Cigarette Smoking Exacerbates Nonalcoholic Fatty Liver Disease in Obese Rats

The prevalence of cigarette smoking (CS) is increased among obese subjects, who are susceptible to develop nonalcoholic fatty liver disease (NAFLD). We investigated the hepatic effects of CS in control and obese rats. Control and obese Zucker rats were divided into smokers and nonsmokers (n = 12 per group). Smoker rats were exposed to 2 cigarettes/day, 5 days/week for 4 weeks. The effects of CS were assessed by biochemical analysis, hepatic histological examination, immunohistochemistry, and gene expression analysis. Phosphorylation of AKT and extracellular signal-regulated kinase (ERK) and quantification of carbonylated proteins were assessed by western blotting. As expected, obese rats showed hypercholesterolemia, insulin resistance, and histological features of NAFLD. Smoking did not modify the lipidic or glucidic serum profiles. Smoking increased alanine aminotransferase serum levels and the degree of liver injury in obese rats, whereas it only induced minor changes in control rats. Importantly, CS increased the histological severity of NAFLD in obese rats. We also explored the potential mechanisms involved in the deleterious effects of CS. Smoking increased the degree of oxidative stress and hepatocellular apoptosis in obese rats, but not in controls. Similarly, smoking increased the hepatic expression of tissue inhibitor of metalloproteinase-1 and procollagen-alpha2(I) in obese rats, but not in controls. Finally, smoking regulated ERK and AKT phosphorylation. The deleterious effects of CS were not observed after a short exposure (5 days). Conclusion: CS causes oxidative stress and worsens the severity of NAFLD in obese rats. Further studies should assess whether this finding also occurs in patients with obesity and NAFLD. (HEPATOLOGY 2010;51:1567-1576.)

The role of nutritional profile in the orexigenic neuropeptide secretion in nonalcoholic fatty liver disease obese adolescents

Background Little progress has been made to identify the central neuroendocrine pathway involved in the energy intake control in nonalcoholic fatty liver disease (NAFLD) patients. Objective To assess the influence of orexigenic neuropeptides in the nutritional aspects of NAFLD obese adolescents submitted to a long-term interdisciplinary approach. Methods Fifty adolescents aged 15-19 years, with body mass index at least 95th percentile, consisting of 25 patients without NAFLD and 25 with NAFLD. The NAFLD diagnosis was determined by ultrasonography. Blood samples were collected to analyze glycemia, hepatic transaminases, and lipid profile. Insulin resistance was estimated by Homeostasis Model Assessment Insulin Resistance Index. Neuropeptide Y (NPY) and agouti related protein concentrations were measured by enzyme-linked immunosorbent assay. Analyses of food intake were made by 3 days recordatory inquiry. Results At baseline conditions, the patients with NAFLD had significantly higher values of body mass, body mass index, visceral fat, triglycerides, VLDL-C, and hepatic transaminases. After the long-term intervention, they presented a significant reduction in these parameters. In both the groups, it was observed a significant decrease in energy intake, macronutrients and dietetic cholesterol. Only the patients with NAFLD presented a positive correlation between the saturated fatty acids intake and the orexigenic neuropeptides NPY and agouti related protein, and carbohydrate with NPY. Indeed, it was observed a positive correlation between energy intake, lipid (%) and saturated fatty acids with visceral fat accumulation. Conclusion Our findings showed an important influence of diet composition in the orexigenic system, being essential consider that the excessive saturated fatty acids intake could be a determinant factor to increase nonalcoholic fatty liver disease. Eur J Gastroenterol Hepatol 22:557-563 (C) 2010 Wolters Kluwer Health | Lippincott Williams & Wilkins.

Treatment of Hepatocellular Carcinoma With Liver Transplantation: A Single-Center Experience From Brazil

Introduction. Orthotopic liver transplantation (OLT) is the treatment of choice of hepatocellular carcinoma (HCC) for patients with cirrhosis, mainly those with early HCC. Herein we have present the clinical characteristics and outcomes of cirrhotic patients with HCC who underwent OLT from cadaveric donors in our institution. Methods. From May 2001 to May 2009, we performed 121 OLT including 24 patients (19.8%) with cirrhosis and HCC within the Milan criteria. In 4 cases, HCC was an incidental finding in the explants. Results. The patients` average age was 55 +/- 10 years, including 82% men. Fifty percent of patients were Child class B or C. The average Model for End Stage Liver Disease for Child A, B, and C categories were 11, 15, and 18, respectively. The HCC diagnosis was made by 2 dynamic images in 16 cases; 1 dynamic image plus alphafetoprotein >400 ng/mL in 4; and 4 by histologic confirmation. Twenty patients received a locoregional treatment before OLT: 6 percutaneous ethanol injection, 9 transarterial chemoembolization, 1 transarterial embolization, and 4 a combination of these modalities. The median follow-up after OLT was 19.7 months (range, 1-51). A vascular invasion was observed in the explant of 1 patient, who developed an HCC recurrence and succumbed at 8 months after OLT. Two further patients, without vascular invasion or satellite tumor displayed tumor recurrences at 7 and 3 months after OLT, and death at 2 and 1 month after the diagnosis. The remaining 25 patients have not shown a tumor recurrence. Conclusion. In the present evaluation, OLT patients with early HCC and no vascular invasion showed satisfactory results and good disease-free survival. Strictly following the Milan criteria for liver transplantation in patients with HCC greatly reduces but does not completely avoid, the chances of tumor recurrence.

Progression of Lipid Peroxidation Measured as Thiobarbituric Acid Reactive Substances, Damage to DNA and Histopathological Changes in the Liver of Rats Subjected to a Methionine-Choline-Deficient Diet

Methionine-choline-deficient diet represents a model for the study of the pathogenesis of steatohepatitis. Male rats were divided into three groups, the first group receiving a control diet and the other two groups receiving a methionine-choline-deficient diet for 1 month (MCD1) and for 2 months (MCD2), respectively. The livers of the animals were collected for the determination of vitamin E, thiobarbituric acid reactive substances (TBARS), GSH concentration, DNA damages, and for histopathological evaluation. The hepatic TBARS and GSH content was higher (P < 0.05) in the groups receiving the experimental diet (MCD1 and MCD2) compared to control diet, and hepatic vitamin E concentration differed (P < 0.05) between the MCD1 and MCD2 groups, with the MCD2 group presenting a lower concentration. Damage to hepatocyte DNA was greater (P < 0.05) in the MCD2 group (262.80 DNA injuries/100 hepatocytes) compared to MCD1 (136.4 DNA injuries/100 hepatocytes) and control diet (115.83 DNA injuries/100 hepatocytes). Liver histopathological evaluation showed that steatosis, present in experimental groups was micro- and macro-vesicular and concentrated around the centrolobular vein, zone 3, with preservation of the portal space. The inflammatory infiltrate was predominantly periductal and the steatosis and inflammatory infiltrate was similar in the MCD1 and MCD2 groups, although the presence of Mallory bodies was greater in the MCD2 group. The study describes the contribution of a methionine-choline-deficient diet to the progression of steatosis, lipid peroxidation and hepatic DNA damage in rats, serving as a point of reflection about the role of these nutrients in the western diet and the elevated non-alcoholic steatohepatitis rates in humans.

Transient disruption of liver gap junctional intercellular communication and induction of apoptosis after administration of 1,4-bis[2-(3,5 dichloropyridyloxy)]benzene in mice

Gap junctional intercellular communication (GJIC) and connexin expression (Cx26 and Cx32) in mouse liver were studied after administration of 4-bis[2-(3,5 dichloropyridyloxy)]benzene (TCPOBOP), a phenobarbital-like enzyme inducer. Female C57BI/6 mice were administered TCPOBOP (5.8 mg/kg BW) and euthanized 0, 24, 48 and 72 hours later. Liver samples were snap frozen, or fixed in formalin, or submitted to GJIC analysis. The proliferating cell nuclear antigen (PCNA) immunohistochemistry and the Western blotting for Cx26 and Cx32 were performed. After 48 and 72 h of drug administration the liver-to-body weight ratio was increased 70% and 117% (p < 0.0001), respectively. There were temporal-dependent alterations in liver histopathology and a significant increase in cell proliferation was noted after 48h and sustained after 72h, though to a lesser extent (p < 0.0001). In addition. TCPOBOP administration induced apoptosis, which appeared to be time-dependent showing statistical significance only after 72h (p < 0.0001). Interestingly, a transient disruption by nearly 50% of GJIC capacity was detected after 48 h of drug ingestion, which recovered after 72 h (p = 0.003). These GJIC changes were due to altered levels of Cx26 and Cx32 in the livers of TCPOBOP-treated mice. We concluded that a single administration of TCPOBOP transiently disrupted the levels of GJIC due to decreased expression of connexins and increased apoptotic cell death in mouse liver. (C) 2009 Elsevier GmbH. All rights reserved.

Liver iron overloading in captive muriquis (Brachyteles spp.)

Background Iron accumulation was investigated qualitatively and quantitatively in the liver of 15 captive Brachyteles spp. Methods Hepatic hemosiderosis index (HHI) was determined as the area percentage of the liver parenchyma occupied by hemosiderin and ferritin deposits, through computerized histomorphometric analysis of Prussian blue-stained histologic sections. Results All studied animals presented liver hemosiderosis, and HHI ranged from 0.2% to 41.7%. There were no significant differences in HHI between muriqui species or genders, and no correlations were detected among HHI and age, time in captivity or body mass. Iron deposits were accompanied by other hepatic disorders. Conclusions This is the first study addressing the occurrence and consequences of iron overloading in the liver of muriquis. We propose that hemosiderosis may act as a contribute factor for the development of hepatic injuries. Further studies are advised to clarify the role of diet in the pathogenesis of hemosiderosis in these atelids.

Neither bovine somatotropin nor growth hormone-releasing factor alters expression of thyroid hormone receptors in liver and mammary tissues

Physiological effects of thyroid hormones are mediated primarily by binding of triiodothyronine to specific nuclear receptors. Organ-specific changes in production of triiodothyronine from its prohormone, thyroxine, have been hypothesized to target the action of thyroid hormones on the mammary gland and play a role in mediating or augmenting a galactopoietic response to bovine somatotropin (bST). Additionally, tissue responsiveness to thyroid hormones may be altered by changes in the number or affinity of nuclear receptors for thyroid hormones. In the present study, effects of bST and bovine growth hormone-releasing factor (bGRF) on thyroid hormone receptors in liver and mammary gland were studied. Lactating Holstein cows received continuous infusions of bST or bGRF for 63 d or served as uninfused controls. Nuclei were isolated from harvested mammary and liver tissues and incubated with [(125)I]-triiodothyronine. Treatments did not alter the capacity or affinity of specific binding sites for triiodothyronine in liver or mammary nuclei. Evaluation of transcript abundance for thyroid hormone receptors showed that isoforms of thyroid hormone receptor or retinoid receptor (which may influence thyroid receptor action) expressed in the mammary gland were not altered by bST or bGRF treatment. Data do not support the hypothesis that administration of bST or bGRF alters sensitivity of mammary tissue by changing expression of thyroid hormone receptors.

Hepatogenous photosensitization associated with liver copper accumulation in buffalos

Four 1.5-year-old, male, Murrah buffalos were maintained during eight months without direct solar exposure during a study of copper toxicosis. Four days after solar exposure, all buffalos presented clinical manifestations consistent with acute photosensitization, including anorexia, apathy, loss of body weight, and generalized cutaneous lesions. Gross lesions were characterized by severe erythema, localized edema, fissures, tissue necrosis, gangrene and crust formation with h serous exudation. Liver copper concentration was evaluated, and cutaneous biopsies were taken when clinical signs were evident. The liver copper concentration before solar exposure was increased in all animals. Histopathologic examination of the skin revealed hepatogenous photosensitization characterized by parakeratotic hyperkeratosis, acantholysis, degeneration of squamous epithelial cells, epidermal necrosis with atrophy of sweat glands, and multifocal superficial and deep dermal edema. These findings suggest that asymptomatic accumulation of copper within the liver might have induced hepatic insufficiency thereby resulting in secondary photosensitization when these buffalos were exposed to sunlight. (C) 2009 Elsevier Ltd. All rights reserved.

Caries-preventive Efficacy and Retention of a Resin-modified Glass lonomer Cement and a Resin-based Fissure Sealant: A 3-year Split-mouth Randomised Clinical Trial

Purpose: This prospective clinical trial compared the retention rate and caries-preventive efficacy of two types of sealant modalities over a 3-year period. Materials and Methods: Using a split-mouth randomised design, 1280 sealants were randomly applied on sound permanent second molars of 320 young patients aged between 12 and 16 years. Half of the teeth (n = 640) were sealed with a resin-modified glass ionomer cement (RMGIC) (Vitremer (TM), 3M ESPE) and the other half (n = 640) with a conventional light-cure, resin-based fissure sealant (LCRB) (Fluoroshield (R), Dentsply Caulk). Teeth were evaluated at baseline, 6-, 12-, 18-, 24-, 30- and 36-month intervals with regard to retention and new caries development. Results: On the sealed occlusal surfaces after 3 years, 5.10% of RMGIC and 91.08% of LCRB sealants were totally intact and 6.37% of RMGIC and 7.65% of LCRB sealants were partially intact. New caries lesions were found in 20.06% of RMGIC sealed occlusal surfaces, compared to 8.91% for LCRB sealants. Conclusions: The findings of the present clinical study suggest that RMGIC should be used only as a transitional sealant that can be applied to newly erupting teeth throughout the eruptive process, whereas LCRB sealants are used to successfully prevent occlusal caries lesions once an effective rubber dam can be achieved. It can be concluded that there are differences between the RMGIC and LCRB sealants over a 3-year period in terms of the retention rate and caries-preventive efficacy. RMGIC can serve as a simple and economic sealing solution, however provisional. Due to its poor retention rate, periodic recalls are necessary, even after 6 months, to eventually replace the lost sealant.

Insulin-like growth factor 2 cDNA cloning and ontogeny of gene expression in the liver of the marsupial brushtail possum (Trichosurus vulpecula)

The cDNA sequence for insulin-like growth factor 2 (IGF-2) was determined from the liver of the marsupial brushtail possum (Trichosurus vulpecula) using reverse transcription followed by polymerase chain reaction (RT-PCR) with gene-specific primers. The 359 bp of possum sequence encompassed the mature peptide, 27 bp of the signal peptide, and 125 bp of the E-peptide. Alignment of the deduced amino acid sequence with those from other species indicated that the mature peptide was 71 amino acids in length, 4 amino acids longer than most other mammals. At both the nucleotide and amino acid levels there was a high degree of sequence identity with IGF-2 from other mammalian and nonmammalian species. Amino acid identity ranged from 94.4% with a variant form of human IGF-2 to 80.3% with zebrafinch IGF-2. Northern analysis revealed that radiolabeled possum IGF-2, cDNA hybridized to multiple transcripts in the liver of both adult possums and 150-day-old pouch young and that the overall level of expression was greater in pouch young. Semiquantitative RT-PCR with total RNA from liver samples of pouch young aged 12 to 150 days postpartum and adults confirmed that IGF-2 gene expression was two to three times more abundant in pouch young than in adults but there was no significant change in the level of expression during pouch life. Unlike other mammalian species, in which there is a decline in levels of liver IGF-2 gene expression around the time of birth, levels in the marsupial brushtail possum remain elevated for at least 150 days after birth. This suggests that the decline in liver IGF-2 expression in marsupials and eutherians occurs at a similar stage of development and may reflect a role for this growth factor during the postnatal growth and development of the marsupial, (C) 2001 Academic Press.

A molecular comparison of T lymphocyte populations infiltrating the liver and circulating in the blood of patients with chronic hepatitis B: Evidence for antigen-driven selection of a public complementarity-determining region 3 (CDR3) motif

Despite a large number of T cells infiltrating the liver of patients with chronic hepatitis B, little is known about their complexity or specificity. To characterize the composition of these T cells involved with the pathogenesis of chronic hepatitis B (CHB), we have studied the clonality of V beta T cell receptor (TCR)-bearing populations in liver tissue by size spectratyping the complementarity-determining region (CDR3) lengths of TCR transcripts. We have also compared the CDR3 profiles of the lymphocytes infiltrating the liver with those circulating in the blood to see whether identical clonotypes may be detected that would indicate a virus-induced expansion in both compartments. Our studies show that in most of the patients examined, the T cell composition of liver infiltrating lymphocytes is highly restricted, with evidence of clonotypic expansions in 4 to 9 TCR V beta subfamilies. In contrast, the blood compartment contains an average of 1 to 3 expansions. This pattern is seen irrespective of the patient's viral load or degree of liver pathology. Although the TCR repertoire profiles between the 2 compartments are generally distinct, there is evidence of some T cell subsets being equally distributed between the blood and the liver. Finally, we provide evidence for a putative public binding motif within the CDR3 region with the sequence G-X-S, which may be involved with hepatitis B virus recognition.

Disposition of naproxen, naproxen acyl glucuronide and its rearrangement isomers in the isolated perfused rat liver

1. An isolated perfused rat liver (IPRL) preparation was used to investigate separately the disposition of the non-steroidal anti-inflammatory drug (NSAID) naproxen (NAP), its reactive acyl glucuronide metabolite (NAG) and a mixture of NAG rearrangement isomers (isoNAG), each at 30 mug NAP equivalents ml(-1) perfusate (n = 4 each group). 2. Following administration to the IPRL, NAP was eliminated slowly in a log-linear manner with an apparent elimination half-life (t(1/2)) of 13.4 +/-4.4 h. No metabolites were detected in perfusate, while NAG was the only metabolite present in bile in measurable amounts (3.9 +/-0.8%, of the dose). Following their administration to the IPRL, both NAG and isoNAG were rapidly hydrolysed (t(1/2) in perfusate=57 +/-3 and 75 +/- 14min respectively). NAG also rearranged to isoNAG in the perfusate. Both NAG and isoNAG were excreted intact in bile (24.6 and 14.8% of the NAG and isoNAG doses, respectively). 3. Covalent NAP-protein adducts in the liver increased as the dose changed from NAP to NAG to isoNAG (0.20 to 0.34 to 0.48% of the doses, respectively). Similarly, formation of covalent NAP-protein adducts in perfusate were greater in isoNAG-dosed perfusions. The comparative results Suggest that isoNAG is a better substrate for adduct formation with liver proteins than NAG.

Relationships between non-occupational cadmium exposure and expression of nine cytochrome P450 forms in human liver and kidney cortex samples

This study was undertaken to assess associations between age, gender, cigarette smoke and non-workplace cadmium exposure, and liver pathology and inter-individual variation in cytochrome P450 (CYP) expression in human tissues. Autopsy specimens of twenty-eight Queensland residents whose ages ranged from 3 to 89 years were analyzed for the presence of nine CYP protein isoforms by immunoblotting. All subjects were Caucasians and their liver cadmium contents ranged from 0.11 to 3.95 kg/g wet weight, while their kidney cadmium contents were in the range of 2 to 63 mug/g wet weight. CYP1A2, CYP2A6, CYP2D6, CYP3A4, and CYP3A5 were detected in liver but not in kidney, and CYP1A1 and CYP1B1 were not found in liver or kidney. Lowered liver CYP2C8/19 protein contents were found to be associated with liver pathology. Importantly, we show elevated levels of CYP2C9 protein to be associated with cadmium accumulation in liver. No mechanism that explains this association is apparent, but there are two possibilities that require further study. One is that variation in CYP2C9 protein levels may be, in part, attributed to an individual's non-workplace exposure to cadmium, or an individual's CYP2C9 genotype may be a risk factor for cadmium accumulation. A positive correlation was found between liver CYP3A4 protein and subject age. Levels of liver CYPIA2 protein, but not other CYP forms, were increased in people more exposed to cigarette smoke, but there was no association between CYPIA2 protein and cadmium. CYP2A6 protein was found in all liver samples and CYP2A6 gene typing indicated the absence of CYP2A6 null allele (CYP2A6(D)) in this sample group, confirming very low prevalence of homozygous CYP2A6(D) in Caucasians. CYP2A6 gene types W/W, WIC, and CIC were not associated with variations in liver microsomal CYP2A6 protein. CYP2D6 protein was absent in all twenty-five kidney samples tested but was detectable in liver samples of all but two subjects, indicating the prevalence of the CYP2D6 null allele (CYP2D6(D)) in this sample group to be about 7%, typical of Caucasian populations. (C) 2001 Elsevier Science Inc. All rights reserved.

Population pharmacokinetics of tacrolimus in children who receive cut-down or full liver transplants

Background. The aim of this study was to investigate the population pharmacokinetics of tacrolimus in pediatric liver transplant recipients and to identify factors that may explain pharmacokinetic variability. Methods. Data were collected retrospectively from 35 children who received oral immunosuppressant therapy with tacrolimus. Maximum likelihood estimates were sought for the typical values of apparent clearance (CL/F) and apparent volume of distribution (V/F) with the program NONMEM. Factors screened for influence on the pharmacokinetic parameters were weight, age, gender, postoperative day, days since commencing tacrolimus therapy, transplant type (whole child liver or cut-down adult liver), liver function tests (bilirubin, alkaline phosphatase [ALP], aspartate aminotransferase [AST], gamma -glutamyl transferase [GGT], alanine aminotransferase [ALT]), creatinine clearance, hematocrit, corticosteroid dose, and concurrent therapy with metabolic inducers and inhibitors of tacrolimus. Results. No clear correlation existed between tacrolimus dosage and blood concentrations (r(2) =0.003). Transplant type, age, and liver function test values were the most important factors (P

Liver transplantation: current status and future prospects

The enormous progress that has been made in liver transplantation over the past two decades has culminated in survival approaching 90% at 12 months. The success of the procedure combined with the widening spectrum of disease processes deemed amenable to liver transplantation has meant that there are too few donors for those awaiting transplantation. This has extrapolated to many patients having such advanced disease by the time a suitable donor liver is available, that they are almost non-transplantable. The immediate options facing the transplant community are to decrease the number of patients listed or to increase the number of living donor transplants. Alternatives to liver transplantation such as hepatocyte transplantation, gene therapy, xenotransplantation and the bioartificial liver are being sought but, at best, are some way from clinical application. It is anticipated that a number of liver diseases that are indications for liver transplantation at this time will have progression arrested or will be cured by medical therapy in the future.