Seiichiro Kamimura

In situ detection of insulin-like growth factor II (IGF2) and H19 gene expression in hepatocellular carcinoma

AbstractTo assess the relationship between insulin-like growth factor II (IGF2) and H19 gene expression at the cellular level, we have examined the distribution of IGF2 and H19 mRNA by means of in situ hybridization in hepatic malignancies consisting of hepatocellular carcinoma (HCC), cholangiocellular carcinoma (CCC), and metastatic liver cancer (MLC). In HCC, 15 of 27 tumors (56%) and 11 of 27 tumors (41%) demonstrated increased IGF2 and H19 gene expression, respectively. Of 16 HCCs with increased expression of either IGF2 or H19, 10 tumors coexpressed both transcripts at comparable levels. Moreover, the spatiotemporal distribution and the cellular localization of the two gene transcripts were almost identical, suggesting the presence of a reciprocal relation between IGF2 and H19. In addition, 5 HCCs showed increased IGF2 expression without concomitant H19 expression, whereas 1 HCC showed increased H19 expression without IGF2 transcripts. However, 11 HCCs showed no IGF2 or H19 expression. On the other hand, neither IGF2 transcripts nor H19 transcripts were detected in 2 CCCs or 10 MLCs studied. The data suggest that IGF2 and/or H19 gene expression may be characteristic of some HCCs.

Appearance of cross linked proteins in human atheroma and rat pre-fibrotic liver detected by a new monoclonal antibody

A new monoclonal antibody against malondialdehyde (MDA)-treated low density lipoprotein (LDL) was raised using homogenate of human atheroma as immunogen. This antibody, DLH2, was obtained by selecting the clones which did not react to native LDL but did react to copper-induced oxidized LDL (OxLDL). DLH2 showed a greater reactivity to MDA-LDL than to OxLDL. When LDL was treated with various aldehyde containing reagents, treatment of LDL with glutaraldehyde or MDA greatly increased the reactivity to the antibody, while LDL treated with 2,4-hexadienal or 4-hydroxynonenal was not reactive. Among many proteins tested, high density lipoprotein, bovine serum albumin and hemoglobin showed significant reactivity to DLH2 after they were treated with MDA or glutaraldehyde. When low density and high density lipoproteins treated with MDA were subjected to immunoblot analysis, newly formed products larger than the original apolipoproteins were detected with the antibody, suggesting that this antibody recognizes aggregated proteins with divalent short chain cross linkers. The antigenic materials were shown by immunohistochemical analysis to be present in foamy macrophages in human atheromatous lesions. DLH2 antigen did not colocalize either with apolipoprotein B. Furthermore, we found a massive accumulation of the antigenic material in Kupffer cells in the liver of rats treated with alcohol and carbonyl iron, a model of hepatic fibrosis due to oxidative stress. These results suggest the presence of cross linked proteins in damaged tissues.

Case report: eosinophilic colitis with high antibody titre against Ascaris suum.

Eosinophilic gastroenteritis (EGE) is an inflammatory disease characterized by eosinophilic infiltration of the gastrointestinal tract accompanied by varying abdominal symptoms and usually by peripheral blood eosinophilia. Although the precise aetiology of EGE remains to be determined, contribution of allergic process to certain allergens, such as foods, drugs and parasites, has been repeatedly proposed as the pathogenesis of the disease. Here we report on a rare case of a woman who had extensive eosinophilic infiltration in the descending and rectal colon with a high titre of IgG antibody against Ascaris suum. The patient was successfully treated with prednisolone.

Hemochromatosis with HFE gene mutation in a Japanese patient

A case of hemochromatosis associated with HFE gene mutation has never been previously reported in a Japanese patient. A 65-yr-old Japanese woman presenting with primary hemochromatosis underwent HFE mutation analyses, which demonstrated a C282Y mutation, this being the definitive gene mutation of Caucasian hemochromatosis.

CASE REPORT: Insulin ‐ like growth factor II expression in hepatocellular carcinoma with alcoholic liver fibrosis accompanied by hypoglycaemia

This case of hepatocellular carcinoma (HCC) with alcoholic liver fibrosis, which was not associated with hepatitis viruses, was accompanied by hypoglycaemia. The immunoreactive insulin level was low and other hormonal examinations were almost normal. Immunohistochemical studies showed a high level of insulin‐like growth factor II (IGF2) peptide in the HCC section and the size heterogeneity of serum IGF2 investigated by western blot revealed a large form at approximately 15 kDa. These results suggest that the HCC with alcoholic liver fibrosis produced IGF2 and that the hypoglycaemia was caused by tumour‐associated IGF2.

Immunohistochemical evidence of insulin-like growth factor II in human small hepatocellular carcinoma with hepatitis C virus infection: Relationship to fatty change in carcinoma cells

It has recently been reported that insulin‐like growth factor II (IGF‐II) may play a role in the pathogenesis of hepatocellular carcinoma (HCC). We studied the relationship between the expression of IGF‐II and fatty change in human small HCC using immunohistochemical staining techniques. Liver biopsy specimens were obtained from 35 patients with HCC (consisting of 15 patients with fatty change and 20 patients without fatty change). All patients had serum markers for the hepatitis C virus (HCV) and histological findings obtained from non‐tumourous lesions showed liver cirrhosis or chronic active hepatitis. Immunohistochemical staining was performed using a monoclonal antibody against rat IGF‐II. A positive immunoreaction was found in 69% (24/35) of HCC. Insulin‐like growth factor II was immunodetected in 80% (12/15) of HCC with fatty change but only in 60% (12/20) of those without fatty change. In most cases, IGF‐II was not found in hepatocytes from non‐tumourous lesions. We believe this to be the first time that IGF‐II has been detected immunohistochemically in small HCC derived from HCV infection. This growth factor was more frequently immunodetected in HCC with fatty change than without. As insulin is an essential factor for the metabolism of fatty acids, IGF‐II may play an important role in both fatty degeneration and in the proliferation of HCC cells. Furthermore, immunohistochemical IGF‐II staining may contribute to the diagnosis of HCC, particularly in early stages accompanied by fatty change.

Putative Hemochromatosis Gene Mutations and Alcoholic Liver Disease With Iron Overload in Japan

It is well known that alcoholic liver disease is associated with iron overload. To study the role of hemochromatosis gene mutations on the pathogenesis of alcoholic liver disease (ALD), we have analyzed C282Y and H63D mutations on the chromosomes obtained from 95 Japanese alcoholics. Patients were divided in two groups [i.e., 64 alcoholic patients with liver damage (group I) and 31 alcoholics without liver damage (group II)]. In group I, biochemical examinations showed that serum levels of iron and ferritin were significantly high, and unsaturated iron binding capacity levels were low, compared with those of group II. An analysis by means of allele-specific polymerase chain reaction demonstrated that C282Y mutation was not observed in both groups I and II. H63D mutation was observed in only two heterozygotes of group I and in one heterozygote of group II. Results could not indicate the relationship between ALD and these mutations. We speculate that other causes of iron overload may exist in ALD with iron overload.

Hepatic fibrosis in rats fed a liquid diet with ethanol and carbonyl iron

Abstract It is known that iron acts as a co-factor to catalyze lipid peroxidation (LP) induced by an hepatotoxic compound, such as alcohol. To investigate the role of iron in the pathogenesis of alcoholic liver disease (ALD), we developed a new experimental rat model. Male Sprague-Dawley rats were pair-fed ad libitum a liquid high-fat diet containing ethanol (36% of total calories) or isocaloric carbohydrate with or without dietary carbonyl iron (0.5% w/v) for 12 weeks. Serum alanine aminotransferase (ALT) levels were greatly elevated in rats fed a highfat diet containing both ethanol and iron (EtOH-Fe). Morphologically, slight fibrosis with fatty infiltration and occasional iron deposits were found in the liver of rats fed EtOH-Fe. Moreover, type 4 collagen was definitely stained in the liver of the EtOH-Fe-fed group. However, no evidence of fibrosis was seen in the liver of rats other than the EtOH-Fe-fed group. Furthermore, there was no evidence of secondary hemochromatosis in the rat fed EtOH-Fe or a high-fat diet containing iron. The hepatic content of hydroxyproline (HP) was significantly increased in EtOH-Fe-fed rats as compared to rats other than the EtOH-Fe-fed group. Similarly, microsomal malondialdehyde (MDA) levels were relatively elevated in EtOH-Fe-fed rats. These results demonstrate the evidence of a synergistic effect between alcohol and iron in producing alcoholic liver fibrosis through the enhancement of LP. This new rat model (Fukudai Model) may be useful for further studies in the pathogenesis of ALD.

Effects of esophageal varices obliteration by endoscopic variceal sclerotherapy on asialoscintigraphy and liver function test

Background: hepatic functional reserve is determined by the function and number of hepatocytes, as well as hepatic blood flow. In Japan, endoscopic injection sclerotherapy is often performed prophylactically in patients with high-risk esophageal varices. We examined the effects of blocking portosystemic shunts by this treatment on hepatic circulation and hepatocyte function as evaluated by 99mTc-GSA scintigraphy (HH15, LHL15), an examination via asialoglycoprotein receptors on hepatocytes. Methods: forty-nine patients who underwent prophylactic treatment of esophageal varices were divided into two groups; one having esophageal varices alone and the second having other collateral circulation in addition to esophageal varices. Asialoscintigraphy and general liver function tests were performed both before and after treatment in each group and the data were statistically analyzed. Results: In the group having esophageal varices alone, serum total bile acid significantly decreased at 62.2% and ICGR15, HH15, and LHL15 were slightly improved at -10.9, -3.0, and +4.7%, respectively, after sclerotherapy (P<0.01). However, Child-Pughs score was not changed after sclerotherapy. In the group having additional collaterals, no parameters were changed after sclerotherapy. Conclusion: we demonstrate that HH15, LHL15, and ICGR15 are partially influenced by hepatic circulation but are mainly determined by the function of hepatocytes, whereas serum total bile acid is markedly influenced by hepatic circulation.

Differential effects between tauroursodeoxycholic and taurochenodeoxycholic acids in hepatic fibrosis: An assessment by primary cultured Ito and Kupffer cells from the rat liver*

The pathogenesis of hepatic fibrosis in cholestasis is still unknown, except for endotoxaemia. There is a possibility that the elevation of serum bile acids in cholestasis may play an important role in hepatic fibrogenesis due to a reaction to perisinusoidal cells, such as Ito or Kupffer cells. To assess the effects of bile acids, we investigated the cell proliferation and collagen formation of primary cultured Ito cells that were incubated with a Kupffer cell conditioned medium (KCCM) treated with either taurochenodeoxycholic acid (TCDCA) or tauroursodeoxycholic acid (TUDCA) in short‐term (8 h) or long‐term (48 h) cultures. KCCM treated with TCDCA (100 μmol/L) but not with TUDCA increased cell proliferation of Ito cells in short‐term cultures and also partially elevated collagen formation by Ito cells in long‐term cultures. The release of tumour necrosis factor‐α (TNFα) from Kupffer cells was increased by TCDCA in short‐term cultures, but not in long‐term cultures. The release of transforming growth factor‐β1 (TGFβ1) from Kupffer cells was increased by TCDCA in long‐term cultures, but not in the short‐term cultures. TUDCA showed no significant effect on the release of TNFα and TGFβ1 from Kupffer cells. TUDCA or TCDCA itself showed no direct effect on the cell proliferation and collagen formation of Ito cells. In conclusion, these findings are thus considered to show the potentially important role of TCDCA on the development of hepatic fibrosis in the early phase of cholestasis without endotoxaemia.