolism by increasing lipolysis, adipogenesis, mitochondrial biogenesis and fat browning. WKYMVm administration also elicited a decrease in leptin levels, but an increase in leptin sensitivity via regulation of hypothalamic endoplasmic reticulum stress and the leptin receptor cascade. Taken together, our results show that WKYMVm ameliorates obesity by improving lipid metabolism and leptin signalling, suggesting that WKYMVm can be a useful molecule for the development of anti-obesity agents.     

Partial leptin deficiency favors diet-induced obesity and related metabolic disorders in mice

Partial leptin deficiency is not uncommon in the general population. We hypothesized that leptin insufficiency could favor obesity, nonalcoholic steatohepatitis (NASH), and other metabolic abnormalities, particularly under high calorie intake. Thus, mice partially deficient in leptin (ob/+) and their wild-type (+/+) littermates were fed for 4 mo with a standard-calorie (SC) or a high-calorie (HC) diet. Some ob/+ mice fed the HC diet were also treated weekly with leptin. Our results showed that, when fed the SC diet, ob/+ mice did not present significant metabolic abnormalities except for elevated levels of plasma adiponectin. Under high-fat feeding, increased body fat mass, hepatic steatosis, higher plasma total cholesterol, and glucose intolerance were observed in +/+ mice, and these abnormalities were further enhanced in ob/+ mice. Furthermore, some metabolic disturbances, such as blunted plasma levels of leptin and adiponectin, reduced UCP1 expression in brown adipose tissue, increased plasma liver enzymes, beta-hydroxybutyrate and triglycerides, and slight insulin resistance, were observed only in ob/+ mice fed the HC diet. Whereas de novo fatty acid synthesis in liver was decreased in +/+ mice fed the HC diet, it was disinhibited in ob/+ mice along with the restoration of the expression of several lipogenic genes. Enhanced expression of several genes involved in fatty acid oxidation was also observed only in ob/+ animals. Leptin supplementation alleviated most of the metabolic abnormalities observed in ob/+ fed the HC diet. Hence, leptin insufficiency could increase the risk of obesity, NASH, glucose intolerance, and hyperlipidemia in a context of calorie overconsumption.     

Garcinia cambogia extract ameliorates visceral adiposity in C57BL/6J mice fed on a high-fat diet

The aim of present study is to evaluate the effects of Garcinia cambogia on the mRNA levels of the various genes involved in adipogenesis, as well as on body weight gain, visceral fat accumulation, and other biochemical markers of obesity in obesity-prone C57BL/6J mice. Consumption of the Garcinia cambogia extract effectively lowered the body weight gain, visceral fat accumulation, blood and hepatic lipid concentrations, and plasma insulin and leptin levels in a high-fat diet (HFD)-induced obesity mouse model. The Garcinia cambogia extract reversed the HFD-induced changes in the expression pattern of such epididymal adipose tissue genes as adipocyte protein aP2 (aP2), sterol regulatory element-binding factor 1c (SREBP1c), peroxisome proliferator-activated receptor gamma2 (PPARgamma2), and CCAT/enhancer-binding protein alpha (C/EBPalpha). These findings suggest that the Garcinia cambogia extract ameliorated HFD-induced obesity, probably by modulating multiple genes associated with adipogenesis, such as aP2, SREBP1c, PPARgamma2, and C/EBPalpha in the visceral fat tissue of mice.     

Adiponectin expression in human epicardial adipose tissue in vivo is lower in patients with coronary artery disease

BACKGROUND: Intra-peritoneal adipose tissue is recognized as a predictor of metabolic syndrome and may contribute to the risk for cardiovascular disease by the production of adipocytokines, including adiponectin. Nevertheless, there is no knowledge on whether other visceral depots of adipose tissue, including the epicardial fat, have any metabolically active role, including production of adiponectin. AIM OF THE STUDY: We sought to evaluate adiponectin protein expression in epicardial adipose tissue in vivo both in patients with severe coronary artery disease (CAD) and in subjects without CAD. METHODS: Twenty-two patients were enrolled for the study. We selected 16 patients who underwent elective coronary artery bypass graft surgery for critical CAD, 5 who underwent surgery for valve replacement and 1 for correction of an interatrial defect. Epicardial adipose tissue biopsy samples were obtained before the initiation of cardiopulmonary bypass. Adiponectin protein level in epicardial adipose tissue was evaluated by Western blotting. RESULTS: Adiponectin protein value, expressed as adiponectin/actin ratio, in epicardial adipose tissue was significantly lower in patients with severe CAD than in those without CAD (1.42 +/- 0.77 vs 2.36 +/- 0.84 p = 0.02, 95% CI 0.64-1.74). CONCLUSIONS: This study showed for the first time that human epicardial adipose tissue expresses adiponectin. Adiponectin expression is significantly lower in epicardial fat isolated from patients with CAD.