Lymphocyte Subsets Studied with Monoclonal Antibodies in Liver Tissues of Patients with Alcoholic Liver Disease

In order to determine the nature of T lymphocytes accumulating in hepatic tissues of patients with alcoholic liver disease, we phenotyped these cells in situ, using monoclonal antibodies to lymphocyte surface antigens and the avidin-biotin-peroxidase complex (ABC) technique. In portal areas of cirrhotic livers, a significant increase (p less than 0.001) in T lymphocytes was observed as compared to livers showing only fatty changes and normal liver tissues. The T cells accumulating in cirrhotic livers comprised T helper-inducer (T4) and T suppressor-cytotoxic (T8) subsets with the T4/T8 ratio of 1.4 +/- 0.5 (SD, n = 14). In contrast, the T lymphocytes in the cellular projections which extended into the parenchyma consisted mostly of suppressor-cytotoxic cells. The relative enrichment in T8+ lymphocytes among the cells invading the parenchyma suggests that they may be important in the mediation of a cytotoxic injury of hepatocytes in advanced alcoholic liver disease and in regulating an immunologic challenge possibly initiated in response to alcoholic hyaline.     

Antibodies to Cytokeratin Filaments in Patients with Alcoholic Liver Disease

Previous studies have suggested that cytoskeletal elements in an altered form in vivo (alcoholic hyalin , AH) are targets for an autoimmune reaction in alcoholic liver disease (ALD). In this study we assayed autoantibodies to cytoskeletal cytoskeletal cytokeratin filaments, intermediate filaments of epithelial cells, by indirect immunofluorescence technique (IIF) using cultured human amnion epithelial cells and PtK 2 cells as targets. Sera from 20 patients with ALD, 38 patients with urogenital cancer, 22 patients with renal diseases, and from 18 healthy controls were studied. Patients with ALD had a significantly increased prevalence of cytokeratin filament autoantibodies using both PtK 2 cells (69%) and human amnion epithelial cells (69%) as substrate, as compared to other groups. The antibodies were mainly of IgA and IgM class and could be demonstrated in all forms of ALD. Interestingly, cytoskeleton antibodies have previously been observed in diseases of viral and "autoimmune" origin.     

Impaired Accommodation of Proximal Stomach in Patients with Alcoholic Liver Cirrhosis

Background: Impaired gastric emptying has previously been detected by ultrasonography in cirrhotic patients, and the role of the type of meal has also been discussed. While these earlier studies dealt with the distal part of the stomach, the aim of our study was to examine the effects of three different types of meal on the proximal stomach in cirrhotic patients. Methods: The proximal stomach was examined by ultrasonography in 15 healthy volunteers and in 21 alcoholic cirrhotic patients. The subjects received a liquid meal with a low calorie content and two different semisolid test meals with a low calorie content or high calorie and fat contents. The proximal gastric size was assessed by ultrasonography in a sagittal area and a frontal diameter. On the basis of assessment of the autonomic nervous function, the cirrhotic patients were divided into two groups: autonomic neuropathy positive and autonomic neuropathy negative. Results: The postcibal gastric size immediately after ingestion of the liquid test meal was significantly lower in the cirrhotic patients than in the healthy controls. In the healthy volunteers, the measures of the proximal gastric size were significantly higher than in either group of cirrhotic patients at t 0 , and at 10, 20 or 30 min after ingestion of a semisolid test meal with low calorie and fat contents. The proximal gastric sizes in the three groups of investigated subjects did not differ when the meal with high fat and calorie contents was tested. When the liquid meal was administered, the proximal gastric size was significantly lower in the cirrhotic patients with autonomic neuropathy. A significant intragroup difference was not observed when the semisolid meals were tested. Conclusions: This study reveals an impairment of the proximal stomach in alcoholic cirrhotic patients. The low calorie liquid meal distinguishes between the two groups of cirrhotic patients and healthy controls.     

Serum Ubiquitin Levels in Patients With Alcoholic Liver Disease

Serum concentrations of free ubiquitin and multiubiquitin chain as determined by immunoassays were compared between 10 healthy subjects, and 11 patients with alcoholic hepatic fibrosis, 10 with alcoholic cirrhosis, and 6 with viral liver cirrhosis. All measurements of multiubiquitin chains were expressed in terms of a standard multiubiquitin chain reference preparation 1. Serum concentrations (mean ± SD) of free ubiquitin and multiubiquitin chains were significantly higher in patients with alcoholic cirrhosis (63.5 ± 33.7 ng/ml and 7.5 ± 4.6 ng/ml) than in the normal subjects (29.6 ± 6.6 ng/ml, p < 0.05 and 4.1 ±1.7 ng/ml, p < 0.05), and those with alcoholic hepatic fibrosis (34.8 ± 16.3 ng/ml, p < 0.05 and 3.0 ± 0.7 ng/ml, p < 0.05) and viral liver cirrhosis (28.8 ± 7.5 ng/ml, p < 0.05 and 4.2 ± 1.3 ng/ml, p < 0.05). Serum levels of both forms of ubiquitin in six patients with alcoholic cirrhosis showed a tendency to decline after 3 months of abstinence. In a total of 14 patients with alcoholic liver damage, 11 with brain atrophy had significantly higher serum levels of both ubiquitin forms than did three patients without brain atrophy ( p < 0.05). No correlation was seen between serum concentrations of either form of ubiquitin and liver function test results in the patients with alcoholic liver damage. However, serum levels of both forms of ubiquitin levels correlated significantly with cumulative alcohol intake ( p < 0.05). A significant correlation ( p < 0.05) also was observed between serum levels of multiubiquitin chains and mean corpuscular volume, a marker of alcohol consumption. These results suggest that the serum concentrations of ubiquitin, especially multiubiquitin chain is a good marker for the diagnosis of alcoholic cirrhosis.     

Treatment of Alcohol Use Disorder in Patients with Alcoholic Liver Disease

Alcohol is a leading cause of liver disease worldwide. Although alcohol abstinence is the crucial therapeutic goal for patients with alcoholic liver disease, these patients have less access to psychosocial, behavioral, and/or pharmacologic treatments for alcohol use disorder. Psychosocial and behavioral therapies include 12-step facilitation, brief interventions, cognitive behavioral therapy, and motivational enhancement therapy. In addition to medications approved by the US Food and Drug Administration for alcohol use disorder (disulfiram, naltrexone, and acamprosate), recent efforts to identify potential new treatments have yielded promising candidate pharmacotherapies. Finally, more efforts are needed to integrate treatments across disciplines toward patient-centered approaches in the management of patients with alcohol use disorder and alcoholic liver disease.     

Alcoholic and Non-alcoholic Liver Disease in Relation to Alcohol Consumption in Scotland, 1978–84.

A common strategy in epidemiological studies linking alcohol consumption in the general population with liver cirrhosis mortality is to use non-specific cirrhosis mortality rates in which alcoholic and non-alcoholic causes of death are not distinguished. Evidence is presented from Scottish mortality data for 1979 to 1984 that the two forms of cirrhosis have quite different epidemiological profiles. Similar findings emerge from morbidity data. The two forms of disease should be distinguished in future studies in which liver cirrhosis is used as a proxy for consumption, despite the manifest shortcomings of currently available data.     

Biotinidase Activity in Patients with Liver Disease

To investigate whether biotinidase deficiency may occur in liver disease, we determined biotinidase activity, biotin levels, and organic acids in patients with liver disease. Serum biotinidase activity in patients with liver disease (2.63 1.40 nmol/min/ml) was significantly lower than in the control group (5.43 1.06 nmol/min/ml). Serum biotinidase activity in decompensated liver cirrhosis (LC) and hepatoma was significantly lower than in acute viral hepatitis (AVH), chronic viral hepatitis (CVH), and compensated LC. The mean serum level of biotin in decompensated LC (1.8 0.6 μg/ml) and hepatoma (1.7 0.8μg/ml) was significantly lower than in the control group (2.5 1.0 μg/ml), and urinary excretion of biotin was increased in patients with liver disease, particularly in decompensated LC. Biotinidase activity correlated positively with serum biotin level and correlated negatively with urinary biotin level. Moreover, in four of five patients with severe liver disease the excretion of propionate, lactate, and 3-hydroxybutyrate decreased after biotin supplementation. The data for patients with severe liver disease so resembled those for late-onset multiple carboxylase deficiency that biotinidase deficiency is likely in patients with severe liver disease.     

Characteristics of Serum Hyaluronate Concentrations in Patients with Alcoholic Liver Disease

It has been reported that serum hyaluronate [hyaluronic acid (HA)] concentrations are increased in liver diseases, especially in alcoholic liver disease (ALD). However, the characteristics of serum HA concentration in patients with ALD have not been studied. In this study, first, we measured serum HA concentrations in patients with different stages of both ALD and non-ALD to clarify the characteristics of serum HA concentration in patients with ALD. Second, we measured serum HA concentrations in patients with ALD sequentially after abstinence. We also measured serum HA concentrations in patients with chronic type C hepatitis before and after treatment with interferon. Finally, we analyzed the relationship between serum HA concentrations and the contents of type IV collagen and laminin in the livers of both ALD and non-ALD patients. Serum HA concentrations in liver disease were higher than the cut-off value, and increased significantly ( p < 0.001) in parallel with the progression of hepatic fibrosis in both ALD and non-ALD patients. Serum HA concentrations in patients actively drinking with ALD were significantly higher ( p < 0.001) than those in non-ALD. After 4 weeks of abstinence, these concentrations fell to the levels of non-ALD. Although serum ALT levels were decreased in 80% of patients treated with interferon, serum HA concentrations were not changed or increased. A significant correlation between serum HA concentrations and hepatic type IV collagen and laminin content was present in ALD, but not in non-ALD. These results clearly suggest that the increase of serum HA concentrations in ALD may be associated with not only hepatic fibrosis, but also alcohol drinking.     

Genetic and Epigenetic Profile of Patients With Alcoholic Liver Disease

Alcoholic liver disease (ALD) is a definition encompassing a spectrum of disorders ranging from simple steatosis to cirrhosis and hepatocellular carcinoma. Excessive alcohol consumption triggers a series of metabolic reactions that affect the liver by inducing lipogenesis, increasing oxidative stress, and causing abnormal inflammatory responses. The metabolic pathways regulating lipids, reactive oxygen species (ROS), and immune system are closely related and in some cases cross-regulate each other. Therefore, it must be taken into account that major genetic and epigenetic abnormalities affecting enzymes involved in one of such pathways can play a pivotal role in ALD pathogenesis. However, recent studies have pointed out how a significant predisposition can also be determined by minor variants, such as relatively common polymorphisms, epigenetic modifications, and microRNA abnormalities. Genetic and epigenetic factors can also affect the progression of liver diseases, promoting fibrogenesis, cirrhosis, and ultimately hepatocellular carcinoma. It is noteworthy that some of these factors, such as some of the cytokines involved in the abnormal inflammatory responses, are shared with non-alcoholic liver disease, while other factors are unique to ALD. The study of the genetic and epigenetic components involved in the liver damages caused by alcohol is crucial to identify individuals with high risk of developing ALD, design personalized protocols for prevention and/or treatment, and select the best molecular targets for new therapies.     

Serum and Ascitic Levels of Soluble Intercellular Adhesion Molecule-1 in Patients with Alcoholic Liver Cirrhosis: Relation to Biochemical Markers of Disease Activity and Alcohol Intake

The overexpression of intercellular adhesion molecule-1 (ICAM-1) has been shown to be involved in the pathogenesis of various necro-inflammatory diseases, including alcoholic hepatitis. Shedding of this molecule from cell surfaces results in a circulating form, soluble CAM-1 (SICAM-1). In this work, the serum and ascitic concentrations of SICAM-1 were studied in relation to clinical and laboratory data in patients with alcoholic liver cirrhosis of different disease activities. Elevated circulating concentrations of this adhesion molecule were found in all cirrhotic patients, the highest in those with superimposed sewere alcoholic hepatitis, and the levels in regularly drinking cirrhotics without severe alcoholic hepatitis were likewise significantly higher than in those who had stopped drinking. The serum SICAM-1 concentration was best related to the serum AST activity, and also exhibited significant correlations with the pro-thrombin activity, serum bilirubin, albumin, peripheral leukocyte count, Maddrey's discriminant function value, Child grading, and antecedent alcohol consumption. Multivariate regression analysis revealed that the serum AST and prothrombin activities were independent predictors of the circulating SICAM-1 concentration. The concentration of SICAM-1 in the uninfected ascitic fluid of cirrhotics was about seven times lower than that in the serum; the ratio of its ascitic and serum levels was lower than that of the ascitic and serum total protein concentrations. These data contradict a significant in-traperitoneal production of the molecule. It is concluded that the serum SICAM-1 level may be useful as a marker for the current disease activity (the severity of underlying acute necroinflammatory reactions) in alcoholic liver cirrhosis.     

T Lymphocytes from Alcoholic Cirrhotic Patients Show Normal Interleukin-2 Production but a Defective Proliferative Response to Polyclonal Mitogens

T lymphocyte proliferation is a complex process involving intra- and extracellular molecules. T cell activation was studied in T lymphocytes from patients with alcoholic cirrhosis. A defective phytohemagglutinin (PHA)-induced T cell mitogenesis was observed in 60% of these patients. Likewise, their blastogenic response to anti-CD3 was also depressed ( p < 0.05). However, the DNA synthesis induced by stimulation with phorbol esters (12-O-tetradecanoil-phorbol-13-acetate) + ionomycin was normal ( p > 0.05). These alterations cannot be ascribed either to decreased interleukin-2 synthesis or to a defective interleukin-2 receptor expression after cellular activation. Moreover, supplementation of the PHA-stimulated T cell cultures with saturant concentrations of recombinant interleukin-2 did not normalize the hypoproliferative response of T cells from alcoholic cirrhotic patients. These results provide evidence that a generalized alteration in the interactions between either mitogens or interleukin-2 and their receptors can explain the T lymphocyte-defective blastogenesis found in patients with alcoholic cirrhosis.     

Increased Circulating Products of Lipid Peroxidation in Patients with Alcoholic Liver Disease

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.
Alcohol, Liver Disease, 4-Hydroxynonenal, F₂-lsoprostanes, Lipid Peroxidation.     

The Duffy Blood System and Alcoholic Liver Disease in Baltimore Black Patients

Determination of the Duffy antigen status was made in a group of Baltimore patients with alcoholic liver disease. A parallel group of patients with chronic alcoholism but who did not show evidence of liver impairment was studied in a similar manner. An association between the presence of a positive Duffy antigen and alcoholic liver disease could not be demonstrated.     

Lymphocyte Transforming Activity in the Serum of Patients with Hepatic Disease

Abstract. The stimulating effect of serum of patients with hepatic disease on normal peripheral blood lymphocytes cultured in vitro was studied. In 25 out of 27 patients serum stimulated lymphocyte transformation as determined morphologically and by increased uptake of ³H-thymidine and ³H-cytidine. Rapidly increasing RNA synthesis was found during the first 24 h of incubation. DNA synthesis was found to be at its maximum between days 3 and 4. The behaviour of the serum of patients found to be positive for Australia antigen and patients with other hepatic disease was comparable. It was demonstrated that the factor in serum which initiates lymphocyte transformation, is in the globulin fraction.     

Upper Gastrointestinal Hemorrhage in Patients with Alcoholic Liver Disease and Esophageal Varices

One hundred and four patients with chronic alcoholic liver disease and esophageal varices were admitted to the hospital because of acute gastrointestinal hemorrhage. All were studied by early esophagogastroscopy to demonstrate the exact source of bleeding.
The most common bleeding site was the varices (41%). Other bleeding lesions included erosive gastritis (29%) and peptic ulceration (16%). The source of bleeding could not be detected in 14% of the patients. Eudoseopic examination proved helpful in providing the admission diagnosis, planning specific management and evaluating prognosis.
The over all mortality rate was 28%. Of those with variceal hemorrhage, 43% died. This was over twice the mortality associated with bleeding from peptic ulcer (18%) or erosive gastritis (20%). Exsanguination, hepatic failure or a combination of both was responsible for three-quarters of the fatalities. Death from exsanguination occurred most often during the first 48 hours and death from hepatic failure occurred either in the first two weeks or after the third week. Surgical intervention was associated with a high mortality rate. Other factors associated with a poorer prognosis were increased age, presence of ascites and abnormalities of serum albumin, bilirubin and transaminase levels.
In those patients who did not die of early exsanguination, hemorrhage often resulted in worsening liver function. When these changes were striking, the clinical course culminated in fatal hepatic failure. Patients who survived had little or no deterioration in liver function. Localization of source of bleeding and the evolution of change in liver function following the bleeding episode provide early prognostic indications of short-term outcome in these patients.     

Plasma Glutamate Dehydrogenase: Clinical Application in Patients With Alcoholic Liver Disease

The time course of plasma glutamate dehydrogenase (GDH) elevation was studied in 42 male alcoholics admitted for alcohol detoxification or for the treatment of medical complications of alcoholism and in 2 volunteers consuming 2 g/kg/day of ethanol under metabolic ward conditions. GDH values were usually highest during or immediately after cessation of drinking; thereafter, they fell rapidly toward the normal range. Early GDH values correlated well with liver histology in 37 patients who subsequently underwent diagnostic liver biopsy.     

Development of Hypoxemia in Alcoholic Liver Disease

The aim of this study was to investigate the arterial hypoxemia in Japanese patients with alcoholic liver disease (ALD) with regard to alcohol consumption and/or disease severity. Hypoxemia was observed in 78% patients with ALD and in all 46 patients with alcoholic liver cirrhosis (ALC) and 33 (56%) of 59 patients with noncirrhotic alcoholic liver disease (NCALD) (P < 0.0001). The partial pressure of oxygen (P_aO₂) was 71.1 ± 5.2 mm Hg in ALC and 78.7 ± 6.3 mm Hg in NCALD (P < 0.0001). The oxygen consumption in ALD was significantly higher than that in control subjects (P < 0.0001), and a high oxygen consumption was seen in 88% of the patients with ALD, in all 46 ALC patients, and in 46 (78%) of 59 NCALD patients (P < 0.01). Following abstinence from alcohol, the P_aO₂ and oxygen consumption significantly recovered after day 2 (P < 0.0001), whereas the prothrombin index did not change in either NCALD or ALC patients. Multivariate analysis showed that alcohol consumption and oxygen consumption were significant independent predictors of P_aO₂. In conclusion, the present findings suggest that increased oxygen consumption due to alcohol ingestion is principally responsible for the hypoxemia in ALD patients.     

Atypical Liver Alcohol Dehydrogenase in the Spanish Population: Its Relation with the Development of Alcoholic Liver Disease

The presence of atypical liver alcohol dehydrogenase (ADH) was determined in samples of liver tissue from 222 alcoholic and nonalcoholic subjects to determine its prevalence in the Spanish population, and to evaluate the possible relationship between the presence of this isoenzyme and the development of alcoholism and alcoholic liver disease. Alcoholic patients were classified into the following groups: control subjects, with normal liver pathology (group 1), patients with noncirrhotic liver disease (group 2), and patients with cirrhosis of the liver (group 3). Nonalcoholic subjects were also divided, following the same criteria, into groups 4, 5, and 6, respectively.
The prevalence of atypical ADH in the population analyzed was 16.2%. Atypical ADH was present in 14.9% of alcoholics and in 17.4% of nonalcoholics ( P = ns ). There were no significant differences when the prevalence of atypical ADH of alcoholic and nonalcoholic patients with similar degrees of liver pathology was compared (group 1 vs. 4, group 2 vs. 5, and group 3 vs. 6). The prevalence of atypical ADH was also similar in cirrhotic patients with respect to those of noncirrhotic liver disease and control patients, either in alcoholic or nonalcoholic groups.
Our findings indicate that the prevalence of atypical ADH in the Spanish population is similar to that reported for other Caucasian groups. Moreover, the presence of atypical ADH does not play a role in the development of alcoholism nor in the development of alcoholic liver disease in the population analyzed.     

Comparison of Hepatocellular Carcinoma Patients With Alcoholic Liver Disease and Nonalcoholic Steatohepatitis

Background: Alcoholic hepatitis and nonalcoholic steatohepatitis (NASH) show different clinical features with similar liver histology, but both disorders may progress to cirrhosis and hepatocellular carcinoma (HCC). HCC arising in alcoholic liver disease (ALD) or NASH, without hepatitis B or C virus infection, has been a rare observation, and there are no studies comparing the characteristics of ALD and NASH patients with HCC. Therefore, we compared the characteristics of ALD and NASH patients with HCC.
Methods: A total of 1202 patients received a diagnosis of HCC at Tokyo Women's Medical University from 1989 to 2003, and their clinical data were collected prospectively. A clinical diagnosis was made to diagnose ALD, and clinical and histological changes were required to diagnose NASH. Of these patients, 88 received a diagnosis of HCC arising from ALD. Among them, a biopsy specimen was obtained in 50 patients (ALD-HCC group). We compared the clinical and histological characteristics of 50 ALD and 8 NASH patients (NASH-HCC group) associated with HCC. They all were negative for hepatitis virus infection by serological methods.
Results: The most significant difference between these groups was sex. Women were significantly more common in the NASH-HCC group (6% vs. 63%; p < 0.0001). The median age was 65 years in the ALD-HCC group and 68 years in the NASH-HCC group. The risk factors for NASH all were high in the NASH-HCC group. However, liver function tests were similar in these groups. In the ALD-HCC group, 46 (92%) patients showed severe fibrosis; 2 had septal fibrosis and 44 had cirrhosis. All patients in the NASH-HCC group showed severe fibrosis, and seven had cirrhosis.
Conclusions: Severe fibrosis might be an important risk factor for HCC. Patients who have ALD or NASH with cirrhosis may develop HCC. This seems to occur in a sufficient number of cases to warrant regular screening for this complication.