To study the pathobiology of hepatitis B virus (HBV) infection, HBV DNA was visualized as grains by an in situ hybridization method, using 3H-labelled probe. All three specimens from HBeAg positive asymptomatic HBsAg carrier showed diffuse distribution of HBV DNA. In 63 specimens from chronic hepatitis B, 47 specimens were HBV DNA positive. Nineteen of 47 specimens showed diffuse distribution. In 17 of 47 specimens, HBV DNA was accumulated in the area of 1/3-2/3 of the sections. Eleven of 47 specimens showed spotty distribution. HBV DNA was usually detected in the areas of focal, piecemeal and bridging hepatic necrosis. Less number of grains were visible in the lobule corresponding to patchy marking area observed by peritoneoscopy, compared with other lobules. These findings indicate that main target cells of immunocytes are the hepatocytes undergoing HBV replication, and HBV replication may be suppressed in regenerating area of hepatocytes.

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To clarify the effects of steroid hormone and cyclophosphamide on alteration of viremia of Duck Hepatitis B Virus (DHBV), experimental transmission of DHBV into each 40 ducks treated with steroid hormone and cyclophosphamide was performed.
The mechanism of elongation of viremia by steroid hormone was assumed to be direct effect on activation of viral replication and this effect appered to be age-dependent. The direct effect of cyclophosphamide on activation of viral replication was not significant. It supposed that cyclophosphamide has a role on elongation of viremia by reducing the hepatitis activity in the liver that secondary inhibit the elimination of DHBV.

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IgG class anti-liver membrane antibodies (LM-Abs) were examined by EIA using isolated rat hepatocytes treated with formaline after cultured in the plastic wells. The preliminary tests showed that the antigenisity of the formalinized cells on each plastic wells was homogeneous in quality and quantity, and no substances of nucleous, mitochondria or other cytoskeletones were detected on the wells. An addition of bovine albumin in PBS-buffer was useful for overcoating the formalinized cells and also absorbing the H-D antibodies in the test sera.
In this methods, IgG class LM-Abs were found with significantly high titers in sera of patients with autoimmune chronic active hepatitis. These antibodies were absorbed out most strongly by the hepatic tissues and were inhibited by the first peak fractions of hepatic tissues separated by Sepharose 6B collumn (McFarlane). Indirect immunofluorescence methods showed the linear membrane staining of the isolated hepatocytes by sera of patients with autoimmune chronic active hepatitis.
These results suggested that IgG class antibodies detected in this methods contained the antibodies directed against liver cell membrane.

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Ninety two biopsied livers from patients with alcoholic or non-alcoholic liver disease were studied concerning ALDH isozymes using isoelectric focusing and enzyme assay methods. The incidence of ALDH-I isozyme (low Km for acetaldehyde) deficiency was lower in alcoholic liver disease (2.3%) than non-alcoholic liver disease (47%). ALDH-I consists of three minor bands and ALDH-II (high Km for acetaldehyde) of four minor bands. However, these minor heterogeneities were same in alcoholic and non-alcoholic liver diseases. In alcoholics the activities of ALDH became to be lower as progress of liver injury, but in non-alcoholics there was no such difference. From these results, a role of acetaldehyde in the progression of alcoholic liver disease was suggested.

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The enhancement of the cellular cytotoxicity to hepatoma cell line PLC/PRF/5 by the three kinds of interferon was investigasted. The cellular cytotoxicity was correlated with the DNA synthesis of the lymphocytes. LAK activity to PLC/PRF/5 was little influenced by INFγ in the incubation time of two days or more, whereas within 18 hours, INFγ enhanced the LAK activity. The enhancement of killer activity against to PLC/PRF/5 was observed in INFα and β from 0, namely co-incubation at cytotoxicity assay, to 48 hours preincubation, but INFγ showed an enhancement only in 48 hours preincubation. Preincubation with the combination of IL2 and IFN could enhance the killer activity in order of IL2 to IFN, than opposite order, especially in using by IFNα and β. So the combination treatment of IL2 and IFN for adoptive therapy, especially α and β seemed to be available for clinical evaluation.

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Type III collagen produced in tumor tissues was quantitated by RIA of N-terminal peptide of type III procollagen (P-III-P) in 6 cases of hepatocellular carcinoma (HCC) and 5 cases of metastatic liver cancer (MLC). In HCCs, the amount of P-III-P was 0.511±0.272μg/g Tissue in tumorS and 4.45±7.34μg/g Tissue in cirrhotic portions. In MLCs, it was 1.13±1.13μg/g Tissue in tumors and 0.064±0.069μg/g Tissue in non-tumorous portions. In normal livers, 0.24±0.21μg/g Tissue and in cirrhotic liverS without cancer, 1.83±0.72μg/g Tissue. These liver tissues with cancer were Stained immunoenzymatic-histochemically with a monoclonal antibody againSt type III collagen. Positive staining was observed in tumor cells in 4 caseS of HCC and in one of MLC, in cirrhotic hepatocytes in 5 cases of HCC and in hepatocytes in 2 cases of MLC. In livers with cancer, collagen production was thought to be influenced by the tumor tisSue even in their non-tumorouS portion.

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To make the definite diagnosis of small hepatocellular carcinoma (HCC), we performed US-guided fine needle aspiration biopsy originally deviced by Majima in 103HCC nodules between May. 1985 andNov. 1986. In conclusions;
1) 100% of 103 nodules in 96 cases were histologically confirmed as HCC nodules.
2) The incidence of well-diffenciated HCC increases with the reduction in size of the tumor, in marked contrast, the incidence of moderately or poorly differenciated HCC increases with the enlargement in size of the tumor.
3) Cytological and histological diagnosis were made at the same puncture, however positive rate of cytological diagnosis was only 35.6% in small HCC nodules less than 20mm in diameter.
4) 58.3% of HCC (from 11 to 15mm in diameter) were hyperechoic nodules, and 2 nodules (1.9%) were histologically confirmed as the natural necrotic nodules.
5) In this series, serious complications and skin metastases were not observed.

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We described a method for flow cytometric determination of nuclear DNA content of paraffinembedded liver tissue and discussed a potential pitfall of analysis of DNA histogram from this method in comparison with that from detergent-trypsin method which had been devised for frozen materials.
In order to obtain the best quality of DNA histogram from paraffin-embedded liver tissue, it was necessary for sample preparation to be carried out as follows. Ten percent formalin fixed and paraffinembedded liver tissue was cut at 50μm thickness. Sections were dewaxed with xylene and were rehydrated through a series of ethanol. Single nuclear suspensions were prepared by incubation in 1.0% pepsin, pH1.5, at 37°C for 90 to 120 min with intermittent vortexing. After that, nuclei were treated with RNase at 37°C for 30 min and were stained with propidium iodide.
It was demonstrated that DNA aneuploidy could be detected from paraffin-embedded materials as well as from frozen materials. HoWever, since there were many nuclear fragments in a sample from paraffin-embedded materials, there was a possibility that S-phase fraction was overestimated.

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To investigate the collagen-producing capacity of Ito cells in human hepatic fibrosis, biopsized liver tissues were examined by light and electron microscopic autoradiography using 3H-proline as a tracer. Following biopsies, liver specimens (22cases) were trimmed immediately and incubated at 37°C for 1 hour in the medium containing 200μCi/ml 3H-proline and then chased for 6 hours. After fixation and epon-embedding, thin sections were prepared for light and electron microscopic autoradiography (2-6 Weeks exposure). After 1 hours labelling, numerous silver grains were seen over fibldblasts and Ito cells particularly located in piecemeal necrosis, in addition to over hepatocytes and bile ducts. After a 6 hour chase, silver grains were also over collagen fibrils adjacent to the labelled Ito cells and fibroblasts, indicating incorporation of proline into these cells, and subsequent synthesis and Secretion of collagen. From these results, Ito cells can produce collagen fibers in human hepatic fibrosis, as do fibroblasts in the portal tract.

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In order to re-examine the metabolism of dehydrocholic acid (DHCA), 1g DHCA was intravenously given to two patients with carcinoma of biliary tract, and the reduced metabolites in serum, urine and bile were analyzed. In serum unconj ugated 3α-hydroxy-7, 12-diketo-cholanoic acid (MonoOH-di-K) level was elevated markedly and rapidly after DHCA loading, and reached to about 80μM. While, consecutive increase of the conj ugated 3α, 7α-dihydroxy-12-keto-cholanoic acid (Di-OH-mono-K) level was noted over the couSe of the precedure.
In bile conjugated Di-OH-mono-K occupied more than 90% of total 3α-hydroxy bile acid, while conjugated Mono-OH-di-K was observed as a minor constituent. Therefore, the composition of the reduced metabolites in serum was completely different from that in bile, and it was estimated that mechanism on elevation of Mono-OH-di-K and Di-OH-mono-K in serum could not be identical.
Additionally, it was presumed that elevation of unconjugated Mono-OH-di-K in serum occured due to unknown mechanism except backflow of bile component from biliary tract to systemic circulation.

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Capacity for antipyrine oxidation and urinary excretion of its major metabolites (3-hydroxymethylantipyrine, norantipyrine and 4-hydroxyantipyrine) in 24 hours were studied in six patients with Wilson's disease (all but one were cirrhotic), who were treated with D-penicillamine during more than a year.
Wilson group presented no significant differences from control group on the plasma half-life and clearance of antipyrine, and urinary excretion and % conjugation of the major metabolites. Singificant prolongation of the half-life and significant decreases in the other indexes were demonstrated in posthepatitic cirrhotic group.
Capacity for antipyrine oxidation and hepatic reserve seem to be well preserverd in the Wilson's disease patients with cirrhosis under appropriate treatment, compared with those in the patients with posthepatitic cirrhosis.

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We measured the width and height of lesions on the liver surface of chronic liver diseases of a new peritoneoscopic measuring system, developed with cooperation of Machida Endoscope Co.. This scope emitted diffraction beams in parallel by an argon laser of 488-nm wave length. The diffraction spots on the lesion was clearly observed and pictured under ordinary illumination. Not only the width but also the height of lesions was easily measured by counting the number of spots by the order of 0.1mm. In this study, chronic diseases were classified in chronic aggressive hepatitis 2A (5 cases), chronic aggressive hepatitis 2B (6 cases), early cirrhosis (6 cases) and cirrhosis (27 cases). The results were as follows: 1) The liver surface of chronic aggressive hepatitis 2A was smooth. 2) The width of patch was 1.6±0.4mm in diameter and the depth was 0.1±0.1mm in chronic aggressive hepatitis 2B. 3) The diameter of the nodule was 3.2±0.9mm and the ratio of height to diameter was approximately 0.2 in any cirrhosis.

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Two alcoholic patients with histological features of chronic hepatitis whose liver biopsies showed a decrease in inflammatory activity after abstinence, were reported. Both cases had a history of drinking more than 80g/day of ethanol for more than ten years, and had no history of blood transfusion, nor had HBsAg in their sera. At the first liver biopsies, mild to moderate lymphocyte infiltration in portal tract and piecemeal necrosis were associated with pericellular fibrosis, perivenular fibrosis and fatty metamorphosis. Both thier clinical and biochemical findings strikingly improved after abstinence. In the follow-up biopsies, marked decrease in inflammatory activity was noted after abstinence.
This is the first report in Japan in which an improvement of inflammatory activity of chronic hepatitis in alcoholics after abstinence is recognized. These results suggest that imrhunological mechanism may play a role, at least in a restricted case, in liver injury of alcohlic liver disease.

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A case of primary biliary cirrhosis (PBC) with high levels of serum transaminases like viral hepatitis is reported. The patient was a 37-year-old woman. Her liver function showed higher levels and larger changes of serum transaminases compared with serum biliary enzymes. Anti-mitochondrial antibody (AMA) was positive (160×). Histology of liver biopsy showed the findings of chronic nonsuppurative destructive cholangitis (CNSDC), but showed no piecemeal necrosis. The subtypes of AMA were anti-M2 and anti-M8, but not anti-M4. Though this case showed changes of serum transaminases like chronic active hepatitis (CAH), it was not CAH-PBC mixed type because of negative anti-M4 and no piecemeal necrosis. High levels of serum transaminases of this case seemed to be related to positive anti-M8. The clinical course of this case should be carefully followed.

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A 37-year-old female suffered from PM, asymptomatic PBC and SjS is reported. At her 25 years age (1967) she felt stiffness in the forearms, and was diagnosed as rheumatoid arthritis. Two years later she was diagnosed as PM because of muscle weakness, increased CPK level and muscle biopsy findings. Oral administration of prednisolone brought a marked improvement of the symptoms and laboratory data derived from PM. Liver dysfunction in laboratory data appeared in 1976. Liver biopsy specimen in 1977 showed CNSDC, compatible with stage I of PBC (Scheuer). Two years later xerostomia due to SjS appeared. From 1976 to 1984, she was medicated on d-penicillamine against her pulmonary fibrosis associated with PM. After cessation of d-penicillamine for its side effect, a marked elevation of GPT and ALP level was observed. She is now under observation as an out-patient with asymptomatic condition.

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A case of hypovascular hepatocellular carcinoma in a 55-year-old female was reported. She was admitted to our hospital under the diagnosis of liver tumor. Abdominal angiography showed hypovascurality, and serum level of AFP was less than 5ng/ml. Left lobectomy was performed. The pathological diagnosis of the tumor was Edmondson's grade IV HCC without fenestration of endothelial lining of the blood space, which led us to believe that it was hypovascular by angiography.
It is a matter of course that abdominal angiography is important for the diagnosis of HCC, but for diagnosis of hypovascular liver tumor, various imaging methods make definite diagnosis possible.

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