Toshiaki Ninomiya

Clinical significance of serum hyaluronic acid as a fibrosis marker in chronic hepatitis C patients treated with interferon‐α: Histological evaluation by a modified histological activity index scoring system

Abstract The aim of the present study was to investigate the histological changes effected by interferon (IFN) treatment and to evaluate the clinical significance of serum hyaluronic acid (HA) as a marker of fibrosis. Forty‐nine patients with chronic hepatitis C treated with IFN‐α were divided into three groups according to the existence of viraemia: sustained complete responders (CR), complete responders with relapse (PR) and non‐responders (NR). Needle biopsy sections of the liver taken before and at the end of IFN treatment were assessed according to the modified histological activitindex (HAI) scoring system. Serum fibrosis markers, including HA, were measured at needle biopsies. Biopsies of CR at the end of treatment showed a significant improvement in fibrosis and necroinflammatory scores. More significant correlation was observed between fibrosis scores and serum levels of HA before IFN treatment (r= 0.607, P < 0.0001) than those between fibrosis scores, on the one hand, and pepride of type III procollagen (PIIIP; r= 0.531, P= 0.0004) or type IV collagen 7S domain (type IV‐C; r= 0.241, P= 0.1062) on the other. Moreover, serum HA levels fell significantly in patients in whom fibrosis improved (P= 0.011). This is the first paper describing the advantages of the modified HAI scoring system over others in estimating the effect of IFN‐α; the results also indicate that serum HA can be useful in monitoring liver fibrosis in chronic hepatitis C patients treated with IFN‐α.

Expression of HNF-1α and HNF-1β in various histological differentiations of hepatocellular carcinoma

Hepatic nuclear factor 1 (HNF‐1) regulates genes in a hepatocyte‐specific manner. It has been previously reported that the ratio of HNF‐1α and HNF‐1β mRNA is related to histological differentiation hepatocellular carcinoma (HCC). In this study, the expression levels of the HNF‐1α and HNF‐1β proteins were analysed relatively and quantitatively in various histologically differentiated HCC and surrounding non‐cancerous tissues, and HNF‐1α binding activity for the AT element of the B domain of the human α‐fetoprotein enhancer was examined. Western blot analysis demonstrated that HNF‐1α protein was expressed at a higher level in well‐differentiated HCC tissues than in the surrounding non‐HCC tissues; on the other hand, the HNF‐1α protein was expressed at lower levels in moderately and poorly differentiated HCCs than in the surrounding non‐HCC tissues. The levels of HNF‐1β expression in well‐differentiated and poorly differentiated HCCs were similar to and higher than those found in the respective surrounding non‐cancerous portions. In binding assays, HNF‐1 binding activity was high in well‐differentiated HCC and lower in moderately and poorly differentiated HCCs. Most well‐differentiated HCC cases showed immunohistochemical expression of HNF‐1α. These findings show that poor histological differentiation of HCC correlates with decreases in the level and activity of HNF‐1α proteins.

Significance of Serum Matrix Metalloproteinases and Their Inhibitors on the Antifibrogenetic Effect of Interferon-Alfa in Chronic Hepatitis C Patients

Objective and Methods: The imbalance between matrix metalloproteinases (MMPs) and tissue inhibitors of matrix metalloproteinases (TIMPs) is considered to be an important determination of deposition and breakdown of the extracellular matrix. To investigate the antifibrogenetic effect of interferon-α (IFN-α) treatment on factors regulating hepatic fibrosis, serum MMP-1, MMP-2, TIMP-1 and TIMP-2 levels were measured by the one-step sandwich enzyme immunoassay in 27 patients with chronic hepatitis C and compared with the histological status of the patients before and at the end of treatment. Results: After 6 months of IFN-α treatment, the histological status of liver fibrosis showed improvement in 9 patients (IF group) and no change or a worsening in 18 patients (NIF group). Compared with pretreatment levels, in the IF group, IFN treatment caused a significant increase in the MMP-1/TIMP-1 ratio. In the NIF group, however, the MMP-1/TIMP-1 ratio tended towards a decrease; moreover, there was not only a significant increase in TIMP-2 levels but also a tendency towards an increase in TIMP-1 levels. Conclusion: These results suggested that an elevated MMP-1/TIMP-1 ratio may ameliorate liver fibrosis by interferon in cases of chronic hepatitis C, whereas elevated levels of TIMP-1 and TIMP-2 might impede improvement.

Expression ratio of hepatocyte nuclear factor-1 to variant hepatocyte nuclear factor-1 in differentiation of hepatocellular carcinoma and hepatoblastoma

BACKGROUND/AIMS Liver-specific protein genes have multiple cis-/trans-acting elements, but those accountable for hepatocytic differentiation are unclear. An AT-rich core sequence (AT motif) is essential as a cis-acting element for the hepatic transcription. Homologous proteins hepatocyte nuclear factor-1 (HNF-1) and variant HNF-1 (vHNF-1) bind to this motif. The ratio of HNF-1 to vHNF-1 mRNA was examined in various liver tissues with respect to their differentiation. METHODS The competitive reverse transcriptional polymerase chain reaction was employed to amplify HNF-1 and vHNF-1 mRNA simultaneously and to examine their expression ratio in total RNA extracted from frozen liver tissues of 37 patients with hepatocellular carcinoma, five patients with hepatoblastoma, and 15 non-neoplastic liver tissues. RESULTS The ratio of HNF-1 to vHNF-1 mRNA was higher in well-differentiated cases than in poorly-differentiated and undifferentiated cases, except that one poorly-differentiated hepatoblastoma displayed a high ratio. Non-neoplastic liver tissues had low ratios similar to poorly-differentiated hepatocellular carcinoma, the reason for which remained unknown. However, chronic hepatitis and liver cirrhosis cases also demonstrated low ratios, and hence degenerative changes themselves displayed no obvious influence on such ratios. Thus, the gene expression of HNF-1 and vHNF-1 seemed to be differentially regulated in neoplastic and non-neoplastic hepatocytes. CONCLUSIONS These results suggested that the ratio of HNF-1 to vHNF-1 mRNA correlated with histological differentiation of HCC and hepatoblastoma.

Histological grading and staging in chronic hepatitis: Its practical correlation

Although the histological features of various causes of chronic liver disease have been well described, usually the inflammatory activity of the disease is important after the cause has been established. Some patients have co‐infection or concomitant liver disease and on occasion it is difficult to decide the treatment. In order to clarify the histological differences, we investigated the inflammatory activity among autoimmune hepatitis (AIH), primary biliary cirrhosis (PBC), chronic hepatitis C (CHC) and chronic hepatitis B (CHB) in a standardized way using the modified histological activity index (HAI). According to the modified HAI, inflammatory activity is divided into four categories; categories A/D explains portal/periportal inflammation and categories B/C explains lobular activity. The inflammatory score of AIH tended to be greater in all categories from the early stage of fibrosis, whereas scores of PBC were lower, except for portal inflammation. Chronic hepatitis C patients had portal or periportal inflammation, and their inflammatory scores were linked to the development of fibrosis. Chronic hepatitis B patients tended to have severe lobular injury, but did not have a relationship between the inflammatory score and their stage. To know the distribution of inflammation using the modified HAI scoring system may be helpful and convenient in evaluating patients with chronic inflammatory liver disease.

Inflammatory pseudotumor of the liver in a patient with chronic hepatitis C: difficulty in differentiating it from hepatocellular carcinoma.

A case of an inflammatory pseudotumor of the liver in a 75‐year‐old female with chronic hepatitis C whose radiologic features simulated that of hepatocellular carcinoma (HCC) is presented. On imaging studies, hypervascularity by CO2 ultrasound (US) angiography, enhancement at an early phase and isodensity at a late phase by incremental dynamic computed tomography (CT), perfusion defect by CT during arteriography (CTAP), and clinical background of hepatitis C virus (HCV) infection strongly suggested HCC. A US‐guided needle biopsy revealed a mainly diffuse and polyclonal proliferation of lymphocytes positive for leukocyte common antigen (pan‐lymphocyte cells), L‐26 (B cell lymphocytes), and UCHL‐1 (T cell lymphocytes), negative for both κ and λ light chains and sparsely distributed neutrophils and histiocytes. No lymphoid follicles were observed. The liver tissue around this tumor showed chronic hepatitis with mild activity and mild fibrosis. These histopathologic findings suggested that the diagnosis of inflammatory pseudotumor of the liver was tenable. As it is difficult to differentiate between inflammatory pseudotumor of the liver and HCC by imaging studies alone, supplemental biopsy, where possible, should be obtained when diagnostic imaging of tumors suggesting HCC is carried out. We emphasize that histopathology is a true gold standard in the diagnosis of this disease.

CEA producing primary hepatic carcinoid

Imaging studies of a hepatic tumor in a 53-year-old woman with elevated serum levels of neuron-specific enolase (NSE), carcinoembryonic antigen (CEA) and 5-hydroxyindole acetic acid (5HIAA) revealed a hypervascular tumor in the right lobe. Grossly, the brownish tumor was measured 13.5x12 cm with four daughter nodules. Microscopically, the majority of these columnar and round tumor cells had ribbon-or rosette-like patterns with the expression of neuroendocrine marker proteins, such as Grimelius, NSE, chromogranin A, and synaptophysin, and moderate expression of CEA but without the expression of cytokeratin nos 7,8,14,18,19 and OV-6; the minority had glandular patterns with a strong expression of CEA but without the expression of cytokeratin nos 7,8,14,18,19 and OV-6. Ultrastructurally, most tumor cells contained populations of electron-dense core granules ranging between 100 and 200 nm in diameter. After hepatectomy, serum CEA, NSE, and 5HIAA reverted to normal ranges and persisted for 19 months. These findings suggested that the diagnosis of primary hepatic carcinoid was tenable and that the tumor might derive from hepatic stem cells which acquired the additional nature of producing CEA without cytokeratins characteristic of hepatocytes or bile duct cells. Some molecular based approaches have attributed unique biological behavior and histogenesis to this carcinoid tumor.

Multiple hypervascular liver nodules in a heavy drinker of alcohol

Abstract  A case of hypervascular nodules in the liver, but without hepatitis B or C virus infection in a 38‐year‐old woman with a history of alcohol abuse is presented. An ultrasound disclosed 1–2‐cm hypoechoic tumors in the right and left lobes. Magnetic resonance imaging showed high‐intensity tumors at both the T1‐weighted and T2‐weighted sequences. Incremental dynamic computed tomography and hepatic angiography revealed hypervascular tumors. Ultrasound‐guided needle biopsy revealed no evidence of hepatocellular carcinoma, metastatic liver cancer, hemangioendothelioma, inflammatory pseudotumors or pseudolymphoma, but demonstrated stellate‐scar fibrosis septa, which contained small unpaired arteries without hyperplasia dividing the nodule. Moreover, marked pericellular fibrosis, neutrophilic infiltration and Mallory bodies were observed in the cytoplasm. There was no evidence of bile duct proliferation. From these findings, the diagnosis of alcohol‐induced fibrosis, distinctly different from focal nodular hyperplasia, was tenable. Further studies may provide insights into the pathogenesis of nodule formation and hypervascularity in heavy drinkers of alcohol.

Response to Interferon-α in Chronic Hepatitis B With and Without Precore Mutant Strain Detected by Mutation Site–specific Assay

Goals: We investigated whether the presence of precore mutant (stop codon mutation at codon 28) affects the response to interferon-α in patients with chronic hepatitis B. Background: Mutations of hepatitis B virus (HBV) may influence the response to treatment. The association of precore mutant with the response to interferon is controversial. Study: Thirty-one Japanese patients with hepatitis B e antigen–positive chronic hepatitis were treated with natural interferon-α. HBV DNA with the precore mutation was assayed in serum using a mutation site–specific assay before and after treatment. Results: Before treatment, precore mutant was detected in 22 cases (group A) and not detected in 9 cases (group B). Serum HBV DNA level before treatment was not different between the 2 groups. At the end of treatment, serum HBV DNA was decreased to undetectable levels in 13% (4 of 31). Six months after treatment, the percentage of cases with loss of hepatitis B e antigen and a decrease in the transaminase level to within the normal range was significantly higher in group B than in group A (67%, 18%, P = 0.015). Conclusions: Chronic hepatitis without precore mutant strain before treatment is more responsive to IFN-α.

Development of multicentric hepatocellular carcinoma after completion of interferon therapy

6 international units (IU) of IFNα, 3 days a week for a total of 24 weeks. After the IFN therapy, the patient demonstrated a normal serum ALT level, and was continuously negative for HCV-RNA, and histology improved from chronic active hepatitis to chronic persistent hepatitis. Follow-up studies with ultrasonography (US) every 3 months and computed tomography (CT) every 6 months revealed no space-occupying lesion (SOL) for 3 years after IFN treatment.US-guided biopsies of two 15-mm hypoechoic SOLs in segments eight (S8) and seven (S7) 34 and 74 months, respectively, after IFN treatment showed well-differentiated hepatocellular carcinoma (HCC). Clinical data, imaging studies, and histologic examinations showed that both tumors were multicentric HCC. Further studies may provide insights into the possible role of HCV in hepatocarcinogenesis in patients demonstrating HCV eradication by IFN treatment.