Serum interleukin-1 (IL-1) activity in alcoholic hepatitis

Interleukin-1 (IL-1) is a monokine which has been demonstrated to produce a variety of seemingly diverse metabolic events including fever, neutrophilia, anorexia, altered mineral metabolism, muscle catabolism, and fibroblast proliferation. Because many of the clinical features of alcoholic hepatitis are metabolic abnormalities that have been shown to be caused by IL-1, we questioned whether patients with alcoholic hepatitis had elevated serum levels of IL-1. Six patients with alcoholic hepatitis had serum IL-1 activity measured by the thymocyte costimulator assay after serum inhibitors were removed. Their values were compared to those of 6 age and sex-matched healthy controls. Patients with alcoholic hepatitis had markedly elevated serum IL-1 activity, with the integrated value of all fractions having serum IL-1 activity being 9.8 times that of controls. IL-1 activity in serum from alcoholic hepatitis patients also was blocked by antibody to IL-1. We conclude that patients with alcoholic hepatitis have increased serum IL-1 activity which may play a role in certain of the metabolic complications of alcoholic hepatitis.     

Intermediate filaments in hyaline material in alcoholic liver disease (Mallory bodies) and in cultured hepatoma cells.

Antibodies specific for cytoskeletal intermediate (10 nm) filaments reacted both with the hyaline deposits in alcoholic liver disease (Mallory bodies) and with perinuclear hyaline material of cultured hepatoma cells used as a model for hyaline formation in vitro. Our results suggest that disorganization and accumulation of intermediate filaments is an important step in the pathogenesis of alcoholic liver injury.     

Intermediate filaments: Detection of lectin-like activity in purified alcoholic hyaline

Alcoholic hyaline, an intracellular, filamentous (10 nm) aggregate isolated from human alcoholic livers, bound the glycoprotein enzyme, horseradish peroxidase, in a specific and reproducible manner. Using a solid-phase assay system consisting of adsorbed alcoholic hyaline, we have shown that this binding is thermolabile, relatively insensitive to both pH extremes and high ionic strength, and highly sensitive to the presence of neutral and amino sugars. The results suggest that the binding of horseradish peroxidase is not a passive adsorption but rather an “active” phenomenon involving carbohydrate groups on the enzyme. The presence of an intracellular, filament-associated lectin is strongly indicated.     

Blood thiamine and thiamine phosphate ester concentrations in alcoholic and non-alcoholic liver diseases.

Thiamine state was investigated in patients with alcoholic liver disease, patients with various non-alcoholic liver diseases, and controls using a direct technique (thiochrome assay) to measure thiamine, thiamine monophospate, and the active coenzyme thiamine pyrophosphate in whole blood after isolating the fractions by ion exchange chromatography. Overall nutrition was similar in all groups as assessed by anthropometry, and no patient had clinical evidence of thiamine deficiency. There was no significant difference among the groups in mean concentration of any form of thiamine. The scatter was much greater in patients with alcoholic liver disease but only 8.7% had biochemical thiamine deficiency (defined as a blood concentration of the active coenzyme greater than 2 SD below the mean control value). An unexpected finding was of abnormally high total thiamine concentrations (greater than 2 SD above the mean control value) in 17.4% of patients with alcoholic liver disease, the highest concentrations being found in two patients with severe alcoholic hepatitis and cirrhosis. The ratio of phosphorylated to unphosphorylated thiamine was calculated as an index of phosphorylation and, although the mean did not differ significantly among the groups, the range was greatest in alcoholic liver disease. The lowest ratios occurred in the two patients with severe alcoholic hepatitis, but neither had evidence of thiamine pyrophosphate deficiency. Contrary to studies using indirect assay techniques, these results suggest that thiamine deficiency is unusual in well nourished patients with alcoholic liver disease. The new finding of unexpectedly high thiamine concentrations in some patients may be due to abnormalities of hepatic storage or release in liver disease, particularly in severe alcoholic hepatitis. There was no convincing evidence of impaired thiamine phosphorylation in any patients with liver disease. Conclusions from studies using indirect assays on the prevalence and mechanisms of thiamine deficiency in liver diseases may not be valid.     

Failure of Electron Paramagnetic Resonance Spectroscopy Studies to Detect Elevated Free Radical Signals in Liver Biopsy Specimens from Patients with Alcoholic Liver Disease

Electron paramagnetic resonance spectroscopy (EPR) was used to study free radicals and transition metal complexes in liver tissue taken from patients with liver disease. Samples were frozen to 77K directly following biopsy to prevent deterioration. Our major aim was to compare signals from patients suffering from alcohol abuse with those from patients having liver damage not induced by alcohol. Samples were obtained from 19 chronic alcohol abusers and 7 non-alcoholic liver disease patients. Of the 19 alcoholic patients, 18 had an increased fat content, 6 had Mallory's hyaline, 12 had an acute inflammatory response, 9 had increased stainable iron and 4 had evidence of fibrosis. A signal derived from free radicals with a spectroscopic splitting factor of g = 2.0045 was found in all samples. This signal in the alcoholic patients had a mean amplitude of 2.96 cm (± 1.42 SD), and in patients with non-alcoholic liver disease 2.12cm (±0.82) (p = 0.10NS), measured under identical instrument settings.

The molar proportion of diene conjugated linoleic acid (DCLA), a free radical marker, in the sera of alcoholic patients was 2.68% (±1.93), but did not correlate with the free radical signals obtained by EPR spectroscopy. Also, there was no correlation between the free radical derived EPR signal and fat content, Mallory's hyaline, inflammatory infiltrate, iron or fibrosis in the liver biopsy specimens. Similarly the concentrations of aspartate transaminase, albumin, and gamma-glutamyl transferase in serum samples showed no correlations with free radical concentrations.

The absence of any significant increase in the stable free radical signal in the presence of alcohol induced liver disease and the lack of correlation between the signal and either histological or serological evidence of liver damage, suggests that alcohol derived free radicals may not be involved in the pathogenesis of alcoholic liver disease.

Unusually large sextet features characteristic of MN(II) complexes were observed for all liver samples. Such signals are very rare in human tissue, showing that there is a strong accumulation of Mn (II) in the liver. However, no systematic trends were observed. In some samples signals characteristic of iron-sulphur cluster units were detected, but again no correlations could be discovered.     

Changing pattern of alcoholic liver disease in Great Britain: relation to sex and signs of autoimmunity.

A survey of 293 patients with alcoholic liver disease showed that women, particularly those aged under 45, had a significantly higher incidence of alcoholic hepatitis, with or without superimposed cirrhosis, than men. The long-term prognosis for both women who continued to drink and those who stopped drinking was worse than that for men. Autoantibodies were more common in women, which suggested that immune mechanisms may play a part in the pathogenesis and progression of alcoholic liver disease in women.     

Cytofluorimetric research on the glycogen content of the hepatocytes in patients with various forms of alcoholic liver lesions

By cytofluorometry employing the cytofluorometric PAS reaction, a study was made of the total glycogen and of its two fractions in liver parenchymal cells, both in the norm and in patients with chronic alcoholism (alcoholic steatosis, chronic alcoholic hepatitis, and mixed forms of alcoholic-viral hepatitis, viral hepatitis with steatosis and also viral hepatitis). The examination was performed on preparations-smears of isolated hepatocytes, obtained from the live puncture liver biopsies. The quantitative analysis has shown the increase in the total glycogen content in hepatocytes of patients with alcoholic hepatitis in comparison with the norm and with chronic viral hepatitis. The transition from a reverse stage--alcoholic steatosis--to alcoholic hepatitis was accompanied by a sharp increase in the total glycogen content and by an obvious change in the ratio of glycogen fractions, towards the hard soluble fraction in liver cells. The quantitative analysis of glycogen fractions in liver cells of patients with chronic alcoholic disease may be an appreciated marker of differential diagnostics of different stages and forms of alcoholic liver disease.     

Importance of markers of hepatitis B virus in alcoholic liver disease.

To determine the importance of the presence of serological markers of hepatitis B virus infection in patients with alcohol related liver disease we compared cumulative alcohol intake and clinical and histological features in patients with markers of hepatitis B virus infection and in those without. Hepatitis B surface antigen (HBsAg) was detected in five (2%) out of 285 patients studied and antibody to HBsAg (anti-HBs) in 41 (14%); one patient had antibody to hepatitis B core antigen alone. The combined prevalence of markers of hepatitis B virus infection was similar in patients with alcoholic cirrhosis (18%) and precirrhotic liver disease (13%). Two patients positive for HBsAg had histological features of both alcoholic liver disease and chronic active hepatitis, with stainable HBsAg. Patients with anti-HBs were, however, histologically indistinguishable from patients without markers, and the mean cumulative alcohol intake of patients with anti-HBs was similar to or even higher than that of patients with liver disease of comparable severity who had no evidence of previous infection. The presence of markers of hepatitis B virus infection was related to former residence in countries with a high prevalence of the infection and to previous parenteral treatment and blood transfusions. Infection with hepatitis B virus does not enhance the development of chronic liver disease in heavy drinkers, except in the small number who remain positive for HBsAg.     

Serum aspartate aminotransferase to alanine aminotransferase ratio in human and experimental alcoholic liver disease: relationship to histologic changes

We studied the relationship between the ratio of serum aspartate aminotransferase (ASAT) to alanine aminotransferase (ALAT) and histologic changes in human and experimental alcoholic liver disease. The patient population included 52 hospitalized patients enrolled in a Veterans Administration Cooperative study. The experimental animal group consisted of male Wistar rats fed an ethanol-liquid diet. Of the 52 patients with alcoholic hepatitis, 33 had evidence of cirrhosis. The mean +/- SD for the ASAT/ALAT ratio in the group with alcoholic hepatitis and no cirrhosis was 1.47 +/- 0.84, the mean +/- SD in the group with hepatitis and cirrhosis was significantly higher (2.68 +/- 1.32, p less than 0.01). There was no difference in the ratio between the rats with and without liver fibrosis. The cause for the increased ASAT/ALAT ratio in serum in the presence of cirrhosis is unknown and may reflect more severe liver damage.     

Clinical, enzymological and histological changes in chronic diffuse liver diseases following (+)-cyanidanol-3 (Catergen) treatment

The effect of (+)-cyanidanol-3 (Catergen) monotherapy was examined in 18 patients with alcoholic liver disease and in 12 patients with chronic active hepatitis. During the administration of the drug the majority of complaints diminished or ceased in both groups. In patients with alcoholic liver disease bromsulphalein retention and gamma glutamyl transferase levels decreased, in chronic active hepatitis serum glutaminic pyruvic acid transaminase (SGPT) and alkaline phosphatase levels increased, pseudocholinesterase level decreased. The histological abnormalities of alcoholic liver injury improved in the majority of cases, on the other hand, it was deteriorated in two third of the cases with chronic active hepatitis. In two cases the histological recovery of acute alcoholic hepatitis was observed. On the basis of this results we conclude that Catergen has an excellent therapeutic effect in alcoholic liver injury while in chronic active liver diseases it can be applied only as a part of combined therapy.     

Rod-shaped inclusions in the heart and brain in the experimental alcoholization of rats

Experiments were made on rat alcoholized by inhalation. Electron microscopy of the myocardium and brain cortex showed the dystrophically altered cells of these organs to contain unusual rod-shaped and sagittal cytoplasmic inclusions unrelated to organelles and having a certain structural similarity to alcoholic hyaline detected in men. The necessity of further studies to identify the inclusions in question is discussed.     

Lymphocytotoxicity in alcoholic hepatitis: evalutation of effector cells

Alcohol induced liver damage has been recently suggested to occur in some cases as a consequence of a true autoimmune reaction. However the nature of the effector cells is still debated. Microlymphocytotoxicity "in vitro" test against rabbit hepatocytes has been performed in 29 patients with alcoholic cirrhosis and 8 with acute alcoholic hepatitis. The diagnosis has been done on clinical and histological grounds. The mean cytotoxicity index (CI) of subjects with alcoholic cirrhosis was 15.12 +/- 14.18 and with acute alcoholic hepatitis 62.49 +/- 16.33 (normal range: 0 -37.90). To further characterize the effector cells in our system, the test has been performed again either using T-enriched fractions of peripheral lymphocytes or after pronase-induced digestion of SmIg of total lymphocytes in 5 patients with acute alcoholic hepatitis and high CI. In both instances a disappearance of citotoxic activity has been observed. These data suggest: a) autoimmune reaction can occur only in patients with acute alcoholic liver damage; b) non T-cells are responsible for cytotoxicity; c) the immune reaction appears to be of cell-mediated antibody-dependent type.     

Cost-Effectiveness Analysis of Sofosbuvir Compared to Current Standard Treatment in Swiss Patients with Chronic Hepatitis C

In clinical trials, sofosbuvir showed high antiviral activity in patients infected with hepatitis C virus (HCV) across all genotypes. We aimed to determine the cost-effectiveness of sofosbuvir-based treatment compared to current standard treatment in mono-infected patients with chronic hepatitis C (CHC) genotypes 1–4 in Switzerland. Cost-effectiveness was modelled from the perspective of the Swiss health care system using a lifetime Markov model. Incremental cost-effectiveness ratios (ICERs) used an endpoint of cost per quality-adjusted life year (QALY) gained. Treatment characteristics, quality of life, and transition probabilities were obtained from published literature. Country-specific model inputs such as patient characteristics, mortality and costs were obtained from Swiss sources. We performed extensive sensitivity analyses. Costs and effects were discounted at 3% (range: 0–5%) per year. Sofosbuvir-containing treatment in mixed cohorts of cirrhotic and non-cirrhotic patients with CHC genotypes 1–4 showed ICERs between CHF 10,337 and CHF 91,570 per QALY gained. In subgroup analyses, sofosbuvir dominated telaprevir- and boceprevir-containing treatment in treatment-naïve genotype 1 cirrhotic patients. ICERs of sofosbuvir were above CHF 100,000 per QALY in treatment-naïve, interferon eligible, non-cirrhotic patients infected with genotypes 2 or 3. In deterministic and probabilistic sensitivity analyses, results were generally robust. From a Swiss health care system perspective, treatment of mixed cohorts of cirrhotic and non-cirrhotic patients with CHC genotypes 1–4 with sofosbuvir-containing treatment versus standard treatment would be cost-effective if a threshold of CHF 100,000 per QALY was assumed.     

Vitamin E content and lipid peroxidation of blood in some chronic inflammatory diseases

Patients suffering from rheumatoid arthritis, spondylosis, coxarthrosis, ankylosing spondylitis, chronic active and chronic alcoholic hepatitis were studied. The plasma vitamin E content remained unchanged. The TBA-reactive plasma substances (malondialdehyde) content of plasma increased in all patients except those with ankylosing spondylitis. Catalase activity of plasma increased in patients of both sexes suffering from rheumatoid arthritis and spondylosis and coxarthrosis, but decreased in the two hepatitis groups. The glutathione-peroxidase activity of RBC (1:9 haemolysate) increased in female rheumatoid arthritis patients and decreased in those suffering from chronic alcoholic hepatitis. The results showed that chronic inflammatory processes affect the rate of lipid peroxidation and the activity of the biological antioxidant mechanism.     

T cells from patients with chronic liver diseases: abnormalities in PHA-induced expression of HLA class II antigens and in autologous mixed-lymphocyte reactions

The expression of HLA Class II antigens by resting and phytohemagglutinin (PHA)-activated T cells and their functional properties in autologous mixed-lymphocyte reactions (MLR) were investigated in patients with chronic active hepatitis, with alcoholic cirrhosis, and with primary biliary cirrhosis. In all groups of patients the percentage of resting T cells expressing HLA Class II antigens was significantly higher than that in controls. The percentage of T cells which acquired HLA Class II antigens following PHA stimulation was reduced in patients with chronic active hepatitis, serum hepatitis B surface antigen (HBsAg) positive, and in those with alcoholic cirrhosis, HBsAg negative, although the level of [3H] thymidine incorporation was within normal limits. The degree of proliferation in autologous MLR with PHA-T cells was significantly reduced in patients with chronic active hepatitis, HBsAb positive, and in those with alcoholic cirrhosis, HBsAg positive. A reduced proliferation was also detected in autologous MLR with non-T cells, in patients with chronic active hepatitis, HBsAg positive. The abnormalities of autologous MLR are selective, since the proliferative and stimulatory activities of cells from patients with chronic liver diseases in allogeneic MLR were within normal ranges. The immunoregulatory role of HLA Class II antigens and of autologous MLR suggests that the abnormalities we have identified may play a role in the immunological dysfunctions underlying chronic liver diseases.     

Prospective study of post-transfusion hepatitis after cardiac surgery in a British centre.

A series of 248 consecutive patients undergoing cardiac surgery were examined in a prospective study of post-transfusion hepatitis in a single British centre. Patients received a total of 1796 units of blood or blood products (mean blood transfusion 6.28 units per patient). During five to 30 days after operation 38 of the patients showed an increase in serum transaminase activities. There was no serological evidence for fresh infection by hepatitis A or B virus, cytomegalovirus, Epstein-Barr virus, or herpes virus in any of these patients. The increase in transaminase activities was unexplained and reached over 100 IU/l (normal less than 40 IU/l) in six patients. The incidence of acute short incubation post-transfusion non-A, non-B hepatitis was therefore thought to be 2.4%. These six patients had normal liver function six months after transfusion but a further two of the surviving 228 patients had raised serum transaminase activities at six months. In one of these, liver biopsy disclosed chronic persistent hepatitis; in the other, alcoholic liver disease was suspected. The incidence of significant chronic liver disease after blood transfusion possibly attributable to a non-A, non-B hepatitis agent was therefore only 0.4%.     

Relative and Combined Effects of Chronic Alcohol Consumption and HCV Infection on Serum Zinc, Copper, and Selenium

In alcoholic hepatitis, Kupffer cells are activated by intestinal gram-bacteria, leading to cytokine production and free radicals release, which, enhancing cytokine secretion, create a positive feedback loop which contributes to liver inflammation. Free radicals also damage the liver in chronic hepatitis C virus (HCV) infection, a condition frequently associated to alcohol consumption. In both situations, activity of antioxidant enzymes and of its cofactors zinc (Zn), selenium (Se), and copper (Cu) is important. This study was performed to assess the relative and combined effects of chronic alcoholism and HCV infection on serum Se, Zn, and Cu, and its relation with serum malondialdehyde (MDA) and tumor necrosis factor-α, interferon-γ, and interleukins (IL) 4, 6, and 8, in 19 HCV? alcoholic patients, 12 HCV+ alcoholic patients, nine HCV+ non-alcoholic patients, and 20 controls. Serum Zn and Se were lower in both HCV+ and HCV? alcoholic patients, whereas serum Cu was lower in HCV+ individuals. Serum Zn and Se were related to liver function derangement. MDA levels were higher in alcoholics, but no relation was observed between trace elements and MDA or cytokines, so that our results do not support a relevant role of the analyzed trace elements in the pathogenesis of chronic liver disease.     

Prolyl hydroxylase activity in normal and diseased human liver.

Prolyl hydroxylase activity was determined in liver biopsy samples obtained from 10 patients. The liver prolyl hydroxylase values in patients with active hepatitis distribute into two numerical populations based on the extent of elevation over control. The first of these groups includes those with enzyme levels elevated approximately 2.5-fold over normal. Included in this group are patients with active (but nonagrressive) hepatitis and patients where advanced portal fibrosis is already established. The second group where prolyl hydroxylase is elevated approximately nine-fold is comprised of two patients with advanced clinical symptoms of active alcoholic hepatitis with evidence of aggressive cirrhosis but with only early minimal evidence of existing fibrosis.     

Therapeutic effect of (+)-cyanidanol-3 in toxic alcoholic liver disease and in chronic active hepatitis

(+)-Cyanidanol-3 is considered to have cytoprotective effect in toxic liver injury. A randomized clinical trial was carried out in alcoholic precirrhotic patients and in chronic active hepatitis with (+)-cyanidanol-3 versus placebo. The daily dose of the drug was 1.5-2.0 g for a one-year period. A: Toxic alcoholic precirrhotic liver disease: 38 patients were treated with (+)-cyanidanol-3, 36 with placebo. We found significant improvement in the subjective symptome like asthenia and anorexia, and in serum aspartate-transaminase (GOT) levels. However, it is possible that the improvement was in part due to abstinence from alcohol. B: Chronic active hepatitis: previously introduced continuous prednisolone therapy (10-15 mg/day) was combined with (+)-cyanidanol-3 in 13 patients and with placebo in 12 controls. The results showed a more favourable, but not significantly better response in patients receiving (+)-cyanidanol-3 versus placebo. We concluded that the drug might be of benefit in some cases with chronic active hepatitis as an adjunct to corticoid therapy.     

From Mallory to Mallory-Denk bodies: what, how and why?

Frank B. Mallory described cytoplasmic hyaline inclusions in hepatocytes of patients with alcoholic hepatitis in 1911. These inclusions became known as Mallory bodies (MBs) and have since been associated with a variety of other liver diseases including non-alcoholic fatty liver disease. Helmut Denk and colleagues described the first animal model of MBs in 1975 that involves feeding mice griseofulvin. Since then, mouse models have been instrumental in helping understand the pathogenesis of MBs. Given the tremendous contributions made by Denk to the field, we propose renaming MBs as Mallory-Denk bodies (MDBs). The major constituents of MDBs include keratins 8 and 18 (K8/18), ubiquitin, and p62. The relevant proteins and cellular processes that contribute to MDB formation and accumulation include the type of chronic stress, the extent of stress-induced protein misfolding and consequent proteasome overload, a K8-greater-than-K18 ratio, transamidation of K8 and other proteins, presence of p62 and autophagy. Although it remains unclear whether MDBs serve a bystander, protective or injury promoting function, they do serve an important role as histological and potential progression markers in several liver diseases.