Search

The role of autophagy in carcinogenesis is controversial and apparently complex. By using mice with hepatocyte-specific knockout of Atg5, a gene essential for autophagy, we longitudinally studied the role of autophagy in hepatocarcinogenesis. We found that impairing autophagy in hepatocytes would induce oxidative stress and DNA damage, followed by the initiation of hepatocarcinogenesis, which could be suppressed by the antioxidant N-acetylcysteine. Interestingly, these mice developed only benign tumors with no hepatocellular carcinoma (HCC), even after the treatment with diethylnitrosamine, which induced HCC in wild-type mice. The inability of mice to develop HCC when autophagy was impaired was associated with the induction of multiple tumor suppressors including p53. Further analysis indicated that the induction of p53 was associated with the DNA-damage response. Tumorigenesis studies using an established liver tumor cell line confirmed a positive role of autophagy in tumorigenesis and a negative role of p53 in this process when autophagy was impaired. Our studies thus demonstrate that autophagy is required to maintain healthy mitochondria and to reduce oxidative stress and DNA damage to prevent the initiation of hepatocarcinogenesis. However, once hepatocarcinogenesis has been initiated, its presence is also required to suppress the expression of tumor suppressors to promote the development of HCC.

Liver cell dysplasia (LCD) is considered a preneoplastic lesion, whose characterization and differentiation from hepatocellular carcinoma (HCC) and from the reactive changes seen in cirrhosis has been controversial. We studied 12 cases of LCD (large cell type) with image analysis techniques (IA) and compared the findings with those of HCC (n = 40), and a spectrum of non-neoplastic hepatic lesions including normal liver and cirrhosis (n = 49). A minimum of 200 Feulgen-stained nuclei were measured from each lesion with the CAS 200 image analysis system. The data were collected with the aid of CellSheet software. Thirty-four variables were measured, including geometric, textural, and photometric nuclear features and DNA ploidy. The data were analyzed with multivariate statistics and a backpropagation neural network (NN). Stepwise statistical analysis selected 22 variables that were statistically significant in the three groups with P values

The present study was designed to demonstrate age- and sex-related differences in immune functions, and to determine whether subchronic elevations in serotonin (5-HT) availability in vivo would alter immune functions assessed subsequently in vitro. Male and female F344 rats (5 and 21 months of age) were administered the 5-HT releaser and reuptake inhibitor, d-fenfluramine (d-Fen), in their drinking water for 30-38 days then killed. The young animals received a higher dose (1.8 mg/kg/day) of d-Fen than the old rats (0.6 mg/kg/day) in order to compensate for age-related decreases in drug biotransformation and clearance. Brain and spleen d-Fen and metabolite concentrations, however, were considerably higher in the young than in the old rats. d-Fen treatment did not affect body weight or fluid intake. Although substantial sex differences in immune function were not discerned, age-related decreases were observed in absolute splenic cellularity, recombinant interleukin-2 (rIL-2) stimulated natural killer (NK) cytotoxicity, LPS stimulated B-cell mitogenesis, and in the level of Ox19 (CD5) positive cells. d-Fen caused an increase in absolute spleen weight and a decrease in absolute splenic cellularity only in the old rats of both sexes. Spleen cells from young male and old female rats receiving d-Fen had relatively more large granular lymphocytes and enhanced baseline and rIL-2 activated killing of YAC-1 cells than their vehicle matched or opposite sex counterparts. The drug also increased Con A-induced T-cell proliferation in young males and LPS induced B-cell proliferation in old females. d-Fen decreased Ox39 (CD25) levels by 19%, but did not affect any of the other phenotypes examined. The results suggest that 5-HT has a selective stimulatory effect on young male and old female NK activity, and that old female rats are more sensitive to the immunological effects of d-Fen than old male rats.

Liposarcoma is a rare mesenchymal malignant tumor, which usually originates in the retroperitoneum and the extremities. Seven cases of primary liposarcoma of the liver have been previously reported. We present the eighth case, which occurred in an adult female patient. Primary liposarcoma of the liver, although extremely rare, must be considered in the differential diagnosis of a hepatic mass that develops in a noncirrhotic liver, especially in patients who are potential candidates for orthotopic liver transplantation. Liposarcoma is an absolute contraindication for liver transplantation.

Hepatitis B is a disease that infects 300,000 people in the United States each year, resulting in 15,000-30,000 cases of chronic hepatitis. Outcomes include death; development of chronic carrier state, chronic hepatitis, fibrosis, cirrhosis, and hepatocellular carcinoma. Hepatitis B surface antigen (HBsAg) carriers, especially those with cirrhosis, are at high risk for development of hepatocellular carcinoma. Histologically, chronic carriers of HBsAg may have a range of degree of tissue changes, ranging from normal, to varying degrees of portal or lobular inflammation, to significant cell injury with widespread necrosis, fibrosis and cirrhosis. Current histological grading systems reflect both graded inflammation and stage of fibrosis.

We have identified a novel human malignancy-associated gene (MAG) expressed in various malignant tumors including glioblastomas and hepatocellular carcinomas (HCCs) and in tumor preexisting conditions such as hepatitis C virus- and hepatitis B virus-induced liver cirrhosis. The expression of MAG was characterized using reverse transcription-PCR (RT-PCR), rapid amplification of cDNA ends PCR, RNA dot blotting, RNase protection assay, and Northern blot analysis. Rapid amplification of cDNA ends PCR yielded a 536-bp MAG fragment in HCC, macroregenerative liver nodules with dysplasia, and liver cirrhosis but not in normal liver or placenta. By RT-PCR, MAG expression was not found in 12 different normal tissues but found in 46 of 51 (90%) premalignant and malignant tissues of various sites. Embryonic liver and brain were positive for MAG expression together with tumors from the same organs, but the corresponding normal adult tissues were negative. By RNase protection assay, MAG mRNA was expressed in the HepG2 liver tumor cell line and in an ovarian carcinoma but not in normal liver. The estimated transcript size from Northern blot analysis was 8.8 kb. This novel gene may play a role in the progression of premalignant conditions and in the development of HCC and other cancers.

To gain insight into liver regeneration mechanisms in fulminant hepatic failure, we compared gene expression of hepatocyte growth factor, its receptor c-met, c-myc, and albumin in human normal (4 cases) and fulminant (14 cases) livers by reverse transcription-polymerase chain reaction. In normal livers, hepatocyte growth factor gene was not expressed, whereas c-met, c-myc and albumin genes were always expressed. In fulminant hepatic failure, hepatocyte growth factor gene was expressed in 1 of 14 cases, c-met in none of 14 cases, c-myc in 10 of 14 cases, and albumin in 3 of 14 cases. By immunofluorescence, c-met protein was revealed in normal but not in fulminant hepatic failure liver tissue. Liver tissue is unlikely to account for high hepatocyte growth factor plasma levels typical for fulminant hepatic failure. Lack of its receptor (c-met) expression may explain a poor response of fulminant hepatic failure livers to exogenous hepatocyte growth factor that normally promotes liver growth and regeneration.

Hepatobiliary and pancreatic mucinous cystadenocarcinomas with mesenchymal stroma are relatively rare neoplasms that occur preponderantly in women, suggesting a role for unidentified sex-specific factor(s) in the pathogenesis of these tumors. We used paraffin tissue immunohistochemical analysis with an appropriate panel of monoclonal antibodies to look for estrogen and progesterone receptors in two cases of hepatobiliary mucinous cystadenocarcinoma with mesenchymal stroma and one case of pancreatic mucinous cystadenocarcinoma. In all three of these cases, the nuclei of tumor stroma and, in the hepatic tumors, the nuclei of tumor epithelium, reacted with both antibodies. These data strongly suggest that a relationship to hormonal functions exists for these tumors. Because of the rarity of these tumors we also investigated the expression of a variety of oncoprotein antigens, epithelial antigens, and cytoskeletal antigens. The oncoprotein antigens, p53 and c-erbB-2, were focally expressed in hepatic and pancreatic tumor epithelium; bcl-2 was focally expressed in hepatic tumor epithelium. Keratin was strongly expressed in most epithelial cells. In addition, epithelial membrane antigen, carcinoembryonic antigen, and chromogranin were focally expressed in epithelial cells. Actin and vimentin were strongly expressed in most stromal cells but not in epithelial cells, and desmin expression was similar but less widespread.

Hepatocellular carcinoma (HCC) is often difficult to diagnose in cytologic material and small tissue biopsies since histomorphologic information is minimal or absent. The potential for misdiagnosis is greatest in attempting to discriminate well-differentiated HCC from dysplastic hepatocytes in cirrhosis. We investigated the feasibility of developing artificial intelligence classification methods based on nuclear image analysis data for use as adjuncts to the morphologic diagnosis of HCC.Ninety hematoxylin-eosin stained histologic slides including 56 with well- to poorly differentiated HCC and 34 showing a morphologic continuum from normal to markedly dysplastic benign hepatocytes were assembled from four laboratories. A relatively inexpensive PC-based image analysis system was used to measure 35 nuclear morphometric and densitometric parameters of 100 nuclei in each specimen. The data were randomized into classification training and testing sets containing equal numbers of benign and HCC samples. Objective diagnostic classification criteria for HCC based on neural networks and multivariate discriminant functions (DFs) were developed for the most discriminatory subsets of morphometric, densitometric, and combined morphometric/densitometric variables as selected by stepwise discriminant analysis of training data.Morphometric parameters provided the best results with the following testing data positive and negative predictive values (PV+ and PV-) for HCC classification: 86.2% PV+ and 81.3% PV- for a linear DF, 85.7% PV+ and 76.5% PV- for a quadratic DF and 100% PV+ and 85.0% PV- for a neural network.Our results demonstrate that nuclear image analysis-based objective classification criteria for HCC can be developed using artificial intelligence methods and that histologic material prepared at different institutions can be reliably classified. Neural networks for HCC classification were superior to linear and quadratic DFs. Morphometric data yielded the best results compared with densitometric or combined morphometric/densitometric data.

In order to study the effects of serotonin (5-HT) on splenic and blood immune functions, male and female F344 rats (5-6 and 21-23 mo old) received the selective 5-HT releaser and reuptake inhibitor, d-FEN (0.6-1.8 mg/kg/day, p.o.) for 30-44 days. d-FEN significantly increased the relative level of splenic as well as peripheral blood large granular lymphocytes (LGL) in not only old females (1.6x) but also young males (2.3x). In addition, d-FEN significantly increased base-line and recombinant interleukin-2 (rIL-2) stimulated YAC-1 killing (NK activity) by young male (1.4x) and old female (1.8x) spleen ocytes. d-FEN only increased (1.7-2.4x) NK cytotoxicity in the old females. The data indicate that in the F344 rat 5-HT modulates the NK component of the immune system in an age and sex dependent manner. The closer correlation of LGL elevation and functional NK activity in the spleen versus peripheral blood may indicate that d-FEN induces a greater compartmental increase in mature LGLs in the spleen than in blood. High concentrations of 5-HT and its metabolite 5-HIAA, 5-HT immunoreactive processes, as well as 5-HT recognition and reuptake sites have been found in the spleen. Morphological and biochemical evidence thus supports the view that 5-HT can play an important role in the regulation of NK-mediated immune functions.

Young (5 mo) and old (21 mo) female Fischer 344 (F344) rats received the serotonin (5-HT) releaser and reuptake inhibitor, d-fenfluramine (d-Fen; 0.6 mg/kg/day, p.o.), in their deionized drinking water for 30–38 days. The control animals had access ad libitum to deionized water. In comparison to the young animals, the old rats showed a significantly higher percentage of splenic large granular lymphocytes (%LGL) and greater concanavalin A (Con-A) stimulated T cell proliferation. The old d-Fen treated rats, furthermore, showed a significant: 1) further increase in %LGL; 2) enhancement of both basal and recombinant interleukin-2 (rIL-2) stimulated natural killer cytotoxicity; and, 3) augmentation of lipopolysaccharide (LPS) induced B cell mitogenesis. d-Fenfluramine did not affect any of the immunological parameters studied in the yound rats, and did not affect the animals’ body weights, 24 h fluid intakes, or the levels of medial frontal cortex monoamines and their metabolites. These data suggest that prolonged 5-HT release and/or reuptake inhibition enhances immune competence in old but not in young female F344 rats.

The sinusoidal endothelial cells of human liver can be identified by light and electron microscopy, but there appear to be no specific immunocytochemical markers of these cells. Among specific markers available for vascular endothelial cells in general, Ulex europaeus I lectin (UEA I) is the most sensitive. In the present study, 37 liver biopsies were examined for UEA I binding and for Factor VIII related antigen (F VIII RAg) to determine if sinusoidal endothelial cells were positive. The material included normal liver, biopsies from patients with cirrhosis and biopsies in a variety of other liver diseases. Three embryonal human livers were also included in the immunocytochemical analysis. Eleven oesophageal rings obtained at mechanical transection for variceal bleeding in cirrhotic patients were used as control tissue. Sinusoidal endothelial cells of normal liver did not stain with UEA I, but six of seven with alcoholic cirrhosis and only one of 25 non-cirrhotic liver specimens (a case of acute hepatitis with bridging necrosis) were positive. In two of the six cirrhoses the sinusoidal endothelial cells were stained for F VIII RAg as well. Embryonal sinusoidal endothelial cells were stained with UEA I but were negative for F VIII RAg. The results of the study confirm that sinusoidal endothelial cells of normal adult human liver are phenotypically different from those lining blood vessels in other sites. In cirrhosis, positive staining may be related to the transformation of hepatic sinusoids into true capillaries and thus be a marker of the severity of physiological disturbance in the liver.