bluuebunny/crossref_metadata_2025 huggingface dataset:
National Toxicology Program Nomenclature for Hepatoproliferative Lesions of Rats
Authors: Maronpot Robert R. , Montgomery Charles A. , Boorman Gary A. , McConnell Ernest E.
Published: September 2009

Diagnostic criteria for hepatoproliferative lesions of Fischer 344 rats are presented to permit more complete categorization of the spectrum of lesions observed in two-year chemical carcinogenicity studies. A nomenclature recently adopted by the National Toxicology Program differs from previous classification schemes in that hepatocellular hyperplasia and hepatocellular adenoma are to be used for lesions which were previously combined under the diagnosis of neoplastic nodule. The term hyperplasia is reserved for proliferative lesions that are perceived to be secondary, nonneoplastic responses to degenerative changes in the liver. Foci of cellular alteration, hepatocellular adenoma, and hepatocellular carcinoma are believed to represent a spectrum of changes that comprise the natural history of neoplasia. This change in nomenclature was made subsequent to a peer review of representative hepatoproliferative lesions from two-year carcinogenicity studies. The revised nomenclature is consistent with traditional pathologic diagnoses for proliferative lesions in other epithelial tissues and should facilitate the interpretation of conventional toxicity and carcinogenicity studies in rats. Morphologic features of other selected rat liver lesions are also presented.
Scanning and Transmission Electron Microscopic Observations of the Acute Morphological Response of the Mouse Urinary Bladder to 4-Ethylsulfonylnaphthalene-1-Sulfonamide
Authors: Frith Charles H. , Ayres Paul H. , Shinohara Yoshitaka , West Robert
Published: September 2009

A total of 75 BALB/cStCrlfC3H/Nctr male weanling mice were administered either 0 or 250 ppm of 4 ethylsulfonylnaphthalene-1-sulfonamide (ENS) in the diet for periods up to 14 days to evaluate the early morphological changes of the transitional epithelium of the urinary bladder with scanning (SEM) and transmission (TEM) electron microscopy. Primary TEM changes included hyperplasia of the epithelium, loosening of the intercellular junctions, autophagic vacuoles and electron dense granules in the mitochondria. Primary SEM changes included sloughing of epithelial cells, irregularity in the size and shape of the transitional epithelial cells and the presence of microvilli. Although pleomorphic microvilli were present after only three days of treatment with ENS, it appears that they are a transient observation in a series of morphological changes. The reversibility or transient nature of the pleomorphic microvilli may indicate that they are an acute toxic response and may not necessarily indicate a preneoplastic change.
Pancreatic Lesions Induced by a Single Dose of N-nitrosobis(2-oxopropyl)amine and Selection by Resistance to Cytotoxicity
Authors: Scarpelli Dante G. , Subbarao Vadrevu , Rao M. Sambasiva
Published: September 2009

The experiments reported here were undertaken to ascertain whether the principle of the selection and rapid emergence of carcinogen-initiated cells based on their resistance to cytotoxicity demonstrated in rat liver is applicable to hamster pancreas. Hamsters injected with a single dose of 70 mg/kg body weight of the pancreatic carcinogen, N-nitrosobis(2-oxopropyl)amine, were subjected to pancreatic injury and regeneration induced by DL ethionine followed by methionine rescue while on a continuous daily regimen of 10 mg/kg body weight of N-nitroso-2,6-dimethylmorpholine by gavage. Randomly selected animals were killed weekly from the 4th week following the induction of regeneration through the 10th week. This carcinogenic schedule significantly decreased the time of emergence of cell injury, cell death, and proliferative, preneoplastic, and neoplastic lesions of the exocrine pancreas. Carcinoma in situ and invasive adenocarcinoma of pancreatic ducts appeared during the 7th and 8th week after induction of pancreatic regeneration, earlier than results obtained with multiple dose carcinogenesis experiments.
The Heterotopically Transplanted Rat Urinary Bladder as a Model for Detection of Tumor-Promoting Urinary Growth Factor(s)
Authors: Oyasu Ryoichi , Konno Seishi , Stevens Julie , Hayashi Osamu
Published: September 2009

The heterotopically transplanted rat urinary bladder (HTB) was developed in our laboratory as a model to study the role of urine in urinary bladder carcinogenesis. With this model, normal urine was found to enhance urinary bladder carcinogenesis initiated by N-methyl-N-nitrosourea or N-butyl-N-(4-hydroxybutyl)nitrosamine. Two crude urinary components (Fractions I and II) were obtained by gel filtration chromatography; they stimulated ornithine decarboxylase (ODC) in a test bladder carcinoma cell line 804G, and promoted carcinogenesis in the HTB system. Fraction I was found to stimulate growth of 804G cells in vitro. Preliminary data indicate that Fraction I contains at least one, and possibly two heat-stable ODC-inducible and mitogenic components. Further characterization of these components is in progress. The HTB system has been demonstrated to be useful for other investigations; for example, α-difluoromethylornithine, an enzyme-activated irreversible inhibitor of ODC, when instilled repeatedly to the bladder lumen, inhibited tumorigenesis in HTBs.
Some Aspects of the Regulation of Hepatocyte Proliferation
Authors: Nadal Glaude
Published: September 2009

Studies of hepatocyte proliferation during physiologic growth of the rat and after experimental stimulations such as partial hepatectomy and injection of irritants show that an inhibitory mechanism is associated with decreasing proliferation and causes hepatocytes to be less sensitive to mitotic stimuli. An inhibitory glycopeptide linked to a high molecular weight protein, α2-macroglobulin in man, has been characterized. This glycopeptide blocks the cell cycle of hepatocytes at the G1-S transition in vivo.
Synthesis of Heat-Shock Proteins and Tumor Growth
Authors: Bardella Lidia , Cajone Francesco , Cairo Gaetano , Schiaffonati Luisa , Bernelli-Zazzera Aldo
Published: September 2009

Exposure to hyperthermia induces the preferential synthesis of a set of proteins, known as heat-shock proteins. The synthesis of heat-shock proteins has been studied in rat liver cells, and human lymphocytes, and in their neoplastic counterparts. Tumor cells synthesize heat-shock proteins essentially as their normal controls, but the response of ascites hepatoma cells depends on the presence of glucose in the medium. Solid hepatoma slices seem to synthesize some heat-shock proteins constitutively, i.e., before exposure to high temperature. Any possible interpretation of this fact must take into account the growth of tumor cells.
Studies on the Mechanisms by Which Tumor Promoters Stimulate the Growth of Primary Neonatal Rat Hepatocytes
Authors: Romano Flora , Andreis Paola G. , Marchesini Cintia , Paccagnella Lucia , Armato Ubaldo
Published: September 2009

A single exposure to a low concentration (10-10 mol/L) of several tumor promoters, namely 12-O-tetra-decanoylphorbol-13-acetate (TPA), phenobarbital (PB), nafenopin, saccharin, teleocidin, benzoyl peroxide, butylated hydroxytoluene (BHT), dichlorodiphenyltrichloroethane (DDT), lindane, clofibrate, and melittin significantly stimulated DNA synthesis of neonatal rat hepatocytes in 4-day-old primary cultures. These cultures were kept in low-calcium (0.01 mmol/L) HiWoBa2000 synthetic medium, thereby evoking a neoplastic phenotype in otherwise normal (i.e., non-initiated) cells. The simultaneous addition of a single dose of alpha-tocopherol (10-4mol/L) or selenous acid (10-5mol/L), just as that of exogenous superoxide dismutase (SOD) (4), together with each of the above agents fully suppressed the stimulation of hepatocytic DNA synthesis by the xenobiotics. Hence, these findings strengthen the view that superoxide anions (or some other oxidizing compounds) act as the common mediators of the mitogenic effects of various tumor promoters in hepatocytes. Inhibition kinetics studies, in which TPA in a single dose (10-10mol/L) was used as the paradigmatic compound together with several kinds of inhibitors of its activity showed that the early mitogenic effects of TPA, i.e., the commitment of quiescent (GO) hepatocytes and the reentry into active cycling of hepatocytes spontaneously poised at the G1/S boundary, required oxidizing compounds, arachidonate metabolism derivatives, and plasmalemmal calcium-binding sites and transmembrane calcium fluxes. Instead, a later TPAs effect, the flow into DNA synthesis of hepatocytes previously committed to cycle, was shown to be controlled by retinoid-modulable activities, by some product(s) of the lipoxygenase pathway, and again by plasmalemmal calcium-binding sites and transmembrane calcium fluxes. Such results reveal that in the neonatal rat hepatocyte the ability to answer to a single mitogenic stimulus and the metabolic pathways by which this answer is enacted depend upon the mitotic cycle setting of the hepatocytes at the moment of the experimental treatment.
Comparison of Central and Peripheral Nervous System Lesions Caused by High-Dose Short-Term and Low-Dose Subchronic Acrylamide Treatment in Rats
Authors: O'Shaughnessy Donald J. , Losos George J.
Published: September 2009

The effects of high-dose subacute acrylamide treatment of up to 50 mg/kg/day for 4 or 10 d were compared to those of subchronic exposure, up to 12 mg/kg/day for 90 d. In the subacute study, Purkinje cells, long ascending tracts of the spinal cord, optic tract terminal or preterminal regions in superior colliculus, sensory ganglion cells, and distal large-caliber peripheral axons were severely affected. Purkinje cells and fasciculus gracilis changes were the earliest lesions. In the subchronic study, the dominant lesion was confined to the distal peripheral axon, with only minor changes occurring in spinal cord and medulla. Paranodal swellings with the characteristic appearance of neurofilament aggregations were not seen. This morphological study suggests a significant difference between high- and low-dose acrylamide-induced lesions. If there is a reduced tendency for long-term low-dose acrylamide exposure to produce CNS lesions, the risk of irreversible nervous system damage would be less than that predicted from subacute studies.
New Data on Kinetics of Lipid Peroxidation in Experimental Hepatomas and Preneoplastic Nodules
Authors: Dianzani Mario U. , Poli Giuseppe , Canuto Rosa A. , Rossi Maria A. , Biocca Maria E. , Biasi Fiorella , Cecchini Giovanni , Muzio Giuliana , Ferro Margherita , Esterbauer Hermann
Published: September 2009

Lipid peroxidation has been found decreased in several hepatomas. The decline has been shown already at the level of preneoplastic nodules obtained after DEN treatment of rats. A substantial exception is represented by the hepatoma cell line MH1C1, deriving from a slightly deviated Morris tumor. Most of the described experiments estimated lipid peroxidation levels in terms of malonaldehyde production by the thiobarbituric acid test. It is now clear that this test does not account for several other aldehydes produced during lipid peroxidation. We now investigated by high performance liquid chromatography (HPLC) the whole range of non-polar aldehydes produced by tumor homogenates and by preneoplastic nodules both in basal conditions and after stimulation with ADP-iron or ascorbate. It was reduced in the preneoplastic nodules as well as in the DEN-induced hepatoma. The susceptibility to the prooxidant effect of ADP-iron or ascorbate was strongly decreased in all hepatomas as well as in preneoplastic nodules. It has been recently published that hepatoma cells are more susceptible than normal liver to the toxic action of aldehydes. This was attributed at least in part to the decreased activity of aldehyde dehydrogenases, as well as to their different distribution in tumor cells. A deeper study on aldehyde metabolism in hepatomas has shown that alcohol dehydrogenase and NADPH-aldehyde reductase also are markedly decreased in Yoshida hepatoma cells and the MH1C1 cell line. However, glutathione transferase, that can use hydroxynonenal as a substrate, is strongly decreased in Yoshida hepatoma cells but not in MH1C1 cells.
Induction of Stress Proteins by Hyperosmolarity in Normal and Transformed Cells
Authors: Borghetti Angelo F. , Petronini Pier Giorgio , Piedimonte Giuseppe , Silvotti Lucia , Tramacere Mariarosaria
Published: September 2009

The synthesis of at least three proteins, with molecular weights of approximately 87, 70, and 53 kd, was enhanced following the exposure of chick embryo fibroblasts to hyperosmolar shock of 30 min at 0.6 osM. Two of these proteins, the 87 and 70 kd, comigrated on one-dimensional gel electrophoresis with the stress proteins induced by heat shock after 30 min at 44°C. In 3T3 cells, the hyperosmolar shock enhanced the expression of two proteins of 88 and 52 kd, whereas the heat shock increased the synthesis of several new polypeptides including the 88 and 52 kd mw. In SV40-transformed 3T3 cells the synthesis of two proteins of 72 and 69 kd was enhanced by heat shock, but no change of the protein pattern was recorded after the hyperosmolar shock.
4-Hydroxynonenal and Other Aldehydes Produced in the Liver In Vivo After Bromobenzene Intoxication
Authors: Benedetti Angelo , Pompella Alfonso , Fulceri Rosella , Romani Andrea , Comporti Mario
Published: September 2009

4-Hydroxynonenal (4-HNE) has been identified as one of the most reactive products in a series of toxic aldehydes originating from lipid peroxidation of cellular membranes. The possibility that this aldehyde plays a role as one of the mediators of the cellular injury induced by pro-oxidants is currently investigated. Mice intoxicated with bromobenzene showed levels of lipid peroxidation in the liver that exceed those induced by hepatotoxic haloalkanes CCl4 and BrCCl3. Hence, we have searched for the presence of 4-HNE and other lipid peroxidation products in the liver of bromobenzene-poisoned mice. We looked for 4-HNE in liver extracts as either free aldehyde or its 2,4-dinitrophenylhydrazone derivative. Using thin-layer chromatography (TLC) and high pressure liquid chromatography (HPLC) we obtained well resolved peaks, corresponding to the standard aldehyde or its 2,4-dinitrophenylhydrazone derivative, respectively. 2,4-dinitrophenylhydrazone derivatization was also used to determine the total carbonyl content in the liver of the intoxicated animals. Three fractions of hydrazones, according to their different polarity (“polar”; “non-polar carbonyls, fraction I”; and “non-polar carbonyls, fraction II”), were obtained using TLC. The UV-visible spectra were recorded for quantitative evaluation. Further fractionation of “non-polar carbonyls, fraction II” provided a fraction containing several alkanals and alk-2-enals, which were analyzed and identified by HPLC. Furthermore, protein bound carbonyls were determined in the liver of the intoxicated animals.
Different Factors Possibly Involved in Post-translational Regulation of Ornithine Decarboxylase Activity
Authors: Zuretti Maria Franca , Gravela Enrico , Papino Fulvia
Published: September 2009

Present results concern a microsome-bound enzymatic system which has been recognized as responsible for the rapid inactivation in vitro of ornithine decarboxylase (ODC). Two different models have been investigated: a) rat liver after a single thioacetamide administration, and b) the 3924 A Morris hepatoma. In both these models we observed variations in the microsome-bound ODC-inactivating capacity. In parallel, changes in ODC properties were observed. The possibility of a causal relationship between the two events is discussed. The actual role of the microsome-bound ODC-inactivating system, in ODC activity regulation in vivo cannot be established, but it remains as a fairly plausible working hypothesis.
Further Studies on the Tumor-Initiating Activity of the Beta-Blocker DL-ZAMI 1305
Authors: Pacces Nicoletta , Braga Marina , Zavanella Teresa , Presta Marco , Ragnotti Giovanni
Published: September 2009

The purpose of this study was to evaluate the initiating activity of the hepatocarcinogen beta-blocker DL-1-(2-nitro-3-methyl-phenoxy)-3- tert-butylamino-propan-2-ol (DL-ZAMI 1305) by the initiation-promotion protocol of Pereira. Female Wistar rats were given a single dose 150 mg/kg of body weight of DL-ZAMI 1305 by gavage 24 hours before or 24 hours after partial hepatectomy. One week later rats were given phenobarbital (0.05%) in the diet for a period of 7 weeks. DL-ZAMI 1305-treatment resulted in the appearance of γ-glutamyltranspeptidase foci and of other preneoplastic lesions in all animals. Preneoplastic lesions were also present in a fraction of DL-ZAMI 1305-treated animals not subjected to partial hepatectomy, whether given or not phenobarbital. Results obtained in a separate experiment demonstrated that DL-ZAMI 1305-treatment inhibits cell proliferation and induces DNA damage in the regenerating rat liver. The results of this study clearly demonstrated that the beta-blocker DL-ZAMI 1305 is an initiating carcinogen for the liver of female Wistar rats.
Studies on DNA Methyltransferase and Alteration of the Enzyme Activity by Chemical Carcinogens
Authors: Cox Ray
Published: September 2009

DNA in mammalian cells is enzymatically methylated at the 5-position of cytosine via S-adenosylme-thionine and DNA methyltransferase. Several chemical carcinogens have been shown to inhibit this reaction, altering DNA methylation. We have been studying the mechanism by which carcinogens alter the methylation of DNA in order to better understand the cellular regulation of DNA methylase activity and to understand the role, if any, of DNA methylation in the carcinogenic process. We have utilized an in vitro assay for DNA methylase isolated from purified rat-liver nuclei. Ethionine, a liver carcinogen, given to rats 17 hr after partial hepatectomy inhibited the incorporation of [methyl-3H]-methionine into 5-methylcytosine residues of DNA. DNA isolated from these ethionine-treated rats was able to accept methyl groups from S-adenosylmethionine 8 times more than control DNA. It was further demonstrated that S-adenosylethionine competitively inhibited the DNA methylase resulting in hypomethylated DNA. N-Methyl-N-nitro-N-nitrosoguanidine reacted with the DNA methylase at the sulfhydryl sites inactivating the enzyme. Methylnitrosourea did not react directly with the methylase enzyme, but when reacted with DNA, the DNA methylase activity was inhibited by the carcinogen alkylated DNA. Sodium selenite also inhibited the enzyme non-competitively with a Ki of 6.7 μM. 5-Azacytidine prevented the 2 to 3 fold increase in DNA methylase seen 2 days following partial hepatectomy. All of these data with various carcinogens, altering DNA methylation by different mechanisms, support the hypothesis that DNA methylation plays a role in the initiation of carcinogenesis.
Immunogenetics of Development and Its Relationship to Carcinogenesis
Authors: Gill Thomas J. , Kunz Heinz W. , Melhem Mona , Rao Kalipatnapu N. , Shinozuka Hisashi , Hassett Andrea L. Cortese , Stranick Kimberly S. , Locker Joseph
Published: September 2009

We have been exploring the classical idea that there is a close relationship between embryogenesis and carcinogenesis. The clinical literature provides evidence for a relationship among the presence of developmental anomalies, an increased incidence of cancer, and genes linked to the major histocompatibility complex (MHC). This relationship was explored experimentally by testing the possibility that rats having the growth and reproduction complex ( grc), which causes developmental abnormalities, might be more susceptible to the development of cancer following exposure to a chemical carcinogen. Two groups of animals which were genetically similar except for the presence or absence of the grc were fed the carcinogen 2-aminoacetylfluorene. The animals carrying the grc developed the morphological changes associated with early cancer of the liver, whereas their normal counterparts did not. These susceptible animals also had a number of biochemical alterations, especially in cholesterol biosynthesis, which may be associated with the development of cancer. Molecular analysis of genomic DNA by blot hybridization revealed restriction fragment length polymorphism differences between the DNA from grc and grc+ rats, and these differences may be related to the differential susceptibility of the strains to cancer.
Focal Eosinophilic Hypertrophic Cells of the Rat Pancreas
Authors: Chiu Taisan
Published: September 2009

Focal spontaneously occurring eosinophilic hypertrophic cells, histomorphologically resembling oncocytes and hepatocytes, were observed in the pancreas from 11 (0.3%) of 4,177 Sprague-Dawley rats, 3 to 26 months old. The age of these eleven rats ranged from 13 to 25 months. The incidence was not different between sexes. The altered cells were usually seen in the peri-insular area. Two types of foci, uniform and pleomorphic, were classified on the basis of uniformity of size of the altered cells within a focus. The origin and nature were not determined for the altered cells/foci reported in this study but they were not considered to be neoplastic.
Immunity, Viral Pathology and Assessment of Immune Dysfunction in Virology and Toxicology
Authors: Krakowka Steven
Published: September 2009

General patterns of tissue injury are recognized as characteristic for certain groups of viruses and certain types of host/virus interactions. Acute viral infections, limited in time and space, tend to be accompanied by florid but brief virus replication cycles in tissues and lesions or signs of disease are largely attributable to the cytolytic effects of the virus on host cells. Chronic or persistent viral diseases tend to have low (or difficult to detect) levels of infectious virus on tissues and the lesions tend to be dominated by inflammation, antiviral immune responses and/or host tissue proliferation. At the cellular level, 3 general categories of response to viral infection are recognized: acute cellular swelling with eventual cytolysis, persistent infection, and transformation (neoplastic) infection. Acute cellular swelling may be accompanied by syncytial giant cell formation and/or viral inclusion bodies. Cells persistently infected with viruses though morphologically normal, are increasingly recognized as functionally deficient and may eventually display degenerative change. Transformation to neoplastic growth can be both benign or malignant in cellular expression. In vivo, the initial neutrophilic inflammatory response to virus-induced cellular necrosis is transient and is rapidly superseded by virus-specific inflammation mediated by both humoral and cellular factors and supplemented by the numerous inflammation amplification pathways. Vasculitis, a common but frequently unappreciated event, may produce nonspecific tissue damage via hemorrhage and ischemia in addition to providing a mechanism for egress of inflammatory factors into the areas of virus-induced cellular damage. During the healing phase, repair by substitution (e.g., fibrosis and scarring) or by proliferation of uninfected replacement cells may dominate sites of viral infection. Functional effects of viruses upon infected hosts are more common than is ordinarily appreciated and, in general, tend to involve control networks such as the nervous, endocrine and immune systems. In this regard, the similarities of mechanisms and effects of the actions of viruses and chemicals on lymphoid tissue are striking. The net systemic effects of these agents may well be enhanced susceptibility to infectious disease, altered responses to toxic agents, and greater sensitivity to chemically-mediated neoplastic transformation.
Free Radical Injury and Liver Tumor Promotion
Authors: Perera Mohan I. R. , Betschart James M. , Virji Mohamed A. , Katyal Sikandar L. , Shinozuka Hisashi
Published: September 2009

One of the underlying mechanisms of tumor promotion both in the skin and liver involves free radical mediated injury to informational macromolecules of target cells. A choline-deficient (CD) diet, which is an efficient liver tumor promoter, induces peroxidative damage of liver cell membrane lipids. By modifying components of a CD diet, we have shown that the efficacy of the promotion is correlated with the extent of lipid peroxidation. The substitution of fats in a CD diet with predominantly polyunsaturated fat and the addition of methapyrilene to a CD diet enhances membrane lipid peroxidation and the promoting effects. An antioxidant (BHT) and hypolipidemic peroxisome proliferators (BR.931 and DEHP) suppress both of these effects. Contrary to these findings, phenobarbital did not induce membrane lipid peroxidation, and its addition to a CD diet inhibited the diet-induced lipid peroxidation, though such a combination exerted a stronger promoting action. Thus, a CD diet and phenobarbital exert their promoting actions through different mechanisms. The consequence of membrane lipid peroxidation in the liver cells induced by a CD diet may be multiple. Our recent study of surface membrane insulin receptors of liver cells of rats fed a CD diet showed a decrease in number and an enhanced binding affinity leading to altered responsiveness of liver cells to insulin mediated glycogen synthesis. It is suggested that CD diet-induced lipid peroxidation leads to functional alterations of membrane receptors involved in cell growth control and may thereby exert its promoting action.
Placental Glutathione S-Transferase (GST-P) as a New Marker for Hepatocarcinogenesis: In Vivo Short-Term Screening for Hepatocarcinogens
Authors: Tatematsu Masae , Tsuda Hiroyuki , Shirai Tomoyuki , Masui Tsuneo , Ito Nobuyuki
Published: September 2009

Our laboratory has developed an in vivo short-term screening test for hepatocarcinogens based on quantitation of γ-glutamyl transpeptidase (γ-GT) positive foci. However, γ-GT positive hepatocytes appear in periportal areas under a variety of circumstances apparently unrelated to hepatocarcinogenesis. Glutathione S-transferase placental type (GST-P), which is hardly detectable in normal rat liver, was recently demonstrated as a new marker protein for preneoplastic liver foci. In experiment I, rats were initially given a single dose (200 mg/kg) of diethylnitrosamine intraperitoneally and 2 weeks later were treated with test compounds for 6 weeks. All rats were subjected to partial hepatectomy at week 3. The long-term development of preneoplastic lesions was followed in rats for 50 weeks. The immunohistochemical investigation of GST-P binding and the histochemical demonstration of γ-GT in serial sections revealed that almost all γ-GT foci were GST-P positive, but 5–10% of GST-P foci could not be detected by γ-GT staining. From week 8, many γ-GT foci partially lost γ-GT activity. However, no comparable disappearance of GST-P was evident in the lesions. All hepatocellular carcinomas (HC) found at week 50 consisted of GST-P positive HC cells. In contrast, 37.9% (11/29) of HC were negative for γ-GT. In experiment II ( in vivo short-term screening test for hepatocarcinogens), rats were treated in the same manner as for experiment I and killed at week 8. Fifty-eight chemicals were investigated for their potential to modify GST-P positive foci development. All hepatocarcinogens and hepatopromoters clearly enhanced the induction of GST-P foci, whereas non-hepatocarcinogens and non-hepatopromoters did not. BHA and acetaminophen inhibited the development of foci. These results suggest the adoption of GST-P as a new and more accurate marker enzyme for rat liver carcinogenesis.
Induction of a Novel Ca2+-Dependent Serine Protease in Rat Liver Treated with Various Promoters of Liver Carcinogenesis
Authors: Enomoto Katsuhiko , Kaneko Aiko , Oyamada Masahito , Oyamada Yumiko , Dempo Kimimaro , Mori Michio
Published: September 2009

The induction of a novel Ca2+-dependent protease in rat liver treated with various liver promoters, as well as its increase in preneoplastic lesions during liver carcinogenesis, was demonstrated. Six groups of male Fischer 344 rats (150 g body weight) were fed separately diets containing one of the following promoters: 0.05% phenobarbital (PB), 0.05% dichlorodiphenyltrichloroethane (DDT), 0.25% ethyl-α-chlorophenoxyisobutyrate (CPIB), 0.5% butylated hydroxytoluene (BHT), 10 ppm 17-α-ethynylestradiol (EE), and 0.05% of the non-promoter diphenylhydantoin (DH). After feeding the indicated diets for 1 week, rats were killed and protease activity in the microsomal fraction of liver tissue was determined using N-benzoyl-L-tyrosine ethyl ester as substrate. The activity of protease increased 3- to 5-fold after treatment with the promoters and compared with normal liver; the non-promoter (DH) induced a slight increase in activity. Hyperplastic nodules were induced according to the method of Solt and Farber. The activity of protease was significantly high in these preneoplastic lesions compared with the surrounding liver tissue. Biochemical characterization of this protease revealed the following properties: 1) high Ca2+ dependency, 2) different molecular weight and optimum pH from previously reported proteases, and 3) preferential distribution in the SER fraction. These results suggest that a novel type of protease is induced specifically in the liver by promoters of liver carcinogenesis. The possible importance of this protease in the carcinogenic process is discussed.
Alteration of DNA Tertiary Structure by Physical and Chemical Carcinogens: Involvement in DNA Repair Processes
Authors: Spadari Silvio , Sutherland Betsy M. , Pedrali-Noy Guido , Focher Federico , Chiesa Maria T. , Ciarrocchi Giovanni
Published: September 2009

Parameters defining the topological state of DNA seem extremely important for describing the reactive state of the same DNA molecules. We have shown that physical and chemical DNA modifying agents alter the tertiary structure of DNA molecules. Variations in the tertiary structure of DNA were studied by one dimensional electrophoresis on an agarose gel of partially relaxed plasmid DNA topoisomers, a technique allowing the measurement of alterations in the degree of supercoiling equivalent to fractions of superhelical turns. Unwinding angles of -10.1° or -8.7“ per pyrimidine or thymine dimer respectively, of -12° per apurinic site, and of -3.4° per methylated purine were obtained by titrating the number of damaged sites necessary to reduce the number of superhelical turns by one in each topoisomer. On the contrary, enzymatic methylation of the C-5 position of cytosine (a modified base present in prokaryotic and eukaryotic DNAs) did not alter the DNA tertiary structure. We have also shown that local alterations in DNA structure caused by UV-irradiation inhibit bacterial DNA topoisomerase I and DNA methylase, and that the topological state of DNA substrate influences the mode of methylation of Hpa II DNA methylase. These findings suggest that the natural topological state of DNA substrate (linear, relaxed, or covalently closed duplex DNA with varying degrees of supercoiling) influences the mode of action of enzymes possibly involved in DNA repair processes, while DNA structural alterations caused by DNA modifying agents might influence DNA repair processes in two ways: either by driving the interaction between repair enzymes and the modified sites of DNA, or by inhibiting or changing the mode of action of enzymes normally acting on unmodified DNA.
Elevated Lipid Peroxidation in Rats Induced by Dietary Lipids and N-Nitrosodimethylamine and its Inhibition by Indomethacin Monitored Via Ethane Exhalation
Authors: Hietanen Eino , Ahotupa Markku , Béréziat Jean-Claude , Bussacchini Valeria , Camus Anne-Marie , Bartsch Helmut
Published: September 2009

The effect of dietary lipids alone or in combination with an administered carcinogen, N-nitrosodimethylamine (NDMA), on whole body lipid peroxidation was studied in rats in vivo. Groups of rats were fed diets containing 2%, 12.5%, or 25% of either saturated or polyunsaturated fat. Lipid peroxidation in individual animals was determined by measuring the concentration of ethane in exhaled air. Increased ethane exhalation was found in rats when the amount of dietary fat was increased from 2% to 12.5%, but animals receiving 12.5% or 25% fat in the diet exhaled ethane at similar rates. Rats consuming polyunsaturated fat exhaled more ethane than those eating saturated fat. In all groups, NDMA administration drastically increased ethane exhalation. Indomethacin completely blocked the increase in ethane exhalation caused by dietary lipids.
Control of Glucose-6-Phosphate Dehydrogenase Deficiency on the Formation of Mutagenic and Carcinogenic Metabolites Derived from Benzo(a)pyrene
Authors: Pirisi Lucia , Garcea Renato , Pascale Rosa , Ruggiu Maria Emilia , Feo Francesco
Published: September 2009

It has been observed that human lymphocytes (HL) and fibroblasts, isolated in vitro from donors carrying the Mediterranean variant of glucose-6-phosphate dehydrogenase (G6PD), show a great decrease in this enzymatic activity, the hexose monophosphate shunt, and the NADPH/NADP+ ratio. This effect is associated with a decreased sensitivity of G6PD-deficient cells to the benzo(a)pyrene (BaP) cytotoxic effect and to a decreased in vitro transformation of BaP-treated fibroblasts. Further, benzo(a)anthracene (BaA)-induced BaP hydroxylase activity is lower in G6PD-deficient cells, when measured in the presence of endogenous NADPH. It has been hypothesized that the NADPH level could be rate-limiting for the NADPH-dependent steps of BaP metabolic activation. To test this hypothesis, the formation of BaP metabolites was studied in normal and G6PD-deficient HL incubated with the carcinogen. HPLC profiles of organic-soluble metabolites revealed that both types of HL produced all the following known BaP metabolites: 9,10-, 4,5- and 7,8-dihydrodiols, quinones, 9- and 3-hydroxy and two peaks of more polar metabolites. There was a great decrease of the various metabolites in the deficient HL. A decrease of total water-soluble BaP metabolites also occurred. HL formed mutagenic metabolites for S. typhimurium TA100 (his-) when incubated with the rat liver S9 fraction. When intact HL substituted S9 fraction, a significant reversed mutation occurred only with normal HL. This could indicate that the NADPH pool is inadequate in G6PD-deficient HL for active BaP metabolism. Accordingly, deficient HL formed lower amounts of BaP:DNA adducts than control during incubation with BaP.
Two-Year Evaluation of Misoprostol for Carcinogenicity in CD Sprague-Dawley Rats
Authors: Dodd David C. , Port Curtis D. , Deslex Paul , Regnier Bernard , Sanders Paul , Indacochea-Redmond Ninfa
Published: September 2009

The carcinogenic potential of misoprostol, a synthetic prostaglandin E, analogue with anti-ulcer potential, was evaluated in CD Sprague-Dawley rats. The compound was given daily by gavage at 24, 240, and 2,400 μg/kg, up to 150 times the daily human dose for 2 years. Necropsies were done on all animals and the incidences of non-neoplastic and neoplastic changes analyzed for significance by life table methods. The only statistically significant non-neoplastic finding was epithelial hyperplasia and hyperkeratosis of the gastric mucosa. These changes, which are characteristic of some prostaglandins, were expected. Other non-neoplastic findings were typical of known spontaneous conditions in this strain of rats. The most frequent neoplasm was the pituitary adenoma, followed by the mammary fibroadenoma, mammary adenoma, mammary adenocarcinoma, and thyroid C-cell adenoma. A rare neoplasm, squamous cell carcinoma of the ovary was found in two rats. There was no evidence that misoprostol is carcinogenic for CD Sprague-Dawley rats.
Twentyone-Month Evaluation of Misoprostol for Carcinogenicity in CD-1 Mice
Authors: Port Curtis D. , Dodd David C. , Deslex Paul , Regnier Bernard , Sanders Paul , Indacochea-Redmond Ninfa
Published: September 2009

Misoprostol, a synthetic prostaglandin E1 methyl ester analogue with anti-ulcer potential, was evaluated for its carcinogenic potential in CD-1 strain mice. The compound was given daily by gavage at 160, 1,600, and 16,000 μg/kg for 21 months. Necropsies were done on all animals and the incidences of non-neoplastic and neoplastic changes analyzed for significance by life table methods. The only statistically significant nonneoplastic compound-related findings were epithelial hyperplasia and hyperkeratosis of the gastric mucosa and hyperostosis of bone in the marrow cavity of sternebrae and femurs. The changes in the gastric epithelium are characteristic of some prostaglandins and were expected. The bone hyperostosis was associated with misoprostol in high dosages, and was considered unique to the mouse. Other non-neoplastic findings were typical of known spontaneous conditions in mice. The most frequent neoplasm was the hepatocellular adenoma followed by lymphosarcoma, lung alveolar carcinoma, and Harderian gland adenoma. Several proliferative lesions of the duodenum were considered to be spontaneous. These were focal avillous hyperplasia, focal atypical hyperplasia, and junctional polyp. There was no evidence that misoprostol is carcinogenic for CD-1 mice.
A Study of the Effects of Vigabatrin on the Central Nervous System and Retina of Sprague Dawley and Lister-Hooded Rats
Authors: Butler William H. , Ford Graham P. , Newberne James W.
Published: September 2009

Vigabatrin ( γ-vinyl GABA), an enzyme-activated, irreversible inhibitor of GABA transaminase, was administered orally to albino Sprague Dawley and pigmented Lister-Hooded rats. A dose-dependent retinal lesion characterized histologically by disruption of the outer nuclear layer was observed in the Sprague Dawley rat but not in Lister-Hooded rats, indicating that this alteration is related to the absence of pigment. The lesion is similar to that induced in albino rats by light and certain drugs. In addition, myelin vacuolation of the brain was observed in both rat strains, consistent with the findings of other toxicity studies with vigabatrin. In all cases, the vacuolation was limited to myelinated tracts and resulted from separation of the myelin sheath at the intraperiod line. There was no evidence of demyelination, axonal degeneration or damage to contiguous structures in the affected areas. The vacuolation is histologically similar to that induced in rats by certain other compounds such as isoniazid, hexachlorophene, and triethyltin, but differs in that it is focal in distribution, it is limited to the brain, and is reversible upon cessation of treatment.
Dietary and Metabolic Manipulations of the Carcinogenic Process: Role of Nucleotide Pool Imbalances in Carcinogenesis
Authors: Rao Prema M. , Laconi Ezio , Vasudevan Shanthi , Denda Ayumi , Rajagopal Sudhasri , Rajalakshmi Srinivasan , Sarma Dittakavi S. R.
Published: September 2009

Perturbations in DNA and/or membranes are considered to be important for the carcinogenic process. A search for nutritional and metabolic means of disturbing the homeostasis of DNA and membranes revealed that nucleotide pools of fer an exciting possibility. An imbalance in nucleotide pools can exert a two-pronged attack on both DNA and membranes. When given to rats, orotic acid, a precursor of pyrimidine nucleotides, results in an imbalance in nucleotide pools (an increase in uridine nucleotides and a decrease in inosine/adenine nucleotides), alterations in both DNA and membranes, and promotion of carcinogenesis in the liver initiated by chemical carcinogens. Agents such as adenine and allopurinol, which inhibit the metabolism of orotic acid and thereby decrease the formation of uridine nucleotides, and galactosamine, which traps uridine nucleotides, inhibited the promotional effects of orotic acid in the liver. These results suggested that orotic acid needs to be metabolized to uridine nucleotides and the creation of a subsequent imbalance in nucleotide pools is important for the promotional effects of orotic acid. To determine whether the creation of a nucleotide pool imbalance is a more general mechanism of tumor-promotion, two lines of approach were investigated. One was to determine the effect of orotic acid on promotion of carcinogenesis in other organs, and the second approach was to determine how to induce nucleotide pool imbalances by means other than orotic acid administration. It is interesting to note that orotic acid promotes carcinogenesis in duodenum initiated by azoxymethane. Regarding the second approach, it became apparent that several metabolic disturbances result in increased orotic acid synthesis and alterations in nucleotide pools. For example, increased administration of amino acids, ammonia, certain disturbances in urea cycle enzymes and/or metabolites, and certain types of liver dysfunction result in increased synthesis of orotic acid. Similarly, folic acid deficiency also results in increased levels of deoxyuridine nucleotide levels.
Effect of Glycine on the Induction of Orotic Aciduria and Urinary Bladder Tumorigenesis in the Rat
Authors: Vasudevan Shanthi , Laconi Ezio , Abanobi Samuel E. , Rao Prema M. , Rajalakshmi Srinivasan , Sarma Dittakavi S. R.
Published: September 2009

The mechanism by which amino acids increase the cellular levels of orotic acid (OA) was investigated. Administration of glycine (2.5 mmoles/100 g) to rats resulted in a 100-fold increase in urinary OA excretion, which was inhibited by pretreatment with cycloheximide or actinomycin D. The induction of OA synthesis from NH4Cl but not from carbamoylaspartate (CA) was inhibited by cycloheximide, indicating that the cycloheximide sensitive step was after the formation of ammonia and before the formation of CA. The glycine-stimulated OA synthesis was not inhibited by acivicin, a potent inhibitor of the cytosolic carbamoylphosphate (CP) synthetase, implicating the mitochondrial CP synthetase in supplying the CP for OA synthesis. Preliminary results indicated that cycloheximide did not inhibit glycine-induced urea synthesis to any significant extent. The results thus suggest that (i) the increased OA synthesis induced by glycine requires a transcription-translation dependent step and (ii) the regulatory step may be the transport of mitochondrial CP to cytosol and/or the synthesis of cytosolic CA. Attempts to determine whether increased exposure of urinary bladder to high concentrations of OA will influence bladder tumorigenesis revealed that chronic administration of glycine (2.5 mmoles/100 g, ip, daily, 5 days a week for 20 weeks) resulted in a 44% increased incidence of hyperplastic, preneoplastic, and neoplastic lesions. Some of these rats also exhibited stones in urinary bladders. The mechanism by which glycine induces tumorigenesis in the urinary bladder is currently being explored.
Induction of the Placental Form of Glutathione S-Transferase by Lead Nitrate Administration in Rat Liver
Authors: Roomi M. Waheed , Columbano Amedeo , Ledda-Columbano Giovanna M. , Sarma Dittakavi S. R.
Published: September 2009

The administration of a single dose of lead nitrate to male Wistar rats caused an increase of a polypeptide in the liver cytosol that cross reacted with the anti-rat antibody of the placental form of glutathione S-transferase (GST-P). GST-P appeared when doses of lead that induced liver cell proliferation were given (5 and 10 micromoles/100 g of body weight). Recently, it has been shown that rat hepatic nodules also exhibited an increased content of the placental form of GST-P. The induction of GST-P by lead together with other biochemical effects exerted in the liver by this metal, suggests that some chemicals may induce in rat liver a biochemical pattern similar, in some aspects, to that exhibited by carcinogen-induced hepatocyte nodules.
Two-Stage Liver Carcinogenesis in the Mouse
Authors: Porta Giuseppe Della , Dragani Tommaso A. , Manenti Giacomo
Published: September 2009

A two-stage protocol for studying liver carcinogenesis was applied to the mouse. The protocol includes the treatment of 7-day-old mice with a single low dose of an initiating agent (diethylnitrosamine, NDEA), promotion starting after weaning and lasting about 20 weeks, and histologic analysis, at 30 weeks of age, of hepatocellular nodules onH&Estained sections. A stereologic analysis of results allows the evaluation of nodule frequency and size. Using this protocol in B6C3 and B6C mice, we have identified the promoting activity of the phenobarbital-like enzyme inducer, 1,4-bis[2-(3,5-dichloropyridyloxy)]benzene (TCPOBOP), which was found to be a strong hyperplaseogenic agent for mouse liver. These studies also indicated that the different susceptibility to hepatocarcinogenesis in B6C3 and B6C mice may be related to a higher susceptibility of B6C3 than B6C initiated liver cells to growth stimulation. A long-term study showed that B6C mice have a low incidence of spontaneous liver tumors but are susceptible to chemical hepatocarcinogenesis and, therefore, they may be an alternative model to B6C3 mice in carcinogenesis bioassays.
Temporal Aspects of Bladder Carcinogenesis
Authors: Vineis Paolo , Estève Jacques
Published: September 2009

A case-control study on 461 males with bladder cancer and 566 hospitalized male controls (all under age 70 and residing in the province of Turin, northern Italy) has been re-analyzed in an attempt to elucidate the role of different temporal variables. Both duration and independently cessation seem to play an important role in the carcinogenic activity of tobacco, whereas occupational exposures to aromatic amines were related to the age at first exposure. Relative risks for current smokers who have smoked less than 20, 20–29, and 40 plus years were 2.8, 6.5, and 8.2, respectively; for ex-smokers such estimates were approximately halved. In the case of occupational exposures, relative risks were 6.7, 3.1, and 1.5 for first exposure ages of less than 25, 25–34, and 35 plus years, respectively.
Effect of 4-Hydroxynonenal on c-myc Expression
Authors: Barrera Giuseppina , Martinotti Stefano , Fazio Vito , Manzari Vittorio , Paradisi Luciana , Parola Maurizio , Frati Luigi , Dianzani Mario U.
Published: September 2009

The 4-hydroxynonenal aldehyde (HNE), a product of lipid peroxidation with high biological activity, inhibits cancerous growths in vivo and in vitro. The mechanism by which this aldehyde acts is not yet understood. The c-myc oncogene seems to be involved in the regulation of cellular multiplication and transformation. We evaluated the c-myc expression and the RNA, DNA and protein synthesis in K562 cells. These cells were incubated for 1 hour in presence of several aldehyde concentrations (range 5.10-7 to 10-4), then washed and kept for 20 hours in a growth medium until used. HNE inhibited both the nucleic acids and protein synthesis in a dose dependent manner, and c-myc expression was evaluated in the K562 cells after incubation with 10-4m or 10-6m HNE. HNE inhibited c-myc expression only at the highest dose. These preliminary results may suggest that the inhibition of c-myc expression is related to nucleic acid synthesis inhibition following HNE exposure.
Autoimmune Mechanisms in Patients and Animal Models
Authors: Talal Norman
Published: September 2009

Autoimmune diseases in patients and animal models are reviewed from the standpoint of immunoregulatory defects which predispose to autoimmunity. New information is provided about autoantibodies which indicates, on the basis of idiotypic cross-reactivity, that they may be closer to normal antibodies than previously thought. The importance of estrogens as a predisposing factor and androgens as a protective factor is discussed in relation to modulation of immune response by sex hormones. Similarities between autoimmune diseases and Acquired Immunodeficiency Syndrome (AIDS) are presented and new therapeutic approaches are discussed.
In Vitro and In Vivo Methods in Immunotoxicology
Authors: Krakowka Steven
Published: September 2009

One approach to assessment of the immunotoxic effects of drugs and chemicals upon the immune system is briefly described. Appropriate in vivo and in vitro methods and techniques are delineated. It is likely that programs implemented at individual institutions will be modified to specific circumstances.