Fatty Liver Stage Research Articles

Effects of Fatty Infiltration of the Liver on the Shannon Entropy of Ultrasound Backscattered Signals

This study explored the effects of fatty infiltration on the signal uncertainty of ultrasound backscattered echoes from the liver. Standard ultrasound examinations were performed on 107 volunteers. For each participant, raw ultrasound image data of the right lobe of liver were acquired using a clinical scanner equipped with a 3.5-MHz convex transducer. An algorithmic scheme was proposed for ultrasound B-mode and entropy imaging. Fatty liver stage was evaluated using a sonographic scoring system. Entropy values constructed using the ultrasound radiofrequency (RF) and uncompressed envelope signals (denoted by HR and HE, respectively) as a function of fatty liver stage were analyzed using the Pearson correlation coefficient. Data were expressed as the median and interquartile range (IQR). Receiver operating characteristic (ROC) curve analysis with 95% confidence intervals (CIs) was performed to obtain the area under the ROC curve (AUC). The brightness of the entropy image typically increased as the fatty stage varied from mild to severe. The median value of HR monotonically increased from 4.69 (IQR: 4.60–4.79) to 4.90 (IQR: 4.87–4.92) as the severity of fatty liver increased (r = 0.63, p < 0.0001). Concurrently, the median value of HE increased from 4.80 (IQR: 4.69–4.89) to 5.05 (IQR: 5.02–5.07) (r = 0.69, p < 0.0001). In particular, the AUCs obtained using HE (95% CI) were 0.93 (0.87–0.99), 0.88 (0.82–0.94), and 0.76 (0.65–0.87) for fatty stages ≥mild, ≥moderate, and ≥severe, respectively. The sensitivity, specificity, and accuracy were 93.33%, 83.11%, and 86.00%, respectively (≥mild). Fatty infiltration increases the uncertainty of backscattered signals from livers. Ultrasound entropy imaging has potential for the routine examination of fatty liver disease.

Staging of Fatty Liver Diseases Based on Hierarchical Classification and Feature Fusion for Back-Scan-Converted Ultrasound Images.

Fatty liver disease is progressive and may not cause any symptoms at early stages. This disease is potentially fatal and can cause liver cancer in severe stages. Therefore, diagnosing and staging fatty liver disease in early stages is necessary. In this paper, a novel method is presented to classify normal and fatty liver, as well as discriminate three stages of fatty liver in ultrasound images. This study is performed with 129 subjects including 28 normal, 47 steatosis, 42 fibrosis, and 12 cirrhosis images. The proposed approach uses back-scan conversion of ultrasound sector images and is based on a hierarchical classification. The proposed algorithm is performed in two parts. The first part selects the optimum regions of interest from the focal zone of the back-scan-converted ultrasound images. In the second part, discrimination between normal and fatty liver is performed and then steatosis, fibrosis, and cirrhosis are classified in a hierarchical basis. The wavelet packet transform and gray-level co-occurrence matrix are used to obtain a number of statistical features. A support vector machine classifier is used to discriminate between normal and fatty liver, and stage fatty cases. The results of the proposed scheme clearly illustrate the efficiency of this system with overall accuracy of 94.91% and also specificity of more than 90%.

Up-regulation of fibroblast growth factor 19 and its receptor associates with progression from fatty liver to hepatocellular carcinoma.

BackgroundHuman fibroblast growth factor 19 (FGF19), its receptor (FGFR4) and EpCAM play an important role in cell proliferation, differentiation, motility, and overexpression have been linked to hepatocellular carcinoma (HCC). The aim of this study was to evaluate the FGF19 signals responsible for the progression of HCC arising from fatty liver.ResultsFGF19 level was significantly increased in the HCC patients' serum compared to non-HCC controls. The IHC results demonstrated significant increases of protein expressions of FGF19, FGFR4 and EpCAM in specimens with fatty liver, NASH, cirrhosis, and HCC compared to healthy liver tissue. There was a significant positive correlation between the protein expressions (FGF19, FGFR4, and EpCAM) and histopathologic changes from FL to HCC. Furthermore, FGF19 was positively correlated with FGFR4 and with EpCAM.Materials and MethodsFGF19 protein levels in serum and tissues were determined by ELISA assay. The FGFR4, and EpCAM expression and tissue distribution were further evaluated by immunohistochemical staining in tissue array samples. FGF19, FGFR4 and EpCAM expressions between the different histologic stages of fatty liver steatohepatitis-cirrhosis-HCC carcinogenesis sequence were compared to healthy hepatic tissue.ConclusionsOverexpression of FGF19/FGFR4 significantly correlated with EpCAM as a marker of hepatic cancer stem cells within the fatty liver-steatosis-cirrhosis-HCC sequence.ImpactThis is the first study to elucidate FGF19/FGFR4 signaling in favor of HCC cells developing as indicated by increased EpCAM within the carcinogenesis sequence from fatty liver to hepatocellular carcinoma. Our study has the potential to yield novel and cost effective screening strategies for HCC patients.

Basic investigation on acoustic velocity change imaging method for quantitative assessment of fat content in human liver

Fatty liver is a disease caused by the excess accumulation of fat in the human liver. The early diagnosis of fatty liver is very important, because fatty liver is the major marker linked to metabolic syndrome. We already proposed the ultrasonic velocity change imaging method to diagnose fatty liver by using the fact that the temperature dependence of ultrasonic velocity is different in water and in fat. For the diagonosis of a fatty liver stage, we attempted a feasibility study of the quantitative assessment of the fat content in the human liver using our ultrasonic velocity change imaging method. Experimental results showed that the fat content in the tissue mimic phantom containing lard was determined by its ultrasonic velocity change in the flat temperature region formed by a circular warming ultrasonic transducer with an acoustic lens having an appropriate focal length. By considering the results of our simulation using a thermal diffusion equation, we determined whether this method could be applied to fatty liver assessment under the condition that the tissue had the thermal relaxation effect caused by blood flow.

Early Detection of Nonalcoholic Fatty Liver Disease Utilizing a Novel QUS Method

Non‐alcoholic fatty liver disease (NAFLD) is the most common liver disease in the Western world, affecting 20% of the population. If left untreated, NAFLD may progress into non‐alcoholic steatohepatitis (NASH), end‐stage liver disease or hepatocellular carcinoma. While conventional ultrasound and other imaging techniques can diagnose advanced stages of fatty liver and hepatic cirrhosis, existing techniques are imprecise and not able to detect early stage NAFLD which may lead to a delayed diagnosis of complications. The goal of this study was to utilize the novel non‐invasive quantitative ultrasound (QUS) method to detect and quantify hepatic fat content at stages of steatosis. In a pilot study, 8‐week‐old male C57BL/67 mice were divided into one of three different dietary groups (n=15/group): control (Con) or one of two different atherogenic diets (Ath or Ath‐Casein). The control diet was a casein‐based purified diet (17.2% fat kcal). The Ath diet was a standard chow diet supplemental cocoa butter, cholesterol and sodium cholate (32.4% fat kcal). The Ath‐casein diet was a modification of the control diet supplemented with cocoa butter, sodium cholate and cholesterol (32.3% fat kcal). The livers of each animal received weekly ultrasound scans starting prior to being placed onto one of the three diets until euthanasia. Each week, five animals were euthanized. At euthanasia, serum was collected and the liver was harvested. Half of the liver's lateral left lobe was stained with H&E (3‐μm section). Animals on the Ath‐Casein diet had significantly higher steatosis grading than animals on the Ath diet (P<0.05) and control diet (P<0.001). Lobular inflammation experienced a similar trend with inflammation being significantly increased in the Ath‐Casein (P<0.001) and Ath (P<0.01) diets compared to the control. Portal inflammation was statistically significant in both of the atherogenic diets compared to the control (P<.001). Interestingly, a significant effect of liver cell injury was only visible in the Ath‐Casein group (P<0.01 against Ath, and P<0.0001 against control). Overall, histology revealed that animals on either of the atherogenic diets were in a worse metabolic condition. In vivo and ex vivo ultrasound of the liver display increased attenuation and backscatter in both of the atherogenic diets compared to the control. Total hepatic lipids were evaluated by the Folch method and will be further compared to ultrasound for trends corresponding to steatosis and inflammation in the liver. Total serum and hepatic cholesterol will be measured. Additionally, serum alanine aminotransferase (ALT) and tumor necrosis factor α (TNFα) will be analyzed to determine liver damage and systemic inflammation. The development of a QUS method to identify early stages of NAFLD and NASH would provide the capability to noninvasively quantify and monitor liver condition and test methods for intervention.Support or Funding InformationThis work was supported by NIH R37EB002641

Value of controlled attenuation parameter in diagnosis of fatty liver using FibroScan

To investigate the value of controlled attenuation parameter (CAP) in the diagnosis of fatty liver using FibroScan in patients with chronic liver disease (CLD). A prospective cohort study was performed for the patients with chronic hepatitis B (CHB), chronic hepatitis C (CHC), and non-alcoholic fatty liver disease (NAFLD) who underwent liver pathological examination followed by CAP measurement within 1 week in The Second People's Hospital of Tianjin from February 2013 to May 2014. According to related guidelines, hepatocyte steatosis was classified as S0: <5%, S1: 5%-33%, S2: 34%-66%, or S3: ≥67%. The receiver operating characteristic (ROC) curves were plotted with positive results as the diagnostic criteria, and the optimal cut-off values were determined at the maximum Youden index. Single linear regression and multiple stepwise regression were applied to analyze the influencing factors for CAP. A total of 427 patients were enrolled, consisting of 19 patients (4.4%) with NAFLD, 383 (89.7%) with CHB, and 25 (5.9%) with CHC. The optimal cut-off values for CAP in the diagnosis of steatosis ≥5%, ≥34%, and ≥67% were 230 dB/m, 252 dB/m, and 283 dB/m, respectively, and the areas under the ROC curve were 0.803, 0.942, and 0.938, respectively (Z = 14.194, 28.385, and 16.486, respectively, all P < 0.01). CAP differentiated S0 from S1, S1 from S2, S0 from S2, S0 from S3, and S1 from S3 (Z = 10.109, 10.224, 47.81, 29.917, and 10.999, all P < 0.01), but was not able to differentiate S2 from S3 (Z = 0.656, P = 0.5116). The single linear regression and multiple stepwise regression analyses showed that only body mass index (BMI; B = 4.001, P < 0.01) and hepatic steatosis (B = 33.015, P = 0.000) were correlated with CAP. The coincidence rates between CAP and liver pathological diagnosis were 77.4%, 81.0%, and 96.2% for S0, S3, and ≥S2, respectively. CAP has a good value in the diagnosis of fatty liver in CLD patients, and can well differentiate between all stages of fatty liver except S2 and S3. CAP is influenced by BMI, but is not found to be associated with liver fibrosis, inflammation, liver stiffness measurement, and etiology.

MTHFR 677C/T and 1298A/C mutations and non-alcoholic fatty liver disease

Common genetic mutations encountered in folate metabolism may result in increased homocysteine (Hcy) levels. It has been reported that increased serum Hcy levels may affect the intracellular fat metabolism and may cause enhanced fatty infiltration in the liver resulting in non-alcoholic fatty liver disease (NAFLD). In total, 150 patients diagnosed with FLD by ultrasound examination and 136 healthy control patients that do not have any fatty infiltration in the liver were included in the study. Patients were grouped as mild (n = 88), moderate (n = 38) or severe (n = 24) according to the stage of fatty liver in ultrasound. Serum liver function tests, Hcy, folic acid and vitamin B12 levels of the patients were studied. The genetic MTHFR C677T and A1298C polymorphisms of the patients were also evaluated. Although there was no significant difference in vitamin B12 and folic acid levels, in the severe group, Hcy levels were significantly higher than that of control and mild groups (p<0.001). By contrast, there was no significant difference in heterozygote MTHFR 677C/T and 1298A/C mutations, both MTHFR 677C/T and MTHFR 1298A/C mutations were more common in NAFLD groups compared with the control patients (p<0.001). We have determined increased Hcy levels and increased prevalence of homozygote MTHFR 677C/T and MTHFR 1298A/C mutations in patients with NAFLD compared with healthy controls. Larger studies are warranted to clarify the etiological role of the MTHFR mutations and Hcy levels in FLD.

Relationship between Ultrasound Backscattered Statistics and the Concentration of Fatty Droplets in Livers: An Animal Study

Ultrasound grayscale B-mode imaging is the most frequently used modality for examining fatty liver. Different concentrations and arrangements of fatty droplets in the liver may produce different statistical distributions of ultrasound backscatter signals, which may be treated as a useful clue for assessing the stage of fatty liver. To verify this point, we investigate the relationship between changes in backscattered statistics and the concentration of fatty droplets in the liver. Fatty liver was induced in rats fed a methionine-choline-deficient diet. Livers were excised from rats for in vitro ultrasound scanning using a single-element transducer. The envelopes of the acquired raw ultrasound signals were used for the analysis of the backscattered statistics by ultrasound Nakagami parametric imaging, which has been shown as a reliable tool to model the statistical distribution of ultrasound backscatter signals. Histological analyses and the measurements of triglyceride and cholesterol in the rat liver were conducted for comparison with the Nakagami parameter. Results show that the ultrasound Nakagami parameter has an excellent correlation with the concentration of fatty droplets, demonstrating that ultrasound backscatter statistics depend on the degree of fatty liver in rats.

Relationship Between Ultrasound Backscattered Statistics and the Concentration of Fatty Droplets in Livers: An Animal Study

Ultrasound grayscale B-mode imaging is the most frequently used modality for examining fatty liver. Different concentrations and arrangements of fatty droplets in the liver may produce different statistical distributions of ultrasound backscatter signals, which may be treated as a useful clue for assessing the stage of fatty liver. To verify this point, we investigate the relationship between changes in backscattered statistics and the concentration of fatty droplets in the liver.

Liver proteomics in progressive alcoholic steatosis

Fatty liver is an early stage of alcoholic and nonalcoholic liver disease (ALD and NALD) that progresses to steatohepatitis and other irreversible conditions. In this study, we identified proteins that were differentially expressed in the livers of rats fed 5% ethanol in a Lieber–DeCarli diet daily for 1 and 3months by discovery proteomics (two-dimensional gel electrophoresis and mass spectrometry) and non-parametric modeling (Multivariate Adaptive Regression Splines). Hepatic fatty infiltration was significantly higher in ethanol-fed animals as compared to controls, and more pronounced at 3months of ethanol feeding. Discovery proteomics identified changes in the expression of proteins involved in alcohol, lipid, and amino acid metabolism after ethanol feeding. At 1 and 3months, 12 and 15 different proteins were differentially expressed. Of the identified proteins, down regulation of alcohol dehydrogenase (−1.6) at 1month and up regulation of aldehyde dehydrogenase (2.1) at 3months could be a protective/adaptive mechanism against ethanol toxicity. In addition, betaine-homocysteine S-methyltransferase 2 a protein responsible for methionine metabolism and previously implicated in fatty liver development was significantly up regulated (1.4) at ethanol-induced fatty liver stage (1month) while peroxiredoxin-1 was down regulated (−1.5) at late fatty liver stage (3months). Nonparametric analysis of the protein spots yielded fewer proteins and narrowed the list of possible markers and identified d-dopachrome tautomerase (−1.7, at 3months) as a possible marker for ethanol-induced early steatohepatitis. The observed differential regulation of proteins have potential to serve as biomarker signature for the detection of steatosis and its progression to steatohepatitis once validated in plasma/serum.

Serum Proteomics and Biomarker Discovery Across the Spectrum of Nonalcoholic Fatty Liver Disease

Nonalcoholic fatty liver disease (NAFLD), ranging from relatively benign simple steatosis to progressive nonalcoholic steatohepatitis (NASH) and fibrosis, is an increasingly common chronic liver disease. Liver biopsy is currently the only reliable tool for staging the subtypes of NAFLD; therefore, noninvasive serum biomarkers for evaluation of liver disease and fibrosis are urgently needed. We performed this study to describe changes in the serum proteome and identify biomarker candidates in serum samples from 69 patients with varying stages of NAFLD (simple steatosis, NASH, and NASH with advanced bridging [F3/F4] fibrosis) and 16 obese controls. Using a label-free mass spectrometry-based approach we identified over 1,700 serum proteins with a peptide identification (ID) confidence level of >75%, 605 of which changed significantly between any two patient groups (false discovery rate <5%). Importantly, expression levels of 55 and 15 proteins changed significantly between the simple steatosis and NASH F3/F4 group and the NASH and NASH F3/F4 group, respectively. Classification of proteins with significant changes showed involvement in immune system regulation and inflammation, coagulation, cellular and extracellular matrix structure and function, and roles as carrier proteins in the blood. Further, many of these proteins are synthesized exclusively by the liver and could potentially serve as diagnostic biomarkers for identifying and staging NAFLD. This proteomic analysis reveals important information regarding the pathogenesis/progression of NAFLD and NASH and demonstrates key changes in serum protein expression levels between control subjects and patients with different stages of fatty liver. Future validation of these potential biomarkers is needed such that these proteins may be used in place of liver biopsy to facilitate diagnosis and treatment of patients with NAFLD.

Differential expression of microRNAs in mouse liver under aberrant energy metabolic status

Despite years of effort, exact pathogenesis of nonalcoholic fatty liver disease (NAFLD) remains obscure. To gain an insight into the regulatory roles of microRNAs (miRNAs) in aberrant energy metabolic status and pathogenesis of NAFLD, we analyzed the expression of miRNAs in livers of ob/ob mice, streptozotocin (STZ)-induced type 1 diabetic mice, and normal C57BL/6 mice by miRNA microarray. Compared with normal C57BL/6 mice, ob/ob mice showed upregulation of eight miRNAs and downregulation of four miRNAs in fatty livers. Upregulation of miR-34a and downregulation of miR-122 was found in livers of STZ-induced diabetic mice. These results demonstrate that distinct miRNAs are strongly dysregulated in NAFLD and hyperglycemia. Comparison between miRNA expressions in livers of ob/ob mice and STZ-administered mice further revealed upregulation of four miRNAs and downregulation of two miRNAs in livers of ob/ob mice, indicating that these miRNAs may represent a molecular signature of NAFLD. A distinctive miRNA expression pattern was identified in ob/ob mouse liver, and hierarchical clustering of this pattern could clearly discriminate ob/ob mice from either normal C57BL/6 mice or STZ-administered mice. These findings suggest an important role of miRNAs in hepatic energy metabolism and implicate the participation of miRNAs in the pathophysiological processes of NAFLD.

Hepatic Cytochrome P450 2E1 Level Rather Than Cecal Condition Contributes to Induction of Early Stage of the Alcoholic Liver Damage in Rats

Intestinal condition and ethanol toxicity have been discussed as predictors of alcoholic liver damage. In this study, we investigated the association of hepatic antioxidant enzymes and cecal condition, including intestinal bacteria estimated by terminal restriction fragment length polymorphism (T-RFLP), in the early stage of alcoholic fatty liver. Three liquid isocaloric diets, control (CT) diet, ethanol (ET) diet, or ethanol diet including modified beet fiber (MBF), were prepared and administered to rats for 4 weeks. At 4 weeks, the plasma alanine aminotransferase (ALT) levels in the ET group were higher and that in MBF groups tended to be higher than the CT group, but endotoxin was not detected in the portal vein in any of the samples. The hepatic thiobarbituric acid reactive substances (TBARS) concentration in the MBF group was higher than that in the CT group. Hepatic cytochrome P450 2E1 (CYP2E1) was induced in ethanol-fed rats. The hepatic catalase levels in the ET and MBF groups were lower than that in the CT group. The hepatic superoxide dismutase-2 (SOD-2) level did not differ among the groups. Plasma ALT level and/or hepatic TBARS level correlated positively with CYP2E1 and negatively with catalase level and SOD-2 level. Indicators of cecal condition, including the bacterial population estimated by T-RFLP, may not give any explanation for the hepatic damage. In conclusion, the hepatic CYP2E1 induced by ethanol, rather than the cecal conditions, may influence the early stage of alcoholic hepatic damage in rats; hepatic catalase may provide protective effects.

Ethanol Self‐Administration and Alterations in the Livers of the Cynomolgus Monkey, Macaca fascicularis

Most of the studies of alcoholic liver disease use models in which animals undergo involuntary administration of high amounts of ethanol and consume diets that are often high in polyunsaturated fatty acids. The objectives of this study were (1) to evaluate whether cynomolgus monkeys (Macaca fascicularis) drinking ethanol voluntarily and consuming a diet with moderate amounts of lipid would demonstrate any indices of alcoholic liver disease past the fatty liver stage and (2) to determine whether these alterations were accompanied by oxidative stress. Six adult male and 6 adult female cynomolgus monkeys were allowed to consume ethanol voluntarily for 18 to 19 months. Additional monkeys were maintained on the same consumption protocol, but were not provided with ethanol. During the course of the study, liver biopsy samples were monitored for lipid deposition and inflammation, serum for levels of liver enzymes, and urine for concentrations of the isoprostane (IsoP) metabolite, 2,3-dinor-5,6-dihydro-15-F(2t)-IsoP, a biomarker for oxidative stress. Liver mitochondria were monitored for respiratory control and liver for concentrations of neutral lipids, adenine nucleotides, esterified F(2) isoprostanes, oxidized proteins, 4-hydroxynonenal (HNE)-protein adducts, and protein levels of cytochrome P-450 2E1 and 3A4. Ethanol consumption ranged from 0.9 to 4.05 g/kg/d over the period of the study. Serum levels of aspartate amino transferase were elevated in heavy-consuming animals compared with those in ethanol-naïve or moderate drinkers. Many of the ethanol consumers developed fatty liver and most showed loci of inflammation. Both hepatic energy charge and phosphorylation potential were decreased and NADH-linked respiration was slightly, but significantly depressed in coupled mitochondria as a result of heavy ethanol consumption. The urinary concentrations of 2,3-dinor-5,6-dihydro-15-F(2t)-IsoP increased as high as 33-fold over that observed in ethanol-abstinent animals. Liver cytochrome P-450 2E1 concentrations increased in ethanol consumers, but there were no ethanol-elicited increases in hepatic concentrations of the esterified F(2) isoprostanes, oxidized proteins, or HNE-protein adducts. Our studies show that cynomolgus monkeys undergoing voluntary ethanol consumption for 1.5 years exhibit many of the features observed in the early stages of human alcoholic liver disease. Ethanol-elicited fatty liver, inflammation, and elevated serum aspartate amino transferase were evident with a diet that contained modest amounts of polyunsaturated lipids. The dramatic increases in urinary IsoP demonstrated that the animals were being subjected to significant oxidative stress that correlated with their level of ethanol consumption.

Heritability and genetic loci of fatty liver in familial combined hyperlipidemia

VLDL overproduction, a process that is driven by an excess amount of hepatic fat, is a well-documented feature of familial combined hyperlipidemia (FCHL). The aims of this study were to investigate whether fatty liver, measured with ultrasound and as plasma alanine aminotransferase (ALT) levels, develops against a genetic background in FCHL and to identify chromosomal loci that are linked to these traits. In total, 157 FCHL family members and 20 spouses participated in this study. Radiological evidence of fatty liver was more prevalent not only in FCHL probands (40%) but also in their relatives (35%) compared with spouses (15%) (P < 0.05). Heritability calculations revealed that 20-36% of the variability in ALT levels could be attributed to genetic factors. Nonparametric quantitative trait locus (QTL) analysis revealed three significant (P < 0.001) loci with either the ultrasound or the ALT trait in the male sample: 1q42.3, 7p12-21, and 22p13-q11; none was found in the female sample or the entire group. Of these QTLs, the 7p region was consistent over time, because reanalysis with ALT levels that were determined during a visit 5 years earlier yielded similar results. This study shows that fatty liver is a heritable aspect of FCHL. Replication of particularly the 7p region is awaited.

Diagnosis and management of alcoholic liver disease, a review

Summary Alcohol consumption in the United Kingdom has risen dramatically over the last fifty years and there has been an associated increase in cirrhosis secondary to alcohol. In western society the most common cause of cirrhosis is secondary to alcohol. Alcoholic liver disease has become an increasing financial burden upon the NHS. Ten percent of hospital admissions are thought to relate in part to alcohol. This review examines the pathogenesis of alcoholic liver disease and the three stages of fatty liver, alcoholic hepatitis and cirrhosis. The clinical presentations of alcoholic liver disease and management will be discussed in detail. Evidence based treatments available for patients with alcoholic liver disease and their clinical effectiveness are also summarized. The systemic complications of alcoholic liver disease primarily malnutrition, ascites, spontaneous bacterial peritonitis, hepatic encephalopathy, hepatorenal syndrome and oesophageal varices will be discussed, as well as new approaches to management of these complications.

Modification of the Mitochondrial Proteome in Response to the Stress of Ethanol-dependent Hepatotoxicity

Mitochondria are particularly susceptible to increased formation of reactive oxygen and nitrogen species in the cell that can occur in response to pathological and xenobiotic stimuli. Proteomics can give insights into both mechanism of pathology and adaptation to stress. Herein we report the use of proteomics to evaluate alterations in the levels of mitochondrial proteins following chronic ethanol exposure in an animal model. Forty-three proteins showed differential expression, 13 increased and 30 decreased, as a consequence of chronic ethanol. Of these proteins, 25 were not previously known to be affected by chronic ethanol emphasizing the power of proteomic approaches in revealing global responses to stress. Both nuclear and mitochondrially encoded gene products of the oxidative phosphorylation complexes in mitochondria from ethanol-fed rats were decreased suggesting an assembly defect in this integrated metabolic pathway. Moreover mtDNA damage was increased by ethanol demonstrating that the effects of ethanol consumption extend beyond the proteome to encompass mtDNA. Taken together, we have demonstrated that chronic ethanol consumption extends to a modification of the mitochondrial proteome far broader than realized previously. These data also suggest that the response of mitochondria to stress may not involve non-discriminate changes in the proteome but is restricted to those metabolic pathways that have a direct role in a specific pathology.

Ursodeoxycholate protects against ethanol-induced liver mitochondrial injury

The purpose of this work was to examine whether ursodeoxycholate (UDC), a hydrophilic bile salt, could reduce mitochondrial liver injury from chronic ethanol consumption in rats. Animals were pair-fed liquid diets containing 36% of calories as ethanol or isocaloric carbohydrates. They were randomly assigned into 4 groups of 7 rats each and received a specific treatment for 5 weeks: control diet, ethanol diet, control diet + UDC, and ethanol diet + UDC. Respiratory rates of isolated liver mitochondria were measured using a Clark oxygen electrode with sodium succinate as substrate. Mitochondria from rats chronically fed ethanol demonstrated an impaired ability to produce energy. At the fatty liver stage, the ADP-stimulated respiration (V3) was depressed by 33%, the respiratory control ratio (RC) by 25% and the P O ratio by 15%. In ethanol-fed rats supplemented with UDC, both the rate and efficiency of ATP synthesis via the oxidative phosphorylation were improved: V3 was increased by 35%, P O by 8%. All the respiratory parameters were similar in control group and control + UDC group. On the other hand, the number and size of mitochondria were assessed by electron microscopy and computer-assisted quantitative analysis. The number of mitochondria from ethanol-treated rats was decreased by 29%, and they were enlarged by 74%. Both parameters were normalized to control values by UDC treatment. These studies demontrate that UDC has a protective effect against ethanol-induced mitochondrial injury by improving ATP synthesis and preserving liver mitochondrial morphology. These UDC positive effects may contribute to the observed decrease in fat accumulation and may delay the progression of alcoholic injury to more advanced stages.

Effect of the Type of Beverage and Meat Consumed by Alcoholics with Alcoholic Liver Disease

We analyzed meat products and alcoholic beverage preference in patients with the three stages of alcoholic liver disease (ALD) compared with controls using diet history data. Daily consumption of total alcohol, types of alcoholic beverages, and types of meat and meat products in grams was obtained by dietary history taken from patients with biopsy proven stage of ALD. A strong association was found between the ALD subjects and total alcohol and beer consumption. There was a significant increase in the consumption of total pig products, pork, and offal in the ALD groups compared with controls. There was a significant positive correlation between beer consumption and pork in alcoholic hepatitis, total pork products in alcoholic hepatitis, and cirrhosis and offal in alcoholic hepatitis and cirrhosis. There was no correlation with the fatty liver stage of ALD. The strongest correlation was between beer and total pig products in the alcoholic hepatitis group. Wine consumption was negatively correlated with the consumption of pig products and beer in the alcoholic cirrhosis group. In conclusion, the association of total pig product consumption with cirrhosis mortality in various countries was validated by personal diet history data obtained from ALD patients in a tested clinical microcosm. The results suggest that this association may be modified by the type of alcoholic beverage that is preferentially consumed.

The effect of the prostaglandin analogue - misoprostol on rat liver mitochondria after chronic alcohol feeding

Rats fed ethanol (36% of total calories in a nutritionally adequate liquid diet) for 5 weeks develop functional alterations of hepatic mitochondria and steatosis of the liver. At the fatty liver stage, ADP-stimulated respiration of mitochondria was depressed in ethanol fed rats by 30% (p< 0.001) with glutamate + malate and by 23% (p< 0.001) with succinate as substrates. A similar decrease was noted in the respiratory control ratio (RCR) (34% and 29%, respectively). The total lipid content of the liver increased 2.6 fold (p< 0.001). Mitochondrial dysfunction could be prevented, in part, by the treatment with a synthetic derivative of prostaglandin E 1, misoprostol, at a mean daily dose of 80 μg/kg of body weight. The RCR with glutamate + malate as substrates was improved by 36% (p< 0.05). We conclude that misoprostol attenuates several functional alterations in liver mitochondria during alcohol feeding.