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Food and Chemical Toxicology : An... Aug 2012Diethylnitrosamine (DEN) is a potent hepatotoxin and hepatocarcinogen in animals and possible in humans. Estrogen has been reported to play a protective role against DEN...
Diethylnitrosamine (DEN) is a potent hepatotoxin and hepatocarcinogen in animals and possible in humans. Estrogen has been reported to play a protective role against DEN exposure. Osteopontin (OPN), a downstream molecular of estrogen, plays a role in many pathophysiological processes. In this study, we evaluate the role of OPN in estrogen-mediated hepatoprotection in DEN-treated mice. DEN was administrated intraperitoneally to C57BL/6 and OPN(-/-) mice. Compared to male mice, female mice exhibited significantly higher hepatic OPN expression with less liver damage 48 h after DEN treatment. Interestingly, enhanced OPN expression was predominantly detected in hepatocytes after DEN treatment. OPN deficiency enhanced the susceptibility to DEN, which was more apparent in females than males. Estrogen-mediated protection against DEN in males was abrogated by OPN deficiency. The protective activities of estrogen could be mimicked by exogenous OPN. Consistent with liver injury, oxidative stress in liver was enhanced with OPN depletion. OPN reduced DEN-induced oxidative stress likely through inhibition of CYP2A5 expression. In conclusion, we demonstrate that OPN may be involved in estrogen-mediated hepatoprotection in DEN-induced liver injury through enhancement of hepatocyte survival and inhibition of DEN biotransformation. Our findings may provide new insight into gender differences in chemical-induced liver injury and related diseases.
Topics: Animals; Diethylnitrosamine; Estrogens; Female; Liver; Male; Mice; Mice, Inbred C57BL; Osteopontin; Real-Time Polymerase Chain Reaction
PubMed: 22609492
DOI: 10.1016/j.fct.2012.05.012 -
Mutation Research Sep 2012Various liver micronucleus assay methods, such as those involving partial hepatectomy, treatment with mitogens, and the use of juvenile animals, have been developed....
Various liver micronucleus assay methods, such as those involving partial hepatectomy, treatment with mitogens, and the use of juvenile animals, have been developed. These assays have been proven to be of high sensitivity and specificity to predict hepatocarcinogenicity of compounds that cannot be detected by bone marrow micronucleus assays. On the contrary, the existing assays have only been evaluated for their use in detecting micronucleus induction in the settings of relatively short-term cell proliferation. However, the integration of in vivo genotoxicity endpoints into routine toxicity studies is increasingly desired from the viewpoint of animal welfare to reduce the number of animals used. In the present study, the rodent hepatocarcinogens diethylnitrosamine (DEN) and 2,4-diaminotoluene (2,4-DAT) were repeatedly administered orally to male Crl:CD (SD) rats (6 weeks old at the beginning of administration) for 5, 14, and 28 days, and changes in the frequency of hepatocytes with micronuclei in liver tissues that had undergone no artificial treatment to accelerate cell proliferation were evaluated. At the same time, a new method of hepatocyte isolation involving the treatment of a portion of the liver with collagenase in a centrifuge tube, without the use of in situ perfusion, was established. The induction of micronucleated hepatocytes was achieved after the repeated administration of DEN for 5 days or longer and of 2,4-DAT for 14 days or longer. Micronucleus frequencies were increased depending on the number of administrations, indicating that micronucleated hepatocytes had possibly remained for a long period of time and accumulated additively. It therefore appears that even in adult rat liver with low mitotic activity, a repeated-dose of a chemical substance for 14 days or longer enables the detection of micronucleus induction. In addition, the establishment of a method to isolate hepatocytes without perfusion using only a part of the liver enables the integration of liver micronucleus assays into general toxicity studies.
Topics: Animals; DNA Damage; Diethylnitrosamine; Dose-Response Relationship, Drug; Liver; Male; Micronucleus Tests; Mutagens; Phenylenediamines; Rats
PubMed: 22677510
DOI: 10.1016/j.mrgentox.2012.05.012 -
Mutation Research Feb 2009We examined the dose-dependency of gene expression changes for 51 genes in mouse liver treated with two N-nitroso genotoxic hepatocarcinogens, diethylnitrosamine (DEN)...
We examined the dose-dependency of gene expression changes for 51 genes in mouse liver treated with two N-nitroso genotoxic hepatocarcinogens, diethylnitrosamine (DEN) and ethylnitrosourea (ENU) by quantitative real-time PCR (qPCR). DEN (3, 9, 27 and 80mg/kg bw) or ENU (6, 17, 50 and 150mg/kg bw) was injected intraperitoneally into groups of five male 9-week-old B6C3F(1) mice and the livers were dissected after 4h and 28 days. Total RNA from pooled livers was reverse-transcribed to cDNA and the amount of each gene was quantified by qPCR. Results were analyzed by hierarchical and k-means clustering and ingenuity pathway analysis (IPA). The most characteristic result was a similar dose-dependency of gene expression changes with DEN and ENU. Twenty-one genes exhibited a distinct dose-dependent increase in expression at 4h for both carcinogens [Bax, Btg2, Ccng1, Cdkn1a, Cyp4a10, Cyp21a1, Fos, Gadd45b, Gdf15, Hmox1, Hspb1, Isg20l1, Jun, Mbd1, Mdm2, Myc, Net1, Plk2, Ppp1r3c, Rcan1 and Tubb2c], although the increase in gene expression due to ENU was generally weaker than that due to DEN. Only Gdf15 showed a dose-dependent increase in expression at 28 days for both carcinogens. The differences between DEN and ENU were in the expression of additional genes (7 for DEN and 8 for ENU). IPA extracted five gene networks: Network-1 included genes related to cancer and cell cycle arrest and associated with Bax, Btg2, Ccng1, Cdkn1a, Gadd45b, Gdf15, Hspb1, Mdm2 and Plk2 and Network-2 was related to DNA replication, recombination, repair and cell death and associated with Cyp21a1, Gdf15, Ppp1r3c, Rcan1 and Tubb2c. The present results show a distinct dose-dependency of gene expression changes induced by DEN and ENU. These changes were associated with cancer, cell cycle arrest, DNA replication, recombination, repair and cell death and were seen not only at 4h but also, for some, at 28 days after administration.
Topics: Algorithms; Animals; Carcinogens; Diethylnitrosamine; Dose-Response Relationship, Drug; Ethylnitrosourea; Gene Expression Regulation; Liver; Male; Mice; Polymerase Chain Reaction
PubMed: 19100860
DOI: 10.1016/j.mrgentox.2008.11.004 -
Zeitschrift Fur Krebsforschung Und... Oct 1977Disulfiram (DSF) delayed the appearance of diethylnitrosamine (DEN)-induced strand breaks in liver DNA of rats. The fragmentation of liver DNA produced by DEN was...
Disulfiram (DSF) delayed the appearance of diethylnitrosamine (DEN)-induced strand breaks in liver DNA of rats. The fragmentation of liver DNA produced by DEN was studied 4 and 24 h after administration of the carcinogen on alkaline sucrose gradients. A single dose of 500 mg/kg DSF given 1 h prior to carcinogen treatment delayed for at least 4 h the DEN--induced strand breaks in liver DNA. A single DSF pretreatment, however, did not protect against carcinogen-induced strand breaks when observed 24 h after DEN injection. It is possible that continuous administration of DSF might inhibit the DEN-produced damage of the genetic material.
Topics: Animals; DNA Repair; Diethylnitrosamine; Disulfiram; Liver; Male; Molecular Biology; Nitrosamines; Rats
PubMed: 145745
DOI: 10.1007/BF00306025 -
Mutation Research. Genetic Toxicology... Sep 2015A repeated-dose micronucleus assay utilizing young adult rat hepatocytes was recently developed to evaluate the genotoxicity. In this assay, accumulation of...
A repeated-dose micronucleus assay utilizing young adult rat hepatocytes was recently developed to evaluate the genotoxicity. In this assay, accumulation of micronucleated hepatocytes (MNHEPs) induced by repeated dosing of genotoxic chemicals is considered to be a key factor in the detection of micronuclei induction. Then, we hypothesized that the period following chemical exposure enable the detection of MNHEP induction in young adult rats, namely that MNHEPs can be generated from chromosomally damaged cells and accumulate following initiation of chemical exposure until sampling. We therefore measured MNHEP induction at 2 or 4 weeks after a single oral administration of 12.5, 50, or 100mg/kg of diethylnitrosamine (DEN) or an intraperitoneal administration of 0.5, 1.0, or 2.0mg/kg of mitomycin C (MMC) to young adult rats. Results showed a statistically significant, dose-dependent increase in the numbers of MNHEPs in DEN- or MMC-treated rats, indicating that prolonged rest period following a single dose of a genotoxic chemical enables the detection of MNHEP induction in the liver of young adult rats. From these results, a single oral administration of 50mg/kg of DEN with a 2- or 4- week rest period can be used as a positive control in repeated-dose liver micronucleus assays. This procedure is superior in terms of labor saving and animal welfare to repeated dosing of DEN.
Topics: Administration, Oral; Animals; DNA Damage; Diethylnitrosamine; Dose-Response Relationship, Drug; Drug Administration Schedule; Hepatocytes; Injections, Intraperitoneal; Micronucleus Tests; Mitomycin; Mutagens; Rats
PubMed: 26338541
DOI: 10.1016/j.mrgentox.2015.07.010 -
Nature Feb 1978
Topics: Adenosine Triphosphatases; Animals; Carcinoma, Hepatocellular; Diethylnitrosamine; Female; Hepatectomy; Liver; Liver Neoplasms; Nitrosamines; Phenobarbital; Phenotype; Rats; gamma-Glutamyltransferase
PubMed: 24178
DOI: 10.1038/271456a0 -
Nanomedicine : Nanotechnology, Biology,... Feb 2017Nano-carriers, especially lipid nanoparticles have been used widely in "a good manner", for instance in the treatment of cancer, by enhancing the targetability and...
Nano-carriers, especially lipid nanoparticles have been used widely in "a good manner", for instance in the treatment of cancer, by enhancing the targetability and reducing required dose. Here in the contrary, we presented a new possibility: nanoDEN, a nanoparticle-packed "bad guy", which is more effective and efficient in generating liver tumor in mice. We have shown that nanoDEN, same as diethylnitrosamine (DEN), induced overexpression of multiple pivotal factors (including COX-2, β-catenin and PCNA) during oncogenesis. Moreover, nanoDEN increased the apoptosis of liver cells compared with DEN alone. This apoptotic effect of nanoDEN is more efficient on normal cells than on cancer cells. Taken into consideration the fact that there are endogenous nanoparticles naturally formed inside our body, our research enlarged our views of all the aspects of oncogenic chemicals, while also established a better method of producing animal model of liver cancer, which has future investigational and therapeutical potential.
Topics: Animals; Carcinogenesis; Cell Proliferation; Diethylnitrosamine; Disease Models, Animal; Humans; Lipids; Liver; Liver Neoplasms; Mice; Nanoparticles
PubMed: 27729235
DOI: 10.1016/j.nano.2016.09.017 -
Cancer Jan 1976Chronic subcutaneous (s.c.) or single intravenous (i.v.) injections of diethylnitrosamine (DEN) to gerbils (Meriones unguiculatus) led to the induction of carcinomas of...
Chronic subcutaneous (s.c.) or single intravenous (i.v.) injections of diethylnitrosamine (DEN) to gerbils (Meriones unguiculatus) led to the induction of carcinomas of the nasal cavities in a high incidence. These neoplasms were multifocal in origin, frequently showing mixed cellular patterns of the following cell types: large cuboidal cells, nonciliated columnar cells, ciliated columnar cells, small cells, and squamous epithelial cells. No sensory cells of the olfactory mucosa or neurofibrillar differentiations were found in these neoplasms, although several showed rosette- and pseudorosette-like formations. Dose and route of administration seemed to influence the site of origin and the patterns of cellularity of these tumors. Whereas chronic s.c. injections resulted in carcinomas of the olfactory region, being mostly composed of large cuboidal cells, single i.v. injections led to the development of carcinomas mainly originating from the respiratory-olfactory mucosal junction and composed preferentially of nonciliated and ciliated columnar cells.
Topics: Adenocarcinoma; Animals; Carcinoma; Carcinoma, Papillary; Carcinoma, Squamous Cell; Diethylnitrosamine; Dose-Response Relationship, Drug; Female; Male; Nasal Cavity; Nitrosamines
PubMed: 1247964
DOI: 10.1002/1097-0142(197601)37:1<346::aid-cncr2820370145>3.0.co;2-# -
Food and Chemical Toxicology : An... Sep 1993The influence of fasting on the potential of diethylnitrosamine (DEN) to initiate liver carcinogenesis was tested in a medium-term assay using the development of...
The influence of fasting on the potential of diethylnitrosamine (DEN) to initiate liver carcinogenesis was tested in a medium-term assay using the development of putative preneoplastic altered foci of hepatocytes (AFH) as the endpoint. Male Wistar rats fasted for 48 hr were given a single ip injection of DEN (200 mg/kg body weight). Partial hepatectomies were carried out at wk 3 and the rats were killed at wk 8. Fasted rats exhibited a small increase in the numbers of AFH with glutathione S-transferase in the placental form and eosinophilic AFH when compared with non-fasted animals. However, after a 6-wk exposure to 0.05% sodium phenobarbital in the diet, there were no differences in the numbers of AFH between fasted and non-fasted animals. Fasting also increased DEN-dependent centrilobular cell necrosis and specifically drug metabolism as indicated in vivo by a decreased time of paralysis of the lower limbs induced by zoxazolamine (40 mg/kg body weight, ip) and by an unaltered sleeping time induced by sodium pentobarbital (40 mg/kg body weight, ip). The results indicate that although fasting during the initiation stage of carcinogenesis increases DEN hepatoxicity, it does not interfere quantitatively with the development of liver preneoplastic lesions.
Topics: Animals; Diethylnitrosamine; Fasting; Immunohistochemistry; Liver; Liver Neoplasms; Male; Rats; Rats, Wistar
PubMed: 8406239
DOI: 10.1016/0278-6915(93)90045-z -
Toxicologic Pathology 1983Comparative studies of enzyme activities during the dedifferentiation of hepatic cells and through their development into overt hepatomas are few and contradictory. This... (Comparative Study)
Comparative Study
Comparative studies of enzyme activities during the dedifferentiation of hepatic cells and through their development into overt hepatomas are few and contradictory. This study was designed to investigate the histochemical, biochemical and morphologic features of the altered liver cells with particular emphasis on the importance and validity of the histoenzymatic behavior of glucose-6-phosphatase (G6Pase) as a marker for the detection of precancerous hepatic cells. Serum and hepatic levels of G6Pase were analyzed and compared with the histoenzymatic behavior of this enzyme. The use of other enzymes, such as adenosine triphosphatase (ATPase) and gamma glutamyl-transpeptidase (GGT) as histochemical markers for malignancy was also tested. The activities of a variety of enzymes commonly used as diagnostic tools were also evaluated in both the liver homogenates and sera of rats treated with 2 mg diethylnitrosamine (DENA)/kg body weight for 2-28 weeks. Using G6Pase as a histoenzymatic marker, precancerous cells appeared after 4 weeks of exposure to DENA in the form of small islets devoid of G6Pase activity. These G6Pase free cells increased in number forming larger islands and finally appeared as tumor nodules after 28 weeks of treatment. The histoenzymatic behavior of ATPase was identical to that of G6Pase. The precancerous cells, as well as the tumor cells appeared devoid of ATPase activity. The application of GGT as a marker, showed significantly increased activity in the altered liver and tumor cells. Increased serum levels of G6Pase were noted after 10 weeks and were greatly elevated in the late stages of the evolution of the precancerous cells.(ABSTRACT TRUNCATED AT 250 WORDS)
Topics: Animals; Diethylnitrosamine; Female; Glucose-6-Phosphatase; Liver; Liver Neoplasms, Experimental; Nitrosamines; Rats; Rats, Inbred Strains
PubMed: 6100219
DOI: 10.1177/019262338301100104