Increased circulating products of lipid peroxidation in patients with alcoholic liver disease.
Aleynik SI. Leo MA. Aleynik MK. Lieber CS.
Alcohol Research and Treatment Center, Bronx Veterans Affairs Medical Center and Mount Sinai School of Medicine, New York 10468, USA.
F2-isoprostanes (F2-IP) and 4-hydroxynonenal (4-HNE), peroxidation products of polyunsaturated fatty acids (PUFA), are considered the most reliable indicators of endogenous lipid peroxidation in vivo. To determine to what extent these are also altered in patients with alcoholic liver disease, plasma free and esterified F2-IP as well as 4-HNE were measured by GC/MS in 49 fasting subjects who underwent diagnostic percutaneous needle biopsies of the liver. Compared to patients with mild steatosis and no fibrosis, free F2-IP and 4-HNE were strikingly increased in individuals with alcoholic hepatitis. There was also a significant but lesser rise of 4-HNE in patients with perivenular fibrosis. An increase of F2-IP was also found in subjects with transition to, or complete, alcoholic cirrhosis, with a comparable trend for 4-HNE. By contrast, in patients who were drinking heavily up to 48 hr before admission, F2-IP were not abnormal, but they increased later (p < 0.005). Contrasting with plasma free F2-IP, esterified F2-IP were not significantly changed with fibrosis. Thus, whereas circulating esterified F2-IP were unchanged in patients with alcoholic liver disease, there was an increase in free F2-IP as well as 4-HNE during recovery from intoxication. The increase was not a result of accompanying hepatitis C but a function of the stage of alcoholic liver injury, possibly reflecting enhanced lipid peroxidation as well as interference with biliary excretion and/or hepatic esterification.
Circulating neutrophils and liver injury in rat models of experimental alcoholic liver disease.
Ross AD. Saldivia V. Oporto B. Carmichael FJ. Cameron R. Israel Y.
Department of Pharmacology, Faculty of Medicine, University of Toronto, Ontario, Canada.
The present study examined the relationship between circulating neutrophils and liver injury in two widely used rat models of chronic ethanol administration. Hematological alterations, liver histopathology, and biochemical indices of liver injury were assessed in rats receiving chronic ethanol by oral liquid diet feeding (Lieber-DeCarli method) or by continuous intragastric infusion (Tsukamoto-French method). Oral administration of ethanol did not affect circulating neutrophil counts, but resulted in minimal liver injury characterized by elevated serum alanine aminotransferase (79%), increased liver mass (15%), and moderate steatosis. In contrast, rats receiving ethanol by continuous intragastric infusion showed an approximately 2-fold increase in circulating neutrophils, and a moderate degree of liver injury, indicated by a 169% elevation of serum alanine aminotransferase and a 2-fold increase in liver mass. Liver biopsies from these rats showed severe steatosis and scattered necrotic hepatocytes, and some neutrophil infiltrates. To determine whether an increase in the number of circulating neutrophils could potentiate liver injury induced by oral ethanol feeding, rats were treated with human recombinant granulocyte colony-stimulating factor at a dose of 100 microg/kg/day (s.c.) for 4 days. Treatment with granulocyte colony-stimulating factor resulted in a 6- to 9-fold increase in circulating neutrophil counts. Nevertheless, this change did not enhance the minor degree of ethanol-induced liver injury in this model. Our results indicate that, whereas neutrophil leukocytosis accompanies more severe manifestations of ethanol hepatotoxicity in rats, this condition per se does not directly induce or exacerbate ethanol-induced liver injury.
Relationship between platelet membrane lipid compositions and platelet aggregability in alcoholic liver disease.
Watanabe M. Shiraishi K. Itakura M. Matsuzaki S.
Third Department of Internal Medicine, Tokai University School of Medicine, Kanagawa, Japan.
We studied the relationship between changes in platelet aggregability and platelet membrane lipid in alcoholic liver disease. The maximal rate of ADP-induced platelet aggregation was significantly increased in the alcoholic liver disease group than in the control group. No significant difference was observed in the maximal rate of collagen-induced platelet aggregation. However, a lag time required for the start of platelet aggregation was significantly shortened in the alcoholic liver disease group, indicating increased platelet aggregability. Results of the platelet aggregation test suggested that alcoholic liver disease patients have their platelet aggregation affected by the abnormality of prostaglandin metabolism. The alcoholic liver disease group was further divided into two subgroups: the hyperaggregation group and the unchanged aggregation group. Both free cholesterol and phospholipid in the platelet membrane were significantly increased in the alcoholic liver disease group. In phospholipid compositions, phosphatidylserine plus phosphatidylinositol were significantly decreased in the alcoholic liver disease group, whereas a significant decrease in phosphatidylserine plus phosphatidylinositol was observed in the hyperaggregation group of alcoholic liver disease. Analysis of fatty acid compositions of platelet membrane showed significantly decreased palmitic acid in the alcoholic group. There was no significant change of arachidonic acid, which directly affects platelet aggregation. Eicosapentaenoic acid significantly decreased in the alcoholic liver disease group, but there was no difference in docosahexaenoic acid. Meanwhile, the thrombogenic index, calculated from the fatty acids of platelet membrane, showed no difference between the alcoholic liver disease group and the control group. However, the thrombogenic index was significantly increased in the hyperaggregation group than in the unchanged aggregation group. These data suggested that platelet aggregation is affected by not only a change in arachidonic acid, but also changes in fatty acid compositions of the platelet membrane.
Influence of alcohol on branched-chain amino acid/tyrosine molar ratio in patients with cirrhosis.
Suzuki H. Mamata Y. Mizuno H. Tominaga T. Suga M. Suemori S. Sato A. Suzuki M.
Department of Internal Medicine, Yokohama City Seibu Hospital, St. Marianna University School of Medicine, Yokohama, Japan.
We investigated whether the reduction of plasma tyrosine in alcoholic liver disease would affect the branched-chain amino acid/tyrosine molar ratio (BTR) measured using an enzymatic assay method in alcoholic cirrhosis. BTR values were higher in patients with compensated and decompensated alcoholic cirrhosis (5.68 +/- 2.29 and 3.28 +/- 0.75) due to reduction of the tyrosine level relative to those in patients with nonalcoholic cirrhosis (3.64 +/- 1.22 and 2.53 +/- 0.99). A decrease in tyrosine level and an increase in BTR value were observed after single ethanol administration to healthy subjects. As significant elevation of serum immunoreactive insulin levels followed elevation of serum glucose levels after alcohol loading, it was thought that insulin accelerated intrahepatic metabolism of aromatic amino acids, resulting in reduction of the tyrosine level. The same mechanism may be applied to tyrosine reduction in patients with alcoholic cirrhosis during heavy drinking.
Correlation of a polymorphism in the interleukin-1 receptor antagonist gene with hepatic fibrosis in Japanese alcoholics.
Takamatsu M. Yamauchi M. Maezawa Y. Ohata M. Saitoh S. Toda G.
First Department of Internal Medicine, The Jikei University School of Medicine, Tokyo, Japan.
The aim of this study was to determine whether there is any association between interleukin-1 receptor antagonist (IL1-Ra) genotype and alcoholic liver disease. The IL1-Ra genotype was assessed in 102 Japanese male alcoholic liver disease patients and 46 healthy subjects by polymerase chain reaction with leukocyte DNA. The distribution of IL1-Ra genotype and the allelic frequencies in Japanese healthy subjects are both significantly different from that previously reported in Caucasians (A1/A1 genotype: 95.7% in Japanese vs. 54.0% in Caucasians, p < 0.001; A1 allele: 97.8% vs. 73.4%, p < 0.001). The frequency of A1 heterozygotes tended to be higher in Japanese alcoholics with fibrosis, compared with those without fibrosis (14.9% vs. 2.9%). Furthermore, within the fibrotic groups, cumulative alcohol intake was significantly lower in A1 heterozygotes than in the A1 homozygotes (877 +/- 118 kg vs. 1369 +/- 90 kg,p < 0.05). In conclusion, a genetic polymorphism in the IL1-Ra gene may influence the risk of developing hepatic fibrosis in Japanese alcoholics. The same study should be conducted in Caucasian patients having more frequency of IL1-Ra A1 heterozygotes.
Immunohistochemical detection of 4-hydroxy-2-nonenal-modified-protein adducts in human alcoholic liver diseases.
Ohhira M. Ohtake T. Matsumoto A. Saito H. Ikuta K. Fujimoto Y. Ono M. Toyokuni S. Kohgo Y.
Third Department of Internal Medicine, Asahikawa Medical College, Nishikagura, Japan.
4-Hydroxy-2-nonenal (HNE) is one of the major components of lipid peroxidation product and has been shown to react with proteins to form HNE-protein adducts. HNE-protein adducts are relatively stable and can be used as a marker of radical-mediated cellular damage. We report herein the immunohistochemical analysis of HNE-protein adducts in human alcoholic liver diseases using a specific monoclonal antibody HNEJ-2. Cytoplasm of hepatocytes and bile duct epithelia was positively stained for HNE-protein adducts, and the nucleus was negligibly stained. The immunohistochemical intensity of hepatocytes was classified into three groups: strong, moderate, and faint staining. Strong staining was found in 43% of alcoholic liver diseases and in 4% of viral liver diseases. Hepatocytes of alcoholic liver diseases contained a higher amount of HNE-protein adducts than those of viral liver diseases, and the difference was statistically significant (p = 0.005; chi2 test). Semiquantitative analysis of the histological intensities of HNE-protein adducts and iron indicated a significant positive correlation (p = 0.084; Spearman's rank correlation). The localization of HNE-protein adducts and iron in hepatocytes appeared to be identical. These data suggested the correlation between HNE-protein adducts and iron. Our results indicate that HNE-protein adducts, a marker of oxidative stress-induced damage, are increased in human alcoholic liver damage, and that hepatic siderosis may act on the production of free radicals.
An enzyme immune assay for serum anti-acetaldehyde adduct antibody using low-density lipoprotein adduct and its significance in alcoholic liver injury.
Nagata N. Nishizaki Y. Watanabe N. Tsuda M. Matsuzaki S.
Third Department of Internal Medicine, School of Medicine, Tokai University, Kanagawa, Japan.
An acetaldehyde (AcH) adduct was prepared using rabbit low-density lipoprotein as carrier proteins. An antibody against this adduct was raised in Watanabe heritable hyperlipidemic rabbits and cross-reacted with human low-density lipoprotein and bovine serum albumin adducts. Using this antibody, serum anti-AcH-adduct antibody levels were measured by a direct ELISA method in 56 Japanese adults (healthy adults and patients with nonalcoholic gastrointestinal diseases, alcoholic liver injury, or alcoholic pancreatitis). The antibody level (mean +/- SD) was 22 +/- 10 microg/ml in healthy adults, 22 +/- 11 microg/ml in nonalcoholic gastrointestinal diseases, and 16 +/- 13 microg/ml in alcoholic pancreatitis. These antibody levels tended to increase with the progression of alcoholic liver injury, starting from fatty liver via hepatitis to cirrhosis, 29 +/- 24 microg/ml in fatty liver, 35 +/- 29 microg/ml in alcoholic hepatitis, and 46 +/- 54 microg/ml in alcoholic cirrhosis. The antibody level in patients taking 100 g or more of ethanol per day tended to be higher, compared with those in people taking less ethanol. A follow-up observation revealed that alcohol abstinence after hospitalization raised serum anti-AcH-adduct antibody level in some patients and kept it constantly low in other patients. The immunohistochemical study using the anti-AcH-adduct antibody revealed the presence of adduct-like substance in hepatocytes of liver biopsy specimens obtained from patients with alcoholic liver disease. The results indicate that the anti-AcH-adduct antibody may be associated with the progress of alcoholic liver diseases.
Hepatitis G virus infection in patients with alcoholic liver disease.
Sobue S. Higashi K. Nakao H. Takahashi Y. Itoh M. Nakajima K.
First Department of Internal Medicine, Medical School, Nagoya City University, Nagoya, Japan.
The recently discovered hepatitis G virus (HGV) is believed to be a single-stranded RNA virus belonging to the Flaviviridae family, similar to hepatitis C virus (HCV), but much remains to be learned about its characteristics and clinical manifestations. Although it has been suggested that alcohol intake might have an effect on liver pathology by promoting the proliferation of HCV, the association between HGV infection and alcohol intake is yet to be elucidated. In the present study, we investigated the prevalence of HGV-RNA and HCV-RNA in 63 patients with alcoholic liver disease, and studied the effects of alcohol on the progression of hepatic damage in HGV-RNA positive patients. Among these 63 patients, 9 (14%) were HGV-RNA-positive and 37 (59%) were HCV-RNA-positive. Seven (78%) of the nine HGV-RNA positive patients were also infected with HCV. The patients showed no significant differences of clinical features in relation to the presence or absence of HGV infection. There were also no differences of liver histology among HCV-RNA-positive patients with or without HGV-RNA. The two patients infected with HGV alone had alcoholic hepatitis and nonspecific reactive hepatitis, respectively. In this study, alcohol seemed to have little influence on the progression of the liver histology in HGV-RNA-positive patients.
Alcohol-related problems in Taiwan with particular emphasis on alcoholic liver diseases.
Liu JD. Leung KW. Wang CK. Liao LY. Wang CS. Chen PH. Chen CC. Yeh EK.
Department of Medicine, Taipei Medical College Hospital, Taiwan.
Socioeconomic development has led to a progressive increase of alcohol consumption in Taiwan, with an accompanying increase in alcohol-related psychiatric problems, traffic accidents, and liver disease. The prevalent rates of alcohol dependence for Han Chinese and Fomosan aborigines were 0.1% and 1%, respectively in 1950. The rate of alcohol dependence increased to 23% for aborigines in 1995. The number of cases of death and serious injuries due to alcohol-related traffic accident has decreased, and the number of fatalities resulting from these accidents has decreased from third to eighth since the inception of a program of random traffic stops with alcohol breath test in 1997. Alcohol liver disease (ALD) was defined as daily alcohol consumption of 60 g, for a duration of longer than 5 years. We classified ALD patients into two groups: (1) those whose average daily consumption of alcohol exceeded 120 g for a duration longer than 15 years (group A); and (2) all other patients (group B). The case records of 33 cases of biopsy-confirmed ALD were obtained for study. The average of daily alcohol consumption in these cases was 160 g. All but one of these patients were male, age ranged from 26 to 69 years, with an average of 43.1. Clinically, ill-defined gastrointestinal symptoms were the most common presentation (61%), and hepatomegaly was the main physical sign (73%). The average mean corpuscular volume values of ALD and non-ALD patients were 102.3 +/- 10.94 and 94.5 +/- 8.1, respectively (p < 0.01). The mean corpuscular volume values of group A and group B were 102.9 +/- 9.7 vs. 96.5 +/- 9.11 (p < 0.05). Result from serum SGOT/SGPT and gamma-glutamyltransferase/alkaline phosphatase for ALD and non-ALD revealed statistically significant differences between these groups. Using the avidin-biotin complex technique, tissue IgA deposition for ALD patients was found to be different from that of non-ALD patients. Ten of 13 ALD patients vs. 2 of 13 non-ALD patients had continuous-form IgA deposition. Histologically, 45.5% of ALD patients had alcoholic cirrhosis, whereas alcoholic hepatitis was present in only 9.1% of patients. Overall, 88% of cases showed various severity of fatty metamorphosis.
Korean status of alcoholics and alcohol-related health problems.
Park SC. Oh SI. Lee MS.
Aging and Physical Culture Research Institute, Seoul National University, Korea.
Recent changes in the socioeconomic status of Korea have caused big differences in alcohol-related social and health problems. The traditional trait of drinking mild fermented beverages with nutritional side dishes and meals has shifted to drinking strongly distilled liquors without any side dishes. Moreover, the alcohol consumption per adult capita of Korea is now 8.1 liters, which parallels the level of other developed countries; it used to be 1.0 liter in 1960 and 7.0 liters in 1980, respectively. But the alcohol consumption per capita of adult males is now 18.4 liters. Consequently, the national incidence of alcohol-related diseases and accidents has rapidly increased. Korean adult males have the highest risk of the incidence of hepatoma. The rate of car accidents caused by drunken driving is about 10-fold higher than in any other developed country.
Hepatic stellate cells and liver retinoid content in alcoholic liver disease in humans.
Hautekeete ML. Dodeman I. Azais-Braesco V. Van den Berg K. Seynaeve C. Geerts A.
Laboratory for Cell Biology and Histology, Free University of Brussels (VUB), Belgium.
Body retinoids are stored in the lipid droplets of hepatic stellate (Ito) cells. In chronic liver disease, the stellate cells differentiate into myofibroblast-like cells, a process whereby they lose their retinoid-containing lipid droplets. We studied the relation between liver retinoid content, the number of lipid droplets per stellate cell, and the number of stellate cells per mm2 in human alcoholic liver disease. Semithin sections of liver biopsies from normal subjects and patients with early (steatosis, inflammation, and mild fibrosis) and late (cirrhosis and cirrhosis with acute alcoholic hepatitis) alcoholic liver disease were morphometrically evaluated. Liver retinoid content was determined by HPLC. In normal patients, liver retinoid content was 901 +/- 213 IU/g of liver (mean +/- SEM). There was a decrease in liver retinoid content in early alcoholic liver disease (409 +/- 50 IU/g) and a further reduction in cirrhosis (153 +/- 50 IU/g). In patients with acute alcoholic hepatitis, retinoid content was strikingly low (5.2 +/- 1.8 IU/g). There was a progressive decrease in the number of stellate cells per mm2 associated with progressive liver damage. We found a fair correlation between the number of stellate cells per mm2 and liver retinoid content in all patient groups (overall correlation: 0.71). In normal subjects, the mean number of lipid droplets per stellate cell was 7.4 +/- 0.7. In patients with early alcoholic liver disease and in patients with alcoholic cirrhosis, this value was increased to 13.6 +/- 0.8 and 10.4 +/- 2.0, respectively. In patients with acute alcoholic hepatitis, only a few lipid droplets were present (4.2 +/- 0.5). There was a good correlation between liver retinoid content and mean number of lipid droplets in normal patients (r = 0.58). In alcoholic cirrhosis, however, correlation was poor (r = 0.34). In early alcoholic liver disease, the correlation was absent (r = 0.004). In conclusion, the major finding of our study is that the correlation between the mean number of lipid droplets per stellate cell and liver retinoid content varies according to the hepatic pathology considered. Marked lipid droplet accumulation occurs in stellate cells in early alcoholic liver disease and, to a lesser extent, in alcoholic cirrhosis, but there is no correlation between the mean number of lipid droplets per stellate cell and liver retinoid content. Therefore, not retinoids but probably lipids are responsible for the accumulation of lipid droplets. We also find that there is a fair correlation between the number of stellate cells per mm2 and liver retinoid content in all patient groups. Finally, we confirm the decrease in hepatic retinoid content that occurs in alcoholic liver disease in humans, even at the early stages of the disease.
A model to examine the validity of the 6-month abstinence criterion for liver transplantation.
Yates WR. Martin M. LaBrecque D. Hillebrand D. Voigt M. Pfab D.
Department of Psychiatry, University of Oklahoma College of Medicine, Tulsa 74129-1077, USA.
Six months of abstinence from alcohol is a commonly used criterion for liver transplantation eligibility for patients with alcoholic cirrhosis. There is limited evidence to document the validity of this criterion with regard to risk of alcoholism relapse. Ninety-one patients with alcoholic cirrhosis were interviewed for relapse risk using the High Risk Alcoholism Relapse (HRAR) Scale. The HRAR model can be used to predict relapse risk independent of duration of sobriety and therefore can be used to examine the validity of the 6 months of abstinence criteria in this clinical population. The two methods demonstrated poor to fair agreement. Agreement was highest with a cutoff allowing a 5% 6-month relapse risk when 79% agreement (c = 0.56) was demonstrated between the two methods. Using the 6-month abstinence criterion alone disallows a significant number of candidates who have a low relapse risk based on their HRAR score. The validity of the 6-month abstinence criterion is supported somewhat by comparison with the HRAR model. However, use of the 6-month abstinence criterion alone forces a significant number of patients with a low relapse risk by HRAR to wait for transplant listing. A relapse risk model based on an estimate of alcoholism severity in addition to duration of sobriety may more accurately select patients who are most likely to benefit from liver transplantation.