Vicência Mara Rodrigues de Lima

Combination of N‐acetylcysteine and metformin improves histological steatosis and fibrosis in patients with non‐alcoholic steatohepatitis

Aim:  There is no proven medical therapy for the treatment of non‐alcoholic steatohepatitis (NASH). Oxidative stress and insulin resistance are the mechanisms that seem to be mostly involved in its pathogenesis. The aim of our study was to evaluate the efficacy of N‐acetylcysteine (NAC) in combination with metformin (MTF) in improving the aminotransferases and histological parameters (steatosis, inflammation, hepatocellular ballooning, and fibrosis) after 12 months of treatment.

A rodent model of NASH with cirrhosis, oval cell proliferation and hepatocellular carcinoma

BACKGROUND/AIMS Hepatocellular carcinoma (HCC) is a well recognized complication of advanced NASH (non-alcoholic steatohepatitis). We sought to produce a rat model of NASH, cirrhosis and HCC. METHODS Adult Sprague-Dawley rats, weighing 250-300g, were fed a choline-deficient, high trans-fat diet and exposed to DEN in drinking water. After 16 weeks, the animals underwent liver ultrasound (US), sacrifice and assessment by microscopy, immunohistochemistry and transmission electron microscopy (TEM). RESULTS US revealed steatosis and focal lesions in 6 of 7. All had steatohepatitis defined as inflammation, advanced fibrosis and ballooning with Mallory-Denk bodies (MDB) with frank cirrhosis in 6. Areas of more severe injury were associated with anti-CK19 positive ductular reaction. HCC, present in all, were macro-trabecullar or solid with polyhedral cells with foci of steatosis and ballooned cells. CK19 was positive in single or solid nests of oval cells and in neoplastic hepatocytes. TEM showed ballooning with small droplet fat, dilated endoplasmic reticulum and MDB in non-neoplastic hepatocytes and small droplet steatosis in some cancer cells. CONCLUSIONS This model replicated many features of NASH including steatohepatitis with ballooning, fibrosis, cirrhosis and hepatocellular carcinoma. Oval cell proliferation was evident and the presence anti-CK 19 positivity in the cancer suggests oval cell origin of the malignancy.

Liver mitochondrial dysfunction and oxidative stress in the pathogenesis of experimental nonalcoholic fatty liver disease

Oxidative stress and hepatic mitochondria play a role in the pathogenesis of nonalcoholic fatty liver disease. The aim of the present study was to evaluate the role of hepatic mitochondrial dysfunction and oxidative stress in the pathogenesis of the disease. Fatty liver was induced in Wistar rats with a choline-deficient diet (CD; N = 7) or a high-fat diet enriched with PUFAs-omega-3 (H; N = 7) for 4 weeks. The control group (N = 7) was fed a standard diet. Liver mitochondrial oxidation and phosphorylation were measured polarographically and oxidative stress was estimated on the basis of malondialdehyde and glutathione concentrations. Moderate macrovacuolar liver steatosis was observed in the CD group and mild liver steatosis was observed in the periportal area in the H group. There was an increase in the oxygen consumption rate by liver mitochondria in respiratory state 4 (S4) and a decrease in respiratory control rate (RCR) in the CD group (S4: 32.70 +/- 3.35; RCR: 2.55 +/- 0.15 ng atoms of O2 min-1 mg protein-1) when compared to the H and control groups (S4: 23.09 +/- 1.53, 17.04 +/- 2.03, RCR: 3.15 +/- 0.15, 3.68 +/- 0.15 ng atoms of O2 min-1 mg protein-1, respectively), P < 0.05. Hepatic lipoperoxide concentrations were significantly increased and the concentration of reduced glutathione was significantly reduced in the CD group. A choline-deficient diet causes moderate steatosis with disruption of liver mitochondrial function and increased oxidative stress. These data suggest that lipid peroxidation products can impair the flow of electrons along the respiratory chain, causing overreduction of respiratory chain components and enhanced mitochondrial reactive oxygen species. These findings are important in the pathogenesis of nonalcoholic fatty liver disease.

Intake of trans Fatty Acids Causes Nonalcoholic Steatohepatitis and Reduces Adipose Tissue Fat Content

We investigated the effects of dietary trans fatty acids, PUFA, and SFA on body and liver fat content, liver histology, and mRNA of enzymes involved in fatty acid metabolism. LDL receptor knockout weaning male mice were fed for 16 wk with diets containing 40% energy as either trans fatty acids (TRANS), PUFA, or SFA. Afterwards, subcutaneous and epididymal fat were weighed and histological markers of nonalcoholic fatty liver disease (NAFLD) were assessed according to the Histological Scoring System for NAFLD. PPARalpha, PPARgamma, microsomal triglyceride transfer protein (MTP), carnitine palmitoyl transferase 1 (CPT-1), and sterol regulatory element binding protein-1c (SREBP-1c) mRNA were measured by quantitative RT-PCR. Food intake was similar in the 3 groups, although mice fed the TRANS diet gained less weight than those receiving the PUFA diet. Compared with the PUFA- and SFA-fed mice, TRANS-fed mice had greater plasma total cholesterol (TC) and triglyceride (TG) concentrations, less epididymal and subcutaneous fat, larger livers with nonalcoholic steatohepatitis (NASH)-like lesions, and greater liver TC and TG concentrations. Macrosteatosis in TRANS-fed mice was associated with a higher homeostasis model assessment of insulin resistance (HOMA(IR)) index and upregulated mRNA related to hepatic fatty acid synthesis (SREBP-1c and PPARgamma) and to downregulated MTP mRNA. Diet consumption did not alter hepatic mRNA related to fatty acid oxidation (PPARalpha and CPT-1). In conclusion, compared with PUFA- and SFA-fed mice, TRANS-fed mice had less adiposity, impaired glucose tolerance characterized by greater HOMA(IR) index, and NASH-like lesions due to greater hepatic lipogenesis. These results demonstrate the role of trans fatty acid intake on the development of key features of metabolic syndrome.

Association of polymorphisms of glutamate‐cystein ligase and microsomal triglyceride transfer protein genes in non‐alcoholic fatty liver disease

Background and Aims:  Although the metabolic risk factors for non‐alcoholic fatty liver disease (NAFLD) progression have been recognized, the role of genetic susceptibility remains a field to be explored. The aim of this study was to examine the frequency of two polymorphisms in Brazilian patients with biopsy‐proven simple steatosis or non‐alcoholic steatohepatitis (NASH): −493 G/T in the MTP gene, which codes the protein responsible for transferring triglycerides to nascent apolipoprotein B, and −129 C/T in the GCLC gene, which codes the catalytic subunit of glutamate‐cystein ligase in the formation of glutathione.

Effects of hepatitis C virus on cardiovascular risk in infected patients: a comparative study.

The role of hepatitis C virus (HCV) in the pathogenesis of atherosclerosis and cardiovascular events is unclear. The aim of this study was to evaluate the direct effect of HCV on cardiovascular risk and correlate it with pro and anti-inflammatory cytokines in patients with HCV. HCV monoinfected patients, genotype 1, naive, non-obese (BMI<30) and non-diabetics were included and compared to controls (blood donors). Patients with prior diagnosis of cardiovascular diseases, hypertension, chronic renal failure, cancer and chronic use of lipid-lowering drugs or immunosuppressants were excluded. Age, BMI, systolic blood pressure (SBP) and diastolic (DBP), fasting glucose and lipid levels were determined. Serum cytokines (IL-6, IL-10 and TNF-α) and Framingham score were also evaluated. 62 HCV patients, 34 (54.8%) were males and none of them was smoking. The Framingham scores (median and 25th and 75th percentiles) were 12% (6.5-14%), showing an intermediate cardiovascular risk in patients with HCV. There was significant direct correlation between Framingham and total cholesterol (p=0.043) and DBP (p=0.007). HDL-C (p=0.002) was inversely correlated with the Framingham score. HCV patients had higher levels of proinflammatory cytokines (IL-6 and TNF-α) compared to controls (p<0.0001) and the relation of proinflammatory/anti-inflammatory TNF-α/IL10 and IL-6/IL10 were higher in HCV patients (p<0.01). The Framingham score was directly correlated to IL-6 and TNF-α, but differences were not statistically significant. Patients with HCV monoinfected, nonobese, naïve and non diabetic have an intermediate cardiovascular risk, as measured by the Framingham score and high levels of proinflammatory cytokines (IL-6 and TNF).

Prevention and Reversion of Nonalcoholic Steatohepatitis in OB/OB Mice by S-Nitroso-N-Acetylcysteine Treatment

Objective: To evaluate the role oral administration of S-nitroso-N-acetylcysteine (SNAC), a NO donor drug, in the prevention and reversion of NASH in two different animal models. Methods: NASH was induced in male ob/ob mice by methionine-choline deficient (MCD) and high-fat (H) diets. Two animal groups received or not SNAC orally for four weeks since the beginning of the treatment. Two other groups were submitted to MCD and H diets for 60 days receiving SNAC only from the 31st to the 60th day. Results: SNAC administration inhibited the development of NASH in all groups, leading to a marked decrease in macro and microvacuolar steatosis and in hepatic lipid peroxidation in the MCD group. SNAC treatment reversed the development of NASH in animals treated for 60 days with MCD or H diets, which received SNAC only from the 31st to the 60th day. Conclusions: Oral administration of SNAC markedly inhibited and reversed NASH induced by MCD and H diets in ob/ob mice.

Pro- and Anti-inflammatory Cytokines in Steatosis and Steatohepatitis

BackgroundFatty liver disease is a problem in both bariatric patients and in patients with moderate obesity. Tumor necrosis factor (TNF)-α has been frequently measured in nonalcoholic steatohepatitis (NASH) with or without diabetes, but less is known about interleukin (IL)-6 and IL-10.MethodsModerately obese patients (n = 80) with histologically proven steatosis (n = 29) and NASH (n = 51) were recruited. Serum levels of cytokines were documented along with clinical information. The aim was to identify the correlates of such biomolecules in a stable population.ResultsDiabetes tended to be more associated with NASH (52.5% instead of 41.4%, P = 0.015), with no difference of age, gender, or body mass index regarding steatosis. For the entire population, cytokine changes were not significant, including TNF-α and IL-6. In diabetics only, all markers tended to diminish with NASH, especially IL-10 (P = 0.000). IL-10 correlated with homeostatic model assessment index (P = 0.000) and other variables of glucose homeostasis in diabetes, thus representing a major marker of the disease.Conclusions(1) Generally inconsistent changes in pro- and anti-inflammatory cytokines occurred when NASH was globally compared to steatosis. (2) In contrast, downregulation of IL-6 and IL-10 was perceived in diabetics with NASH. (3) Arterial hypertension did not play a role in these circumstances. (4) IL-10 maintained strong correlations with glucose metabolism indices. (5) TNF-α could not be incriminated for progressive liver damage, as values failed to increase in NASH. (6) Investigations of IL-10 and other counterregulatory cytokines are lacking in this context and deserve further studies.

Yo Jyo Hen Shi Ko, a novel Chinese herbal, prevents nonalcoholic steatohepatitis in ob/ob mice fed a high fat or methionine–choline-deficient diet

Background: Oxidative stress plays a role in the pathogenesis of nonalcoholic steatohepatitis (NASH). Yo Jyo Hen Shi Ko (YHK) is a complex compound purported to reduce reactive oxygen species (ROS) by blocking the propagation of radical‐induced reactions. The aim of this study was to evaluate the role of the effect of YHK in experimental NASH.

Decreased immunoexpression of survivin could be a potential marker in human non-alcoholic fatty liver disease progression?

Background/aim: Regulation of apoptosis in non‐alcoholic fatty liver disease (NAFLD) has been a theme of growing debate. Although no other study assessed the role of survivin in NAFLD, its expression has been reported in hepatic carcinogenesis because of other aetiological factors with relevant discrepancies. The aim of this study was to assess the pattern of survivin immunoexpression by tissue microarray along the whole spectrum of NAFLD, including non‐alcoholic steatohepatitis (NASH)‐related hepatocelular carcinoma (HCC).