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Differential Oncogenic Effects of Methylnitrosourea2
Abstract
We investigated differences in the oncogenic effects of methylnitrosourea (MNU) which were induced by varying dose schedules and changing administration routes. The nervous system represented the target organ when MNU was given intravenously to rats. Carcinogen by other routes resulted in decreased numbers of neurogenic tumors and the appearance of neoplasms at the injection site. Increased oral doses of MNU caused shorter survival times, a decreased incidence of neuroglial tumors, and increased numbers of thymic lymphomas and mesenchymal tumors of the nervous system. The results suggest that many tissues are susceptible to the oncogenic effects of MNU, but the degree of exposure necessary for neoplastic transformation varies.
... Although the alkyl adducts at guanine-O 6 , thymine-O 2 and -O 4 , and cytosine-O 2 together represent only a minor fraction (<15%; Beranek 1990), these lesions exhibit by far the highest mutagenic efficiency of all ENU-or MNU-induced DNA alkylation products. This is due to their mispairing character during DNA replication and correlates well with the carcinogenic potential of the respective chemicals (Singer 1983). Under in vivo conditions, ENU undergoes non-enzymatic heterolytic decomposition with a half-life t ½ of ! 8 min (Goth and Rajewsky 1972). ...
... Similar experiments with MNU in BDIX rats showed a linear correlation between the total carcinogen dose administered in weekly fractions and tumor incidence as well as an inverse correlation to latency time (Druckrey et al. 1965). Additionally, the route of application of MNU appears to be important with the neurogenic tumor incidence being highest after intravenous administration (Swenberg et al. 1975). ...
The alkylating substances N-ethyl-N-nitrosourea (ENU) and N-methyl-N-nitrosourea (MNU) are the most potent systemically acting neurocarcinogens in rodents. They induce tumors of the central
(CNS) and peripheral nervous system (PNS), which are similar to their human counterparts. The neuro-oncogenic effect of both
chemicals depends on the dose, the rodent species, the strain, and the developmental stage at exposure. Neuro-oncogenesis
induced by MNU and ENU has been widely used as a model system including studies of molecular mechanisms of malignant transformation,
screening of therapeutics, and lately, the genetics of predisposition toward the development of these tumors. The induction
of malignant tumors in the PNS of differentially susceptible rat strains made it possible to identify gene loci influencing
tumor risk in an allele- and sex-specific way and to gain insight into molecular and cellular mechanisms underlying tumor
susceptibility and resistance.
... Among hematolymphatic tumors induced in rats (14,15), unlike those in mice (20), thymic lymphomas are rare, but erythroleukemias and myelocytic leukemias are prevalent in most strains. However, it is reported that thymic lymphomas are induced by methylnitrosourea at a high incidence in Sprague-Dawley rats (9,21) and by PNU3 in Fischer 344 rats (22). In an attempt to analyze PNUinduced thymic lymphomagenesis, we studied the strain differ ence of susceptibility to PNU-induced leukemias among 6 strains of rats and subsequently analyzed the genetic segregation among crosses between highly susceptible Fischer and poorly susceptible Long-Evans rats. ...
... These alternatives cannot be formally excluded until the mechanism of action of these genes is elucidated. It is reported that Sprague-Dawley rats are highly susceptible to thymic lym phoma induction by methylnitrosourea (9,21 ), whereas they are less susceptible to PNU in this study. Currently, it is not known whether this discrepancy is due to distinctive genetic control for leukemogenesis by structurally different nitrosoureas or merely substrain differences among Sprague-Dawley rats. ...
Administration of propylnitrosourea p.o. by our protocol induced a high incidence of hematolymphatic neoplasms in all six rat strains studied. Remarkable strain differences in susceptibility to thymic lymphomas were observed. The incidence of thymic lymphomas was high in Fischer 344 (98%) and Wistar/Furth (71%) but low in Sprague-Dawley (29%), ACI/Ms (23%), Donryu (24%), and Long-Evans (10%) strains. Segregation of thymic lymphoma incidence among crosses between highly susceptible Fischer and poorly susceptible Long-Evans rats indicated that the increased susceptibility to thymic lymphomas of Fischer rats was determined by a dominant gene TIs-1 (thymic lymphoma susceptible) and that this gene was linked to the coat color loci, p and c, in Linkage Group I in the order of TIs-1 - c - p. The presence of another independently assorting dominant gene, TIs-2, was also suggested to accelerate the thymic lymphoma-genesis. Expression of the group-specific antigen of murine leukemia virus as well as infectious viruses was not detected in nine propylnitrosourea-induced thymic lymphomas of Fischer rats.
... 23) Therefore, the susceptibility of the rodent thymus to develop earlier and more aggressive MNU-dependent tumors seems to depend particularly on its limited capability for DNA repair through the enzyme O 6 -alkylguanine-DNA alkyltransferase. 23,26) Since MNU has a broad spectrum of target organs, the possibility exists that if the experiment had been extended in time, tumors would have developed in other tissues. 10,26) The present study, however, focused only on LHS tumors because they could be used as early markers of the carcinogenic influence of MNU on the Wistar strain. ...
... 23,26) Since MNU has a broad spectrum of target organs, the possibility exists that if the experiment had been extended in time, tumors would have developed in other tissues. 10,26) The present study, however, focused only on LHS tumors because they could be used as early markers of the carcinogenic influence of MNU on the Wistar strain. The development of putatively preneoplastic liver GST-P + foci of hepatocytes was also evaluated, because this has been an important parameter in the rat alternative medium-term bioassays for carcinogenesis. ...
The Brazilian Agency for the Environment (IBAMA) recently adopted an alternative medium-term multiple-organ assay system with the Wistar rat strain for detection of the carcinogenic potential of pesticides. Originally, this initiation-promotion protocol was established in Japan with the isogenic Fischer 344 male rat. Among the initiating agents used in that assay, N-methyl-N-nitrosourea (MNU) rapidly induces malignant lymphoma and leukemia and early mortality of rats from different strains. This study was developed to evaluate whether the outbred Wistar rats are also similarly susceptible to MNU. Particularly, it aimed to evaluate the dose-response relationship and to register the MNU-induced pre-neoplasia and neoplasia that may develop in the lympho-hematopoietic system (LHS) of the Wistar rat within a medium-term period. Four groups of male Wistar rats were treated during 2 weeks with vehicle or with MNU (80, 160 or 240 mg/kg body weight, i.p.). After sacrifice at the 12th and 20th weeks, the thymus, spleen, bone marrow, cervical and mesenteric lymph nodes and liver were collected for analysis. At the 20th week, LHS malignant tumors and benign vascular tumors occurred only in the high- and intermediate-dose MNU-treated animals. Four animals treated with 240 mg/kg developed diffuse thymic lymphomas; two others, treated respectively with 240 mg/kg and 160 mg/kg, developed spleen hemangiomas. The present observations indicate that the Wistar strain is as susceptible as other strains to the early development of MNU-induced LHS (pre)neoplasia. Therefore, this strain seems suitable to be used as test system in bioassay protocols that adopt MNU as an initiating agent for carcinogenesis.
... The type of cancer developed is dependent on the age, sex and strain of the laboratory animals, route of administration, concentration and the total dose of MNU, as well as the duration of the experimental period. Rats induced with MNU at young age (4 to 6-weekold) were reported to develop higher incidence of thymic lymphoma (Swenberg et al., 1975;Koestner et al., 1977;Mizoguchi et al., 1993) compared to middle-aged (52-week-old) and old age (98-week-old) rats (Mizoguchi et al., 1993). In particular, the middle-aged rats were found to likely to develop higher (97 -100%) incidence of adenocarcinomas in the small intestine compared to young and old age rats (Mizoguchi et al., 1993). ...
This study described the histopathological features of peripheral T-cell lymphoma in male Sprague Dawley rats
following intraperitoneal (i.p.) injections of N-methyl-N-nitrosourea (MNU) at a dose of 60 mg/kg body weight
per injection, administered twice weekly for 2 consecutive weeks, and followed by a five-month’s observation
period. Control rats were injected with normal saline, i.p. All the rats treated with MNU had enlargement
of lymph nodes, with 30% had hepatosplenomegaly and 7% had enlarged kidneys at necropsy. Malignant
lymphoma was observed in the lymph nodes, spleen, liver, lung, heart, and kidneys. The neoplastic cells were
characterised as undifferentiated, and small to large size with bizarre pleomorphic nuclei. The severity was
further described as mild, moderate and severe, based on the diffuseness of the lesions. Nonetheless, similar
lesions were not observed in the thymus of the rats. Immunohistochemistry staining of the organs was positive
for CD3 antibody, which is consistent with T-cell lymphoma.
... The type of cancer developed is dependent on the age, sex and strain of the laboratory animals, route of administration, concentration and the total dose of MNU, as well as the duration of the experimental period. Rats induced with MNU at young age (4 to 6-weekold) were reported to develop higher incidence of thymic lymphoma (Swenberg et al., 1975;Koestner et al., 1977;Mizoguchi et al., 1993) compared to middle-aged (52-week-old) and old age (98-week-old) rats (Mizoguchi et al., 1993). In particular, the middle-aged rats were found to likely to develop higher (97 -100%) incidence of adenocarcinomas in the small intestine compared to young and old age rats (Mizoguchi et al., 1993). ...
This study described the histopathological features of peripheral T-cell lymphoma in male Sprague Dawley rats following intraperitoneal (i.p.) injections of JV-methyl-./V-nitrosourea (MNU) at a dose of 60 mg/kg body weight per injection, administered twice weekly for 2 consecutive weeks, and followed by a five-month's observation period. Control rats were injected with normal saline, i.p. All the rats treated with MNU had enlargement of lymph nodes, with 30% had hepatosplenomegaly and 7% had enlarged kidneys at necropsy. Malignant lymphoma was observed in the lymph nodes, spleen, liver, lung, heart, and kidneys. The neoplastic cells were characterised as undifferentiated, and small to large size with bizarre pleomorphic nuclei. The severity was further described as mild, moderate and severe, based on the diffuseness of the lesions. Nonetheless, similar lesions were not observed in the thymus of the rats. Immunohistochemistry staining of the organs was positive for CD3 antibody, which is consistent with T-cell lymphoma.
... The route of application of MNU has a reproducible influence upon the incidence of neurogenic tumors. The incidence is highest with intravenous administration (95%), followed by oral (52%), intnperitoneal(36%) and subcutaneous (1 2%) administrations of MNU (16,29). ...
Two decades of research with resorptive neurocarcinogens firmly established the high potency of methyl and ethylnitrosourea (MNU and ENU) as neurocarcinogens, particularly in rats. There are significant differences in susceptibility to these agents among species. There are also differences among age groups. Fetuses are between 50 to 100 times more susceptible than adult rats. One single iv inoculation of 20-50 mg/kg ENU into pregnant rats may produce neurogenic tumors in 100% of the offspring. The tumors produced by these compounds have been well characterized morphologically, biologically, biochemically and histochemically. Tumors produced by both compounds are mostly gliomas and neurinomas (Schwannomas), however, clear differences exist between ENU and MNU produced neoplasms. Transplacental exposure to ENU generally results in a high number of anaplastic neurinomas and mostly differentiated gliomas (astrocytomas, oligodendrogliomas or mixed gliomas). In contrast, multiple exposures of adult rats to MNU result in a moderate number of mostly differentiated neurinomas and a high number of anaplastic gliomas. Tumors usually start out as well differentiated oligodendrogliomas or astrocytomas. As they grow larger, they become more mixed and anaplastic. In contrast to spontaneous gliomas in old rats, MNU and ENU-induced astrocytomas can be readily identified with well established biomarkers such as the S100 protein and particularly GFAP (glial fibrillary acidic protein). Neurinomas are also strongly positive for S100 protein. No reliable markers exist for oligodendrogliomas. Neurogenic tumors induced by MNU or ENU, as well as derived cell lines and clones from such tumors, have been successfully used as models for neurocarcinogenesis and therapeutic screening.(ABSTRACT TRUNCATED AT 250 WORDS)
We have previously reported that streptozotocin (STZ) produces islet-cell tumors in rats. In this study small doses of STZ were administered to young male Wistar rats to clarify their diabetogenic and oncogenic action on endocrine pancreas.
The plasma glucagon (IRG) levels 4, 24 and 48 hr after administration of 30 mg/kg body weight (BW) of STZ were significantly elevated compared to controls, while typical triphasic curves of blood glucose and plasma insulin (IRI) were not observed.
Oral glucose tolerance tests were performed serially in rats treated with 30 mg/kg BW of STZ One week after the treatment, hyperglycemia and suppression of IRI responses to glucose load were not observed but IRG levels were elevated paradoxically following glucose load. At the ninth month, the IRI responses to glucose rose markedly in some of the rats.
One year after treatment, pancreatic islet-cell tumors were demonstrated in 8 of 9 and 5 of 7 rats which had been treated with 30 and 40 mg/kg BW of STZ, respectively. The IRI responses to glucose were increased significantly in 10 of the 13 tumor-bearing rats and the others had low insulin responses. The former showed low and the latter high blood glucose levels during oral glucose tolerance tests and no significant difference in IRG levels was observed between the two groups.
The tumors contained large amounts of insulin. Furthermore, radioimmunoassayable glucagon was demonstrated in the tumor extracts. Some of the dilution curves for extracted insulin and glucagon, however, were slightly different from those of rat insulin and porcine glucagon standard, respectively.
The above data indicate that:(1) small doses of STZ exert an effect on circulating glucagon levels in rats and also a potent oncogenic action on endocrine pancreas, (2) STZ-induced tumors are insulin-producing islet-cell tumors, and (3) the tumors produce insulin and glucagon which are, in part, immunochemically heterogeneous.
The study of primary tumors of the nervous system has been greatly complicated by the prevailing nonhomogeneity of these growths and the frequent existence of multiple cell types within a single tumor. Rigorous histological controls have been essential, and special biochemical techniques have been utilized to cope with inherent difficulties of the problem. The mixture of multiple cell types within a tumor sample is a common difficulty. While fairly uniform composition may be found in some of the benign intracranial human tumors (meningiomas and Schwannomas) or well differentiated gliomas (oligodendrogliomas), many of the most common and biologically malignant neoplasms are not uniform. Normal cellular constituents may be intermingled with neoplastic cells. A normal biological characteristic of glial cells is the ability to insinuate their cellular extensions among neural fibers and processes, and the comparable gliomatous cells continue to do this in the neoplastic condition (Fig. 1). The advancing front of a glioma is poorly demarcated and often impossible to identify grossly; yet, the infiltrated and incorporated neurons, myelinated fibers, or reactive astrocytes may differ greatly in biochemical composition from the tumor cells. Even tumors of different primary embryonic cell layers may coexist within the same tumor mass.
Tumors of the nervous system can be induced in experimental animals by chemical compounds from different classes. With regard to the subject of this survey, methods of inducing brain tumors in experimental animals by intracranial (topical) administration of carcinogenic agents will not be discussed in detail. As shown in Table 1, different measures, e.g., administration of aromatic hydrocarbons, of RNA-and DNA-tumor viruses, or of radioactive elements like 60Co or 241Am, have been successfully used to induce brain tumors upon topical administration (Zimmerman 1969, 1982; Bigner and Swenberg 1977).
This survey of spontaneous and induced cardiac tumors of rats, mice, guinea pigs and hamsters is based on a critical review of the literature and the hitherto unpublished data of a study of the material accessed of the NCI Registry of Experimental Cancers. The frequency of cardiac tumors in laboratory rodents, the location within the heart, the histologic classification and heredity aspect are compared with these features in humans with cardiac tumors.
Ten patients in whom tissue proliferation akin to angioglioma occurred within the brain are described; seven of the lesions were supratentorial and three infratentorial. Only 31 accepted instances of such neoplasms have been found in the literature. The combined lesions usually become symptomatic in the second and third decades. In all 10 cases, the angiomatous part of the combined tumors showed characteristic vascular malformation such as severe hyalinization, tortuosity, and some were even calcified. The number of abnormal blood vessels were excessive in all examples. The glial portion consisted of either astrocytoma, oligodendroglioma, or mixtures of these gliomas. Dedifferentiation of the neuroglia combined with neoplastic endothelial proliferation indicates the true neoplastic nature rather than reactive gliosis associated with a vascular anomaly. © 1996 Wiley-Liss, Inc.
An experimental model for induction of gliomas corresponding to human anaplastic astrocytomas and glioblastomas is reported. Eleven week old F344 and ACI rats were given 100 or 200 p.p.m. N-methyl-N-nitrosourea (MNU) solution as their drinking water for 42 weeks. Gliomas were induced at very high incidences (82.5-92.5%) in each group. Induced gliomas showed apparent evidence of morphologic malignancy by an analysis based on diagnostic criteria of human astrocytomas. All of the gliomas from the killed animals were classified histologically into subtypes according to the classification scheme used in the diagnosis of human gliomas. The majority of macrotumors more than 1 mm in diameter in both strains were diagnosed as anaplastic astrocytomas and glioblastomas. lmmunohistochemically, tumor cells in these tumors were almost negative for glial fibrillary acidic protein, while ultrastructurally neoplastic astrocytes contained glial filaments. A strain difference was observed in the ratio of histological subtypes of macrotumors. In F344 rats, astrocytic tumors diagnosed as anaplastic astrocytomas and glioblastomas of an astrocytic type formed the majority, whereas glioblastomas of mixed oligo-astrocytic type predominated in ACI rats. The results indicate that MNU-administration to adult F344 rats may provide a suitable experimental model for gliomas which occur in adult humans.
Brain cancer accounts for approx 1.4% of all cancers and 2.3% of all cancer-related deaths. The incidence of primary cerebral
malignancies varies between 4 and 10/100,000 in the general population. This incidence tends to increase with age (4/100,000
up to the age of 12 yr; 6/100,000 up to the age of 35 yr; 18/100,000 up to the age of 55 yr; 70/100,000 up to the age of 75
yr).
Carcinogenic agents such as N-methyl-N-nitrosourea can cause tumors. The aims of the present study were to evaluate and classify a subtype of AML (acute myeloid leukemia) that was induced by NMU. According to previous publications, NMU induces not only mammary cancer but also leukemia in Sprague-Dawley (S-D) rats. However, the subtype of leukemia involved in NMU-treated rats is unknown. We found that both organ weight and relative organ weights were significantly higher in NMU-exposed rats than in controls. Morphological changes of rat livers and spleens were assessed by histological evaluation (H&E staining), which found that these tissues were abnormal in appearance. Also, cytological examination of the blood showed immature white blood cells in a smear using Liu's and Papanicolaou stains, indicating that gross abnormalities and histopathological changes were pathologically observed. NMU leukemia incidence was 97.1%. In this study, immunohistochemical (IHC) analysis was valuable in classifying the leukemia of poorly differentiated blasts induced by NMU. Paraffin blocks were stained for MPO, CD3, CD15, CD20, and CD34 markers. The NMU-induced group was positive for MPO, but negative for CD3, CD15, CD20, and CD34. These CD markers suggest that they are useful in helping diagnose APL (M3) leukemia. The model of NMU-induced leukemogenesis in an S-D rat suggests a more definite way to classify APL. This APL will provide an important tool for chemical carcinogenesis and leukemia studies.
Intragastric administration of N-nitroso-N-methylurea to strain 13 male guinea pigs, at a weekly dosage of 7.5 mg/kg for 15 weeks and then twice weekly for a subsequent 15 weeks, induced high toxicity, as evidenced by weight loss and mortality and a high incidence of malignant neoplasms, over a total observational period of 40 weeks. The neoplasms included hepatic angiosarcomas, cholangiocarcinomas, and generalized lymphoblastic lymphomas. © 1977, American Association for Cancer Research. All rights reserved.
Methylation of guanine at its O6 position occurred in both the unique and the repetitive DNA sequences in several rat organs following an in vivo pretreatment with the carcinogen N-methyl-N-nitrosourea. Little or no difference was found in the distribution of O6-methylguanine between the 2 DNA fractions.
The carcinogen 3-methyl-1-phenyltriazene (MPT) was administered subcutaneously to normal or pregnant BD VI rats and DNA and RNA were isolated from various tissues after 8 h or 15 h, respectively. Sephadex G-10 chromatography of DNA hydrolysates showed the presence of 7-methylguanine in all tissues examined including that of the brain, one of the target organs for tumour induction. The amounts of the minor product, O6-methylguanine, were characteristic of an SN1 reaction mechanism. Dowex-50 chromatography of RNA hydrolysates showed the presence of 7-methylguanine and of the minor product, 3-methylcytosine. The relative amounts, both of the methylated bases in the individual nucleic acids and of 7-methylguanine in DNA and RNA, were similar to those found previously after administration of 3,3-dimethyl-1-phenyltriazene (DMPT). This suggests the involvment of a common alkylating intermediate. De novo incorporation of radioactivity into purine bases was detected in both DNA and RNA although the levels were not related to the amounts of methylation. The results show that MPT is sufficiently stable to alkylate nucleic acids in vivo and are consistent with the hypothesis that this reaction is a prerequisite for tumour induction. Futhermore, they support the proposal that MPT is the active intermediate in the induction of tumours by DMPT.
Dose-response studies in chemical carcinogenesis are a valuable method for evaluating the risk involved by a certain carcinogen. This article deals with the methods that are presently applied for dose-response studies. It is emphasized that there are many possible mistakes that are already hidden in the method itself; a critical approach is made to the indicative value of dose-response studies. It is also emphasized that the conclusions which may be drawn from dose-response studies can always only be extrapolative but never mathematically calculable. Finally, some practical examples for dose-response studies with n-nitroso compounds are described.
The carcinogenicity of N-methyl-N-nitrosourea (MNU) has been demonstrated in several species for a variety of tissues, particularly the nervous system. Boxer dogs, a breed with a high incidence of spontaneous tumors (including brain tumors), were exposed to MNU to determine their susceptibility to MNU and to compare the induced tumors with the types of tumors reported in the literature. Twenty purebred Boxer dogs were given weekly i.v. injections of MNU, 5 mg/kg, for 36 weeks and were observed for approximately 3 years for the development of neoplasia. Nineteen of 20 dogs developed neoplasms 19 to 36.5 months after the first injection. The mean latent period was 30 ± 5 (S.D.) months. All tumor-bearing dogs had 2 or more neoplasms, including metastases, and 14 dogs had tumors of more than 1 tumor type. Malignant neurinomas of the small intestine were present in 11 dogs, and poorly differentiated sarcomas of the small intestine were found in 6 dogs. Metastasis occurred to liver, spleen, lymph nodes, lung, and adrenal. Two cases of malignant neurinoma developed in the heart, and 1 each developed in the stomach and colon. Two well-differentiated neurinomas occurred in the small intestine, and 1 each occurred in a spinal root, the stomach, and the colon. The high incidence rate, young age of development, and unique tumor types clearly distingiushed the tumors from spontaneous tumors of Boxers. Although the lack of brain involvement was unexplained, MNU had a neurooncogenic effect on the peripheral nerves of the gastrointestinal tract.
Intragastric intubation of MNU (20 mg/kg twice weekly for 9 weeks) into young Srague-Dawley rats resulted in a 100% incidence of thymic lymphomas and gastric carcinomas in animals surviving more than 14 weeks. Progressive thymic atrophy followed the successive MNU administration culminating in marked lymphoid depletion by the 5th week of MNU treatment. At 6 weeks, repopulation of the thymus with lymphoblastic cells was observed, which progressed to thymic neoplasia. The first thymic lymphoma appeared at the 6th week. By the 15th week all rats were either moribund or had succumbed to anoxia caused by pleural effusions associated with thymic neoplasia. Twenty-six of 31 thymic tumors were classified as lymphocytic lymphomas, 2 as histiocytic, 1 as mixed lymphocytic-histiocytic, 1 as bimorphic and 1 as an epithelial thymoma. All but 2 lymphomas were restricted to the thymus. Of these 2 (both histiocytic lymphomas), 1 involved the spleen, liver and abdominal lymph nodes and the other infiltrated locally into cervical and thoracic regions. Significant thymic enlargement occurred only after MNU treatment was stopped (MNU acting as an anti-cancer drug). The gastric carcinomas were restricted to the non-glandular portion of the stomach and developed as a final step following ulceration, hyperplasia and neoplastic transformation. The short latency period and the high incidence of thymic neoplasms represent valuable criteria for future use of this model for pathomorphogenetic as well as immuno- and chemotherapeutic investigations.
Bladder cancer can be induced in the rat by the intravesicular administration of N-methyl-N-nitrosourea. DNA damage in rat bladder epithelial cells after administration of methylnitrosourea has been examined by measuring the change in sedimentation of the DNA in alkaline sucrose gradients. A dose response of DNA damage in the urothelium was observed with single intravesicular doses of 0.1, 0.3, and 0.5 mg of methylnitrosourea. Larger doses of methylnitrosourea damaged the epithelium so extensively, that biochemical studies were not feasible. DNA repair, measured by the return to a normal sedimentation pattern of DNA on alkaline sucrose gradients, was followed over a period of 9 days with the use of 0.5 mg of methylnitrosourea to initiate the damage. Bladder epithelial cells were able to repair the DNA damage induced by methylnitrosourea. However, the possibility of persistent damage not detectable by sedimentation of DNA on alkaline sucrose gradients cannot be ruled out.
The present study reports the induction, histopathology, immunocytochemistry, growth pattern and proliferative behaviour of mesenchymal tumours of the urinary bladder of rats induced by a single intravesical dose (5 mg/kg/body weight) of N-methyl-N-nitrosourea (MNU). In 14 of 283 female Wistar rats (incidence: 4.9%). 16 non-epithelial tumours had developed after an experimental period of 14 months. The most common histological type induced was of fibrohistiocytic origin (eight benign-appearing and three malignant fibrous histiocytomas). Furthermore, two pure histiocytomas (one benign and one malignant) and three capillary and cavernous haemangiomas were produced. Since no metastases occurred and no clear-cut distinction between a merely expansive and a truly invasive growth was possible, the diagnosis of malignancy was based on the degree of cellular atypia and the mitotic activity. The benign-appearing fibrous histiocytomas showed a mean mitotic index of 0.06% and the malignant fibrous histiocytomas of 0.34%. The mitotic activity of the malignant histiocytoma was threefold (0.78%) as high as in the benign-appearing histiocytoma (0.25%). There exist close morphological similarities between the induced mesenchymal tumours in the rat and those occurring in the human bladder. Although the spectrum of histological types of mesenchymal tumours seen in the rat bladder was narrower than that encountered in man, MNU seems suitable for further studying the histogenesis, histopathology and biology of experimentally induced non-epithelial bladder neoplasms to gain information for a better understanding of the pathogenesis of human disease.
From the standpoint of hazard identification of neurooncogenic substances, the specificity of chemical structure for neurooncogenic activity, mode of administration, and influence of host factors such as the strain, sex, and age on the incidence, location, or type of spontaneous or chemically induced rat neurogenic tumors are reviewed and discussed. In addition, diagnostic problems on some of the neurogenic tumors in the practical differential diagnoses are briefly pointed out
The cytotoxic activity of N-methyl-N-nitrosourea (MNU), streptozotocin, and N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) was determined in cell culture by using a P388 cell growth rate inhibition assay. These agents appear to have very different activities when inhibition is related to the agent concentration in the culture medium: ED50(C0) = 40 microM for MNNG to 875 microM for streptozotocin. The mechanism of action of these three agents involves conversion to the active methanediazonium ion and subsequent methylation of cellular macromolecules. As a consequence, the rates of conversion of the parent agent to the methylating species in the medium and within the cell are important parameters that also need to be considered to reach a more detailed understanding of the mechanism of action. In order to do this, a kinetic model has been developed to calculate the concentration of drug that is converted to active methylating species within the cell during the assay incubation period. The use of cell culture kinetic models was extended from simple compounds activated through solvolytic reactions (nitrosoureas) to an agent that undergoes selective intracellular activation (MNNG). By use of measured values for initial drug concentration, incubation time, and cell volume, as well as extracellular and intracellular chemical activation rate constants, the intracellular concentration, [P4], which represents the cumulative intracellular reaction products formed during the incubation period, was calculated and related to cytotoxicity. All three agents showed an ED50[P4] between 140 and 180 microM, and for MNNG, this ED50 was independent of extracellular sulfhydryl concentration.(ABSTRACT TRUNCATED AT 250 WORDS)
Properties of estrogen and progestin receptors present in cytosol and of prolactin receptors in membrane fractions of nitromethylurea (NMU)-induced mammary tumors in the rat were studied. Sucrose gradient analysis and specific studies revealed a similarity to properties of mammary tumors induced by 7,12-dimethylbenz(a)anthracene (DMNA). Dissociation constants (K(D)) at 0.32 and 2.33 nM were observed for receptors for estradiol and progesterone, respectively. In addition, prolactin receptors had specificities and binding constants similar to those found in DMBA-induced mammary tumors. The hormonal dependence of these tumors has also been studied in ovariectomized rats and intact rats given injections of antiestrogens. In the first experiment, rats were given injections beginning on the day of the first NMU injection with 11α-methoxyethinylestradiol or tamoxifen (1-p-β-dimethylaminoethoxyphenyl-trans-1,2-diphenylbut-1-ene) or were ovariectomized. All treatments resulted in a diminution the initiation of tumor growth compared to the control group as had been observed for DMBA-induced mammary tumors. In a second experiment, the effect of these same treatments was studied in animals with established tumors. In control rats, there was an increase in tumor number and size, while ovariectomy and tamoxifen or 11α-methoxyethinylestradiol treatments resulted in a prevention of further tumor growth, although none of these treatments induced tumor regression as occurs with DMBA-induced mammary tumors. Ovariectomy and tamoxifen significantly reduced specific binding of 17β-[3H]estradiol and [3H]-17,21-dimethyl-19-norpregna-4,9-diene-3,20-dione to cytosol of NMU-induced mammary tumors. Hormone receptor levels for estradiol, progesterone, and prolactin in NMU-induced tumors were of a magnitude similar to those observed in DMBA-induced tumors. Antiestrogen treatment did not affect specific binding of 125I-labeled ovine prolactin, but ovariectomy did result in a reduction in prolactin binding. Plasma prolactin values were unchanged by ovariectomy or tamoxifen and increased by 11 α-methoxyethinylestradiol. These results suggest that NMU-induced mammary tumors are more responsive to endocrine treatment coincident with NMU injection but that treatment of established tumors is somewhat less effective.
The high susceptibility of certain organs, for example rat brain, to induction of cancer by N-nitroso-N-alkyl-ureas, has been related to a low ability to remove O6-alkylguanine (O6AG) from DNA. It is therefore reasonable to ask why mouse brain, in which there is also a slow disappearance of O6AG from DNA after treatment with nitroso-alkyl-ureas, is not susceptible and why, in mice, thymus and lung are the main target organs. The explanation of the species difference could lie in the fact that replication of alkylated DNA is an essential event in initiation. If nitroso-alkyl-ureas had a greater inhibitory effect in some organs than in others, replication might be inhibited until after the O6AG had been removed, so preventing replication of DNA while still alkylated. This concept was tested by comparing the effect of N-nitroso-N-methyl-urea (NMU) on incorporation of [3H]TdR into DNA of relevant organs in Wistar rats and C57BL mice, and by determining ability to remove O6AG from DNA by measuring the alkyl acceptor protein (AAP) concentrations in these organs. No evidence was obtained that the AAP content was lower or inhibition of replication was less extensive in the organ of the species more susceptible to carcinogenesis than in the same organ of the less susceptible species.
Male and female F344/DuCrj rats were administered N-butyl-N-nitrosourea at a concentration of 400 ppm in their drinking water. By the 50th week of the experiment, the cumulative incidence of upper-digestive-tract tumors was as high as 35/39 (90%) and 34/39 (87%) in male and female rats, respectively. Among these, esophageal and forestomach tumors occurred most frequently. Except one female rat with fibroma, upper-digestive-tract neoplasms were of the epithelial type -- papilloma, squamous-cell carcinoma or adenocarcinoma. In female rats, vaginal tumors were induced in 16 (41%) animals. Ear-duct tumors and hematopoietic neoplasms were also induced in both sexes of rats, with incidence of less than 21%.
The susceptibility of rat brain to induction of cancer by N-nitroso-N-ethyl-urea (NEU) increases dramatically from a very low level in the 12 day foetus to a maximum at the time of birth, and then decreases with age. Liver tumors are rarely induced by a single treatment with NEU at any age. If induction of cancer by nitroso-alkyl-ureas depends on replication of DNA containing the mispairing base O6-alkylguanine, susceptibility would reflect the balance between the protective effect of removal of the base by repair mechanisms and the potentiating effect of cell replication. The capacity of tissues to remove O6-alkyl-guanine from DNA depends on the amount of alkyl acceptor protein (AAP) present. To study the concept that carcinogenesis results from replication of alkylated DNA, the AAP contents and relative rates of DNA replication were studied in brain and liver of rats at various stages of development, from the 12 day foetus to the 48 week old rat. Replication in liver was approximately ten times higher than in intra-cranial contents at each stage of development studies. The AAP content was higher in the 12 day foetal brain than later, decreasing to low levels as susceptibility to NEU increased until the time of birth, and then remaining low in the adult. The peak sensitivity of brain therefore corresponds to the time at which AAP content is low and rate of DNA replication is high. With liver, AAP levels are low in the foetus, although higher than in brain, and increase after birth. The higher level of AAP in the foetal liver compared with that of brain is possibly sufficient to explain why cancer is not induced in liver in spite of the high rate of DNA replication. The results are consistent with the concept that replication of alkylated DNA is an essential event in initiation. There may be no quantitative relationship between replication and repair and susceptibility to cancer on comparison of different tissues, owing to the fact that, at the cellular level, cancer is a rare event. The amount of mispairing at replication necessary to bring it about may depend on the detailed organisation of the genome, and hence on cell type.
The metabolism-independent or “direct-acting” alkylating agents comprise a structurally diverse group of chemically reactive organic compounds that include some of the most potent known mutagens and carcinogens. Their conversion to reactive intermediates requires no enzyme-mediated catalysis, and in this respect these compounds differ from the great majority of organic chemical carcinogens. Many of the direct-acting alkylating agents have been used in chemical syntheses for decades, and their chemistry is accordingly well known. The carcinogenicity of a number of these compounds has been studied extensively in both rodent and non-rodent species, and the results of such studies provide a remarkable illustration of qualitative interspecies differences in susceptibility to carcinogenesis that cannot be ascribed to differences in metabolism of xenobiotic compounds.
It is well known that oral administration of N-methylnitrosourea (NMU) induces carcinomas in the forestomach of experimental animals. As the rat forestomach is lined with an orthokeratinized squamous epithelium the purpose of the present study was to elucidate whether carcinogenesis in this localization following intragastric application of NMU was comparable to oral carcinogenesis in experimental animals and humans. Furthermore, the development of oral mucosal lesions in the rats was studied. It is concluded that the morphologic changes seen during carcinogenesis in the forestomach do not seem to differ essentially from those seen during experimental and human oral carcinogenesis. Furthermore, the number of oral lesions found in the present study are higher than reported in other strains of rats following administration of NMU.
Acrylonitrile (ACN) produces tumors in rats, particularly gliomas of the brain, but tests for genotoxicity have yielded mixed results and no ACN-DNA adducts have been identified in the brain. To examine the possibility that ACN-related brain tumors were not a consequence of binding of ACN to brain DNA, experiments were conducted to investigate possible epigenetic mechanisms. Male Sprague-Dawley rats were exposed to 0, 3, 30, and 300 ppm ACN in drinking water for 21 days, a range that includes doses associated with brain tumorigenesis. In the 30 and 300 ppm ACN groups, 8-oxodeoxyguanosine (8-oxodG) levels were two fold greater than in the controls. Measures of glutathione levels, glutathione peroxidase and catalase were not significantly changed, but cyst(e)ine was somewhat increased. No changes were found in brain cytochrome oxidase activity, which indicates a lack of metabolic hypoxia. Also, no effects on thiobarbituric acid reactive substances were found, indicating a lack of lipid peroxidation. In an additional experiment, male Sprague-Dawley rats were exposed to 0 or 100 ppm ACN in drinking water for 94 days; interim sacrifices were conducted at 3, 10, and 31 days. Levels of brain nuclear DNA 8-oxodG were significantly increased in ACN-exposed rats compared with controls. Another group of animals were given weekly i.v. injections of 5 mg/kg methylnitrosurea and no increases in 8-oxodG were found. These studies suggest the possibility that ACN-induced tumors may be produced by a mode of action involving 8-oxodG. The formation of 8-oxodG is not understood, but does not appear to involve lipid peroxidation or disruption of antioxidant defenses.
Brain tumors seemed to have increased in incidence over the past 30 years, but the rise probably results from use of new neuroimaging techniques. Treatments have not improved prognosis for rapidly fatal brain tumors. Established brain tumor risk factors (exposure to ionizing radiation, rare mutations of penetrant genes, and familial history) explain only a small proportion of brain tumors, and only one of these potentially is modifiable. Genetic and environmental characteristics likely play a role in familial aggregation of glioma and these factors are not identified. New concepts in brain tumor etiology and clinical management are the goal of research, with an aim at eradicating this devastating disease.
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