Alcoholic Fibrosis Research Articles

Molecular mechanism of the reversibility of hepatic fibrosis: with special reference to the role of matrix metalloproteinases.

The participation of matrix metalloproteinases (MMP) and their specific inhibitors, the tissue inhibitors of matrix metalloproteinases (TIMP), in both the formation and degradative recovery processes of liver fibrosis were mainly reviewed from the molecular biological aspect. Since authors first reported increased activity of interstitial collagenase in the early stage of hepatic fibrosis in rats induced by chronic CCl4 intoxication, in baboons fed alcohol chronically and in patients with alcoholic fibrosis, other investigators have also demonstrated increased activity biologically and histochemically. However, species-specific differences in response have been found and gene-level research on the rat model has not demonstrated increased mRNA transcription of collagenase. It has also been clarified that activated stellate cells can also produce matrix components. Very recently, authors observed the participation of interstitial collagenase in the recovery from experimental hepatic fibrosis by using polymerase chain reaction northern blotting and in situ hybridization. The in situ hybridization findings not only demonstrated the cells responsible for interstitial collagenase, but also suggested a great deal about the mechanism of recovery from fibrosis. Hepatic stellate cells are activated via the expression of c-myb and nuclear factor-kappaB (NFkappaB) which is induced by oxidative stress, and inhibited by antioxidant (1-alpha-tocopherol) and butylated hydroxytoluene. The activation mechanism is now being revealed. The relationship between the activation mechanism of stellate cells and the production and secretion of MMP and TIMP in the formation and recovery process of hepatic fibrosis should be investigated from the promoter gene level. This approach might help develop a new strategy for the treatment of liver fibrosis.

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.

Periacinar collagenization in patients with chronic alcoholism.

To examine the development of pancreatic fibrosis in alcoholics, the fibrosis types grouped according to Martin's classification were examined by immunohistochemistry using an antibody against alpha-smooth muscle actin (alpha-SMA). The initial stage of periacinar collagenization was also investigated by electron microscopy. The total incidence of pancreatic fibrosis at autopsy of the 29 alcoholics was significantly higher than that of the 40 non-alcoholics. Intralobular sclerosis was observed to be the most frequent type of fibrosis regardless of alcohol intake. No differences in the enhancement of alpha-SMA expression in each type of fibrosis were found between the alcoholics and non-alcoholics. Electron microscopically, myofibroblasts were found around acini in the early stage of periacinar collagenization, and were accompanied by numerous fine filaments (8-15 nm in diameter). The various changes in zymogen granules (ZG), lysosomes and lipid droplets were augmented in the acinar cells of alcoholics. Medium-density materials were also found in dilated rough endoplasmic reticulum (RER). The contents of ZG and RER occasionally leaked out. In conclusion, pancreatic fibrosis was increased in alcoholics; myofibroblasts may play an important role in the initial stage of periacinar collagenization; and the intracellular transport blockage of protein as represented by abnormalities of ZG, ER and lysosomes may contribute to the development of periacinar collagenization.

Correlation of a polymorphism in the interleukin-1 receptor antagonist gene with hepatic fibrosis in Japanese alcoholics.

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.

Correlation of a Polymorphism in the Interleukin-1 Receptor Antagonist Gene with Hepatic Fibrosis in Japanese Alcoholics

Correlation of a Polymorphism in the Interleukin-1 Receptor Antagonist Gene with Hepatic Fibrosis in Japanese Alcoholics

Clinical Significance of Hepatitis GB Virus C Infection in Alcoholic Liver Disease

Recently, hepatitis GB virus C (HGBV-C) has been recovered from patients with non-A-E hepatitis. However, it has been unclear whether HGBV-C may be related to the development of alcoholic liver disease (ALD) or not. In this study, we determined HGBV-C RNA in sera from alcoholic patients without markers for hepatitis C and B viruses to evaluate the role of HGBV-C in ALD. Serum samples were obtained from 68 patients with ALD and 40 nonalcoholic patients with chronic type C liver disease. HGBV-C RNA was detected in only 3 of 68 (4.4%) patients with ALD, in 2 of 27 patients with hepatic fibrosis, and in 1 of 5 patients with chronic hepatitis. There was no HGBV-C RNA in sera from patients with fatty liver, alcoholic hepatitis, or cirrhosis. Serum levels of AST, ALT, and γ-glutamyltranspeptidase in alcoholic patients with, as well as without, HGBV-C RNA decreased to normal levels after abstinence. In addition, an inflammatory change was not observed in liver biopsy specimens obtained from two HGBV-C-positive patients with alcoholic hepatic fibrosis. Our results clearly suggest that the prevalence of HGBV-C infection in patients with ALD is rare and that HGBV-C may not play an important role in the development of liver disease in alcoholics.

Clinical Significance of Hepatitis GB Virus C Infection in Alcoholic Liver Disease

Recently, hepatitis GB virus C (HGBV-C) has been recovered from patients with non-A-E hepatitis. However, it has been unclear whether HGBV-C may be related to the development of alcoholic liver disease (ALD) or not. In this study, we determined HGBV-C RNA in sera from alcoholic patients without markers for hepatitis C and B viruses to evaluate the role of HGBV-C in ALD. Serum samples were obtained from 68 patients with ALD and 40 nonalcoholic patients with chronic type C liver disease. HGBV-C RNA was detected in only 3 of 68 (4.4%) patients with ALD, in 2 of 27 patients with hepatic fibrosis, and in 1 of 5 patients with chronic hepatitis. There was no HGBV-C RNA in sera from patients with fatty liver, alcoholic hepatitis, or cirrhosis. Serum levels of AST, ALT, and gamma-glutamyltranspeptidase in alcoholic patients with, as well as without, HGBV-C RNA decreased to normal levels after abstinence. In addition, an inflammatory change was not observed in liver biopsy specimens obtained from two HGBV-C-positive patients with alcoholic hepatic fibrosis. Our results clearly suggest that the prevalence of HGBV-C infection in patients with ALD is rare and that HGBV-C may not play an important role in the development of liver disease in alcoholics.

In situ nucleic acid hybridization of cytokines in primary biliary cirrhosis: predominance of the Th1 subset.

Primary biliary cirrhosis (PBC) is an autoimmune liver disease characterized by destruction of the intrahepatic bile ducts. It is generally believed that cellular immune mechanisms, particularly involving T cells, result in this bile duct damage. The relative strength of Th1 and Th2 responses has recently been proposed to be an important factor in the pathophysiology of various autoimmune diseases. In this study, we have attempted to identify the Th subset balance in PBC, by detection of cytokines specific to the two T-cell subsets, i.e., interferon gamma (IFN-gamma) for Th1 cells and interleukin-4 (IL-4) for Th2 cells. We analyzed IFN-gamma and IL-4 messenger RNA (mRNA) positive cells in liver sections from 18 patients with PBC and 35 disease controls including chronic active hepatitis C, extrahepatic biliary obstruction (EBO), and normal liver, using nonisotopic in situ hybridization and immunohistochemistry. Mononuclear cells expressing IFN-gamma and IL-4 mRNA were aggregated in inflamed portal tracts in PBC livers, but were rarely present in extrahepatic biliary obstruction, alcoholic fibrosis, or normal liver sections. The IFN-gamma and IL-4 mRNA positive cells in PBC livers were detected in significantly higher numbers than in control livers (P < .01). Moreover, IFN-gamma mRNA expression was more commonly detected than IL-4 expression in PBC livers, and the levels of IFN-gamma mRNA expression were highly correlated with the degree of portal inflammatory activity. IFN-gamma mRNA-positive cells were detected primarily around damaged bile ducts that were surrounded by lymphoid aggregates. The data indicate that Th1 cells are the more prominent T-cell subset in the lymphoid infiltrates in PBC.

Pathogenesis and treatment of liver fibrosis in alcoholics: 1996 update.

Fibrosis is a common end stage for most chronic liver diseases. It results from an imbalance between collagen production and degradation. One promising approach for prevention and treatment is the stimulation of collagenolytic processes. In nonhuman primates it was found that polyenylphosphatidylcholine (PPC), extracted from soybeans, protects against alcohol-induced fibrosis and cirrhosis and prevents the associated hepatic phosphatidylcholine (PC) depletion by increasing 18:2-containing PC species; it also attenuates the transformation of lipocytes into collagen-producing transitional cells. Furthermore, it increases collagen breakdown, as shown in cultured lipocytes enriched with pure dilinoleoyl PC (18:2-18:2 PC), the main PC species present in the extract, which may be the active ingredient. Since PC appears to promote the breakdown of collagen, there is reasonable hope that this treatment may affect not only the progression of the disease, but may also reverse preexisting fibrosis, as demonstrated for CCl4-induced cirrhosis in the rat. Therefore, PPC may be useful for the management of fibrosis of alcoholic and nonalcoholic etiologies as well. S-Adenosylmethionine opposes CCl4-induced fibrosis and can affect some of the consequences of the ethanol-induced oxidative stress in experimental animals and in man. Anti-inflammatory medications (corticosteroids, colchicine) are also being used and agents that interfere with collagen synthesis, such as inhibitors of prolyl-4-hydroxylase and antioxidants, are being tested.

Serum hyaluronate reflects hepatic fibrogenesis in alcoholic liver disease and is useful as a marker of fibrosis.

The high levels of hyaluronic acid (HA), a glycosaminoglycan of the liver extracellular matrix, which is synthesized and degraded in the liver sinusoidal cells, have been related with a decreased function of the endothelial sinusoidal cells. The relevance of HA in alcoholic liver disease has not been sufficiently evaluated, and therefore the current study was addressed to assess whether serum HA reflects the severity of liver fibrosis and fibrogenesis as well as the potential usefulness of hyaluronic acid as a marker of early fibrosis in alcoholics with liver damage. Serum HA and aminoterminal propeptide of collagen III (PIIIP) levels, a marker of liver fibrogenesis in alcoholics with liver disease, were assessed in 45 chronic alcoholic patients (31 men and 14 women, age: 44.1 +/- 1.5 years) (normal liver = 7; fatty changes = 8; fibrosis = 7; alcoholic hepatitis = 6; cirrhosis = 6; and cirrhosis plus alcoholic hepatitis = 11). The severity of liver inflammation and fibrosis were scored in liver specimens as: 0, no lesion; 1+ mild; 2+ moderate; and 3+ severe. Twenty-seven patients (60%) had HA above normal values (1 patient with fatty changes, 3 patients with fibrosis, and all patients with alcoholic hepatitis or cirrhosis). Hyaluronic acid and (PIIIP) levels increased in parallel with the severity of liver damage. Hyaluronic acid levels were higher in those patients with more liver inflammation (0, 128 +/- 38; 1+, 553 +/- 141; 2+, 668 +/- 259; 3+, and 1,073 +/- 419 microg/L; P = .004) and of fibrosis (0, 79 +/- 32; 1+, 156 +/- 70; 2+, 219 +/- 105; and 3+, 695 +/- 114 microg/L; P < .001). Procollagen III peptide levels were related with fibrosis (0, 17 +/- 1; 1+, 25 +/- 6; 2+, 47 +/- 13; 3+, and 55 +/- 9 ng/mL; P = .002) but not with inflammation (0, 29 +/- 7; 1+, 45 +/- 7; 2+, 54 +/- 9; 3+, and 66 +/- 30 ng/mL, P: not significant). Moreover, a direct linear correlation was observed between HA and PIIIP (r = .72, P < .001). A receiver operating characteristic (ROC) curve analysis revealed that HA was similar to PIIIP levels in discriminating between alcoholics without fibrosis and those with fibrosis (area under the ROC curves) .913 +/- .042 vs. .867 +/- .054; P: n.s). In conclusion, serum HA reflects the severity of liver inflammation, fibrosis, and fibrogenesis in patients with alcoholic liver disease and is useful as a marker of precirrhotic and cirrhotic stages.

Serum hyaluronate reflects hepatic fibrogenesis in alcoholic liver disease and is useful as a marker of fibrosis

The high levels of hyaluronic acid (HA), a glycosaminoglycan of the liver extracellular matrix, which is synthesized and degraded in the liver sinusoidal cells, have been related with a decreased function of the endothelial sinusoidal cells. The relevance of HA in alcoholic liver disease has not been sufficiently evaluated, and therefore the current study was addressed to assess whether serum HA reflects the severity of liver fibrosis and fibrogenesis as well as the potential usefulness of hyaluronic acid as a marker of early fibrosis in alcoholics with liver damage. Serum HA and aminoterminal propeptide of collagen III (PIIIP) levels, a marker of liver fibrogenesis in alcoholics with liver disease, were assessed in 45 chronic alcoholic patients (31 men and 14 women, age: 44.1 +/- 1.5 years) (normal liver = 7; fatty changes = 8; fibrosis = 7; alcoholic hepatitis = 6; cirrhosis = 6; and cirrhosis plus alcoholic hepatitis = 11). The severity of liver inflammation and fibrosis were scored in liver specimens as: 0, no lesion; 1+ mild; 2+ moderate; and 3+ severe. Twenty-seven patients (60%) had HA above normal values (1 patient with fatty changes, 3 patients with fibrosis, and all patients with alcoholic hepatitis or cirrhosis). Hyaluronic acid and (PIIIP) levels increased in parallel with the severity of liver damage. Hyaluronic acid levels were higher in those patients with more liver inflammation (0, 128 +/- 38; 1+, 553 +/- 141; 2+, 668 +/- 259; 3+, and 1,073 +/- 419 microg/L; P = .004) and of fibrosis (0, 79 +/- 32; 1+, 156 +/- 70; 2+, 219 +/- 105; and 3+, 695 +/- 114 microg/L; P < .001). Procollagen III peptide levels were related with fibrosis (0, 17 +/- 1; 1+, 25 +/- 6; 2+, 47 +/- 13; 3+, and 55 +/- 9 ng/mL; P = .002) but not with inflammation (0, 29 +/- 7; 1+, 45 +/- 7; 2+, 54 +/- 9; 3+, and 66 +/- 30 ng/mL, P: not significant). Moreover, a direct linear correlation was observed between HA and PIIIP (r = .72, P < .001). A receiver operating characteristic (ROC) curve analysis revealed that HA was similar to PIIIP levels in discriminating between alcoholics without fibrosis and those with fibrosis (area under the ROC curves) .913 +/- .042 vs. .867 +/- .054; P: n.s). In conclusion, serum HA reflects the severity of liver inflammation, fibrosis, and fibrogenesis in patients with alcoholic liver disease and is useful as a marker of precirrhotic and cirrhotic stages. (Hepatology 1996 Dec;24(6):1399-403)

Effect of Alcohol Intake on the Efficacy of Interferon Therapy in Patients with Chronic Hepatitis C as Evaluated by Multivariate Logistic Regression Analysis

The effect of alcohol intake on the efficacy of interferon (IFN) therapy was evaluated retrospectively in patients with chronic hepatitis C diagnosed by liver histology and positive serum hepatitis C virus (HCV)-RNA. Patients included 119 given IFN therapy and 11 no IFN therapy. Serum HCV-RNA was measured 6 months after discontinuation of IFN therapy in 92 treated patients, 27.2% of whom showed disappearance of serum HCV-RNA. Multivariate logistic regression analysis revealed that this disappearance was affected by alcohol intake, the presence of its history (p < 0.05) or cumulative alcohol consumption (kg) (p < 0.01), and serum HCV-RNA levels (p < 0.001). The odds ratio associated with serum HCV-RNA still positive at 6 months was 7.018 (95% confidence interval: 1.444-34.062) and 1.004 (1.001-1.007) for the presence of alcohol intake history and the cumulative alcohol consumption, respectively. Other predictor variables-such as sex and age of patients, history of blood transfusion, HCV genotype, histological findings of the liver, and types of IFN-had no influence on the efficacy of the therapy. Cumulative alcohol consumption showed a negative correlation with serum HCV-RNA levels pretreatment, when the outcome variable was divided into two categories based on serum HCV-RNA levels: 10(6) copy/ml or less and 10(7) copy/ml or more. Alcohol intake was positively correlated with histological extent of alcoholic fibrosis, but affected neither grading nor staging of chronic viral hepatitis. We conclude that alcohol intake was a risk factor on the efficacy of IFN therapy in chronic hepatitis C patients. This effect was independent of serum HCV-RNA levels and histological findings specific for viral hepatitis in the liver.

Effect of Alcohol Intake on the Efficacy of Interferon Therapy in Patients with Chronic Hepatitis C as Evaluated by Multivariate Logistic Regression Analysis

The effect of alcohol intake on the efficacy of interferon (IFN) therapy was evaluated retrospectively in patients with chronic hepatitis C diagnosed by liver histology and positive serum hepatitis C virus (HCV)-RNA. Patients included 119 given IFN therapy and 11 no IFN therapy. Serum HCV-RNA was measured 6 months after discontinuation of IFN therapy in 92 treated patients, 27.2% of whom showed disappearance of serum HCV-RNA. Multivariate logistic regression analysis revealed that this disappearance was affected by alcohol intake, the presence of its history (p < 0.05) or cumulative alcohol consumption (kg) (p < 0.01), and serum HCV-RNA levels (p < 0.001). The odds ratio associated with serum HCV-RNA still positive at 6 months was 7.016 (95% confidence interval: 1.444-34.082) and 1.004 (1.001-1.007) for the presence of alcohol intake history and the cumulative alcohol consumption, respectively. Other predictor variables–such as sex and age of patients, history of blood transfusion, HCV genotype, histological findings of the liver, and types of IFN– had no influence on the efficacy of the therapy. Cumulative alcohol consumption showed a negative correlation with serum HCV-RNA levels pretreatment, when the outcome variable was divided into two categories based on serum HCV-RNA levels: 106 copy/ml or less and 107 copy/ml or more. Alcohol intake was positively correlated with histological extent of alcoholic fibrosis, but affected neither grading nor staging of chronic viral hepatitis. We conclude that alcohol intake was a risk factor on the efficacy of IFN therapy in chronic hepatitis C patients. This effect was independent of serum HCV-RNA levels and histological findings specific for viral hepatitis in the liver.

Experimental liver fibrosis induced in rats receiving high doses of alcohol and alternating between regular and vitamin-depleted diets.

Liver fibrosis was induced in rats by simulating human alcoholic eating and drinking patterns. Alcohol addiction was established by gradually increasing the ethanol concentration in the drinking water; salts were added at the terminal stage. The hepatocytes of rats receiving alcohol concentrations exceeding 50% (v/v) (similar to vodka) exhibited alcoholic hyaline (Mallory bodies). Alcoholic liver fibrosis was induced by alternating between regular and autoclaved (vitamin-depleted) diets, simulating the irregular eating habits of human alcoholics. In the livers of rats receiving 70% (v/v) ethanol (comparable to absinthe) with 25% saline and fed the alternating diets, pericellular fibrosis was induced. No significant difference in calorie intake between control and alcohol rats was detected except when rats underwent drinking bouts (heavy drinking phase). This indicates that neither a high-fat diet nor a choline-depleted diet is necessary to induce the alcoholic fibrosis seen in human alcoholics.

National survey of alcoholic liver disease in Japan (1968-91).

National surveys of alcoholic liver disease (ALD) in Japan were performed in 1978 and 1985 by a previous Japanese study group for ALD (the Takeuchi group). In the present study, a subsequent nationwide survey of ALD in Japan was conducted from 1986 to 1991 and the results compared with the previous studies. In order to clarify the aetiological relationship between hepatitis C virus (HCV) infection and ALD, results were also analysed according to new diagnostic criteria for ALD proposed by the current ALD study group (the Takada group). According to the diagnostic criteria of the Takeuchi group, the incidence of ALD did not differ significantly from 1986 to 1991. However, the incidence of hepatocellular carcinoma (HCC) in alcoholic cirrhosis (AL-LC) clearly increased during this period. The analysis, which included analysis of results from the previous studies, indicated that the incidence of ALD reached a plateau in 1980 and then stabilized. However, HCC in AL-LC continued to show a linear increase from 1976 to 1991. The new diagnostic criteria of the Takada group were used to analyse cases from 1990 and 1991. Approximately two out of every three cases of ALD were caused by alcohol alone, and the remainder were caused by a combination of alcohol and HCV. Cases caused only by HCV were very rare. The main aetiology in patients with alcoholic hepatitis and fibrosis was alcohol alone, and in the case of chronic hepatitis, in heavy drinkers, it was a combination of alcohol and HCV. In half the patients with AL-LC the aetiology was alcohol alone, and in the other half it was a combination of both alcohol and HCV. In the majority of patients with HCC, the aetiology was a combination of alcohol and HCV, indicating that HCV infection may be important in the development of HCC in alcoholics.

MALONDIALDEHYDE STIMULATES COLLAGEN PRODUCTION BY HEPATIC LIPOCYTES ONLY UPON ACTIVATION IN PRIMARY CULTURE

Lipid aldehydes have been proposed as mediators of hepatic fibrosis in alcoholics. In this study we examined whether hepatic lipocytes, the principal matrix-producing cells in liver, exhibit enhanced collagen synthesis in response to the lipid aldehyde malondialdehyde. Lipocytes isolated from normal rat liver and plated in primary culture for 3 days were not affected by malondialdehyde in concentrations ranging from 2 to 200 microM. Cells cultured for 7 days displayed a modest increase in collagen synthesis (137% of control levels) in response to malondialdehyde, but only at a concentration of 200 microM. The malondialdehyde-induced increase in collagen synthesis was paralleled by a rise in type I procollagen mRNA. Subcultured rat fibroblasts at confluent density responded better to malondialdehyde than did 7-day lipocytes. The results indicate that lipocytes respond to the fibrogenic effects of malondialdehyde only after activation in primary culture. This delayed response suggests that lipid aldehydes may enhance, but do not initiate, alcoholic liver fibrosis in vivo.

Detection of antigens related to hepatitis C virus RNA encoding the NS5 region in the livers of patients with chronic type C hepatitis

Hepatitis C virus is a positive single-strand RNA virus distantly related to flaviviruses. Therefore RNA replicase, an RNA-dependent RNA polymerase, may be essential for the replication of hepatitis C virus, as well as other RNA viruses. In this study we synthesized the recombinant polypeptide (HCV-NS5 antigen) with a 576 bp cDNA encoding a part of the NS5 region of the HCV genome that has the Gly-Asp-Asp motif. The antibody against this polypeptide was obtained from rabbit serum. In Western-blot analysis with NS5 IgG HCV antibody, an 84-kD protein was clearly detected as a single band in the microsomal fraction but not in the nuclear and mitochondrial fractions or in the cytosol fraction. Immunohistochemically, HCV-NS5 antigen was clearly stained in the cytoplasm of hepatocytes but not in the nucleus or cell membrane. Moreover, as determined on immunoelectron microscopy, HCV-NS5 antigen was demonstrated with fine granular distribution along the endoplasmic reticulum but not in other organelles, including the nucleus and mitochondria. Immunoreaction in other cell types was negative. These results indicate that replication of HCV may occur only in hepatocytes and that HCV-NS5 may be produced in the endoplasmic reticulum of these cells. HCV-NS5 antigen was stained only in the livers of hepatitis C virus–positive patients but not in sections from patients with chronic type B hepatitis or alcoholic fibrosis. In chronic type C liver disease, the overall detection rate of HCV-NS5 antigen was 56 (33 in chronic persistent hepatitis, 52 in chronic active hepatitis and 86 in cirrhosis). These results indicate that the replication of HCV may occur more frequently in the advanced than in the early stages of type C hepatitis. Recently, interferon has been used for treatment of chronic type C hepatitis. However, it is very difficult to determine when HCV is eliminated from the liver, even if hepatitis C virus RNA is not detected in serum. Immunostaining of HCV-NS5 antigen in liver biopsy sections may be helpful in evaluating the cessation of HCV replication. (Hepatology 1994;19:265–272).

Detection of antigens related to hepatitis C virus RNA encoding the NS5 region in the livers of patients with chronic type C hepatitis

Hepatitis C virus is a positive single-strand RNA virus distantly related to flaviviruses. Therefore RNA replicase, an RNA-dependent RNA polymerase, may be essential for the replication of hepatitis C virus, as well as other RNA viruses. In this study we synthesized the recombinant polypeptide (HCV-NS5 antigen) with a 576 bp cDNA encoding a part of the NS5 region of the HCV genome that has the Gly-Asp-Asp motif. The antibody against this polypeptide was obtained from rabbit serum. In Western-blot analysis with NS5 IgG HCV antibody, an 84-kD protein was clearly detected as a single band in the microsomal fraction but not in the nuclear and mitochondrial fractions or in the cytosol fraction. Immunohistochemically, HCV-NS5 antigen was clearly stained in the cytoplasm of hepatocytes but not in the nucleus or cell membrane. Moreover, as determined on immunoelectron microscopy, HCV-NS5 antigen was demonstrated with fine granular distribution along the endoplasmic reticulum but not in other organelles, including the nucleus and mitochondria. Immunoreaction in other cell types was negative. These results indicate that replication of HCV may occur only in hepatocytes and that HCV-NS5 may be produced in the endoplasmic reticulum of these cells. HCV-NS5 antigen was stained only in the livers of hepatitis C virus-positive patients but not in sections from patients with chronic type B hepatitis or alcoholic fibrosis. In chronic type C liver disease, the overall detection rate of HCV-NS5 antigen was 56% (33% in chronic persistent hepatitis, 52% in chronic active hepatitis and 86% in cirrhosis).(ABSTRACT TRUNCATED AT 250 WORDS)

Alcoholic hepatitis and fibrosis in rats

Alcoholic hepatitis and fibrosis in rats

Serum bile acids and cholestasis in alcoholic hepatitis. Relationship with usual liver tests and histological features

Cholestasis is a biochemical and/or histological feature observed in some patients with alcoholic liver disease and is mainly related to alcoholic hepatitis. Accumulation of bile acids in the liver could be pathogenic in alcoholic hepatitis. The aim of this study was to assay serum bile acids in patients with alcoholic hepatitis and to assess the relationship between these parameters, the usual liver tests and the histological features of alcoholic hepatitis. Thirty-six patients (median 51 years, 19 females and 17 males) with biopsy-proven alcoholic hepatitis were included in the study. Cirrhosis was present in 27 patients. Serum bile acids were assayed by high performance liquid chromatography. Three histological scores (alcoholic hepatitis, fibrosis, and cholestasis) were established on each liver sample by two independent pathologists. Serum bile acid concentrations were increased in 35 patients (97%). The median concentration of total serum bile acids was 41.6 mumol/l (range 3-293), with an increase in primary bile acids (95.7% of total bile acids), mainly chenodeoxycholic acid (median 27.5 mumol/l, range 3-184). In contrast, serum bilirubin levels were increased in only 26 patients (72%). Histological cholestasis was present in 14 patients (38%). There was no significant correlation between the alcoholic hepatitis and cholestasis scores (r = 0.01, p = 0.9). A significant correlation was noted between the alcoholic hepatitis score and serum total bile acid (r = 0.34, p = 0.04), cholic acid (r = 0.38, p = 0.03) and chenodeoxycholic acid (r = 0.32, p = 0.05) levels.(ABSTRACT TRUNCATED AT 250 WORDS)