Phenotypic and karyotypic changes induced in cultured rat hepatic epithelial cells that express the “oval” cell phenotype by exposure to N-Methyl-N,-nitro-N-nitrosoguanidine (original) (raw)
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Cellular and molecular changes in the early stages of chemical hepatocarcinogenesis in the rat
Cancer research, 1990
The early cellular and molecular changes in the Solt-Farber model of hepatocarcinogenesis with and without initiation was studied by using histochemical, immunohistochemical, and in situ hybridization techniques. Increased cellularity was observed in the periductal space in both models 32 to 56 h after partial hepatectomy. These periductal cells and Ito cells were the only cells that became labeled with tritiated thymidine in the uninitiated liver model. Forty-five to 60% of the labeled periductal cells were positive for gamma-glutamyltranspeptidase. From the periductal area the cells that were positive for antibody raised against oval cells (OV-6) infiltrated into liver parenchyma and were followed by desmin-positive Ito cells. The number of Ito cells in the uninitiated model 6 days after partial hepatectomy was 3.5 times higher in the area occupied by oval cells than elsewhere in the liver. The first alpha-fetoprotein (AFP)-positive cells appeared either as individual cells or as ...
Cancer Research
Clonal subpopulations of a chemically induced tumorigenic rat liver epithelial cell line were analyzed for their cellular, biochemi cal, and in vitro growth properties and their tumorigenicity after injection into day-old newborn isogeneic rats. The phenotypic properties studied included DMA content; growth rate in culture; activities of 7-glutamyl transpeptidase, NADH diaphorase, pyruvate kinase, glucose-6-phosphate dehydrogenase, and lactate dehydrogenase; ability to grow in calcium-poor medium; and ability to form colonies in soft agar. The results show that none of these phenotypes cosegregates with tumorigenicity and there fore is not reliable as a "marker" phenotype for neoplastic trans formation in cultured rat liver epithelial cells. The poor correla tions, either qualitatively or quantitatively, between paratumorigenic phenotypes and tumorigenicity suggest that neoplastic transformation in these cells involves a specific transforming gene locus or loci and that in vitro paratumorigenic phenotypes are merely epiphenomena of neoplastic transformation and pro gression. This study further reveals that the efficiency of the tumorigenicity assay of cultured rat liver epithelial cells in isoge neic newborn rats can be considerably improved by incubating the cells in medium containing only trace amounts of serum prior to transplantation into the host animals.
Sequential changes in growth kinetics and cellular phenotype during hepatocarcinogenesis
Journal of Cancer Research and Clinical Oncology, 1989
lar adenomas and carcinomas. The fact that the labelling index of the extrafocai liver tissue of N-nitrosomorpholine-treated rats was also significantly higher than that of the normal parenchyma of untreated controls might indicate an involvement of extrafocal hepatocytes, in addition to that of loci of altered hepatocytes, in hepatocarcinogenesis.
Phenotypic Modulation during Tumorigenesis by Clones of Transformed Rat Liver Epithelial Cells
Cancer Research, 1987
From nine clonal subpopulations (strains) of chemically transformed cultured rat hepatic epithelial cells which were tumorigenic when im planted into 1-day-old isogeneic rats, a cell line was reestablished from each tumor and the cellular properties of the tumor-derived cell lines were compared to those of the corresponding progenitor cells that were implanted to produce the tumors. In seven of eight instances, the cellular DNA content of the tumor-derived cells was virtually identical to the DNA content of the respective progenitor cells, but in one case the tumor cells had twice as much DNA as did their progenitor cells. During the development of tumors in viva, other cellular phenotypic properties often underwent considerable, but variable changes. These changes included the activity of •v-glutamyl transpeptidase, the growth properties on plastic surfaces, and the expression of LDH isozymes. Although there was a relative enhancement in the ability of most of the tumor-derived cells to proliferate or to form colonies in calcium-poor medium, several tumorderived cell lines had very low colony-forming efficiencies in media containing either normal or low levels of calcium. The most consistent association between phenotypes expressed in vitro and tumorigenicity was the ability of cells to form colonies in soft agar; all tumor-derived lines expressed this phenotype, and with some of them this phenotype was acquired only during the process of tumor formation in vivo. These results demonstrate that further phenotypic and genotypic alterations may occur in vivo during tumor formation by chemically transformed cultured cells following their implantation into isogeneic animals; and some of the alterations that occur in vivo may be necessary for the complete expression of tumorigenicity. Although anchorage-independent growth capacity cannot be used to predict the tumorigenicity of clones of rat liver epithelial cells chemically transformed in vitro, this growth property appears to be invariably induced prior to or during the formation of tumors in vivo by these cells.
Environmental Health Perspectives, 1983
Basophilic hepatic foci, nodules, and trabecular hepatocellular carcinomas, collectively referred to as focal hepatic lesions, were induced by single injections of 5.0 jig of diethylnitrosamine (DEN) per gram body weight in 15-day-old C57BL/6J x C3HeBIFeJ F, (B6C3 F,) mice. Groups of eight experimental and eight control mice were killed at 3 days and at 1, 2, 4, 10, 20, 28, 36 and 44 weeks after injection. The only observable acute hepatic toxic effect of DEN, a mild steatosis, was noted at 3 days, but this had disappeared by 7 days following injection. Basophilic foci, composed entirely of altered hepatocytes, were first noted, when very small, at 10 weeks. At later times, some of the foci also contained small collections of proliferated ductules, apparently a result of secondary ingrowth from nearby interlobular bile ducts. The hepatocytes within basophilic foci were characterized by their abundant cytoplasmic RNA, a high nuclear to cytoplasmic ratio (two times greater than normal), which gave them a "crowded appearance," and decreased glucose-6-phosphatase activity. During the course of the study, basophilic foci appeared to increase in size and number. Cytologic anaplasia also became more evident, ultimately culminating in the development of typical trabecular hepatocellular carcinomas by 44 weeks. Invasion of hepatic veins by basophilic foci, first noted at 10 weeks, was prominent by 20 weeks and indicated that many of the lesions manifested this characteristic of malignancy well in advance of the anaplastic features that are also diagnostic of hepatocellular carcinoma. The high growth rates of basophilic foci were confirmed by their greatly increased 3H-thymidine labeling indices, which were 20 times greater than background hepatocytes at 20 weeks following DEN injection. Tumor progression during the course of the study was also suggested by a doubling of labeling indices of hepatocytes in the basophilic foci between 20 to 28 weeks. (The term tumor progression is used in a broad biological sense to encompass any or all of the qualitative and quantitative changes describing the stepwise development of initiated cells to highly malignant neoplasms. This definition differs from the more clinical usage which restricts the process to qualitative changes during the late stages in the development of fully autonomous neoplasms.) An analysis of the number and size of transections through basophilic foci and in some cases, actual reconstructions of the foci from serial sections, indicated that, in aggregate, they grew exponentially between 10 to 36 weeks, with a volume doubling time of 2.5 weeks. The combined morphologic and kinetic data support the view that trabecular hepatocellular carcinomas develop from basophilic foci. Because of their ease of quantitation on conventional H&E stained sections, their rather uniformly spherical shapes, and the high proBability of their clonal origin, the induced focal hepatic lesions should provide a useful model for studying tumor growth kinetics during carcinogenesis.
Chemoprevention of hepatocarcinogenesis
Alcohol, 2002
Accumulation of genetic changes characterizes the progression of cells, initiated by carcinogens, to full malignancy. Various epigenetic mechanisms, such as high polyamine synthesis, aberrant DNA methylation, and production of reactive oxygen species, may favor this process by stimulating growth and inducing DNA damage. We observed a decrease in S -adenosyl-L -methionine (SAM) content in the liver, associated with DNA hypomethylation in rat liver, during the development of preneoplastic foci, and in neoplastic nodules and hepatocellular carcinomas, induced in diethylnitrosamine-initiated rats by "resistant hepatocyte" (RH) protocol. Reconstitution of the methyl donor level in the liver by SAM administration inhibits growth and induces phenotypic reversion and apoptosis of preneoplastic cells. A 6-month SAM treatment results in a sharp and persistent decrease in development of neoplastic nodules, suggesting a long duration of SAM chemopreventive effect. Various observations support the suggestion of a role of DNA methylation in chemoprevention by SAM:
Oncotarget, 2017
Hepatocellular carcinoma (HCC) is the fifth most common cancer worldwide and the second cause of cancer-related death. Search for genes/proteins whose expression can discriminate between normal and neoplastic liver is fundamental for diagnostic, prognostic and therapeutic purposes. Currently, the most used in vitro hepatocyte models to study molecular alterations underlying transformation include primary hepatocytes and transformed cell lines. However, each of these models presents limitations. Here we describe the isolation and characterization of two rat hepatocyte cell lines as tools to study liver carcinogenesis. Long-term stable cell lines were obtained from a HCC-bearing rat exposed to the Resistant-Hepatocyte protocol (RH cells) and from a rat subjected to the same model in the absence of carcinogenic treatment, thus not developing HCCs (RNT cells). The presence of several markers identified the hepatocytic origin of both cell lines and confirmed their purity. Although morphologically similar to normal primary hepatocytes, RNT cells were able to survive and grow in monolayer culture for months and were not tumorigenic in vivo. On the contrary, RH cells displayed tumor-initiating cell markers, formed numerous colonies in soft agar and spheroids when grown in 3D and were highly tumorigenic and metastatic after injection into syngeneic rats and immunocompromised mice. Moreover, RNT gene expression profile was similar to normal liver, while that of RH resembled HCC. In conclusion, the two cell lines here described represent a useful tool to investigate the molecular changes underlying hepatocyte transformation and to experimentally demonstrate their role in HCC development.