Carcinogenicity of hydroxyalkylnitrosamines in F344 rats: contrasting behavior of beta- and gamma-hydroxylated nitrosamines.

The carcinogenicity of N-nitrosomethyl-(2-hydroxyethyl)amine (NMHEA), N-nitrosomethyl-(3-hydroxypropyl)amine (NMHPA), and the p-toluenesulfonate (tosylate) ester of NMHEA (NMHEATs) was tested in male and female F344 rats. The chemicals (25.6 mumol per application) were administered by twice-weekly gavage in corn oil (0.2 ml) for the lifetime of the animals. NMHEA was found to be an effective carcinogen under those conditions. The median survival time for the females was 9 mo after treatment was initiated, while for the males it was 12 mo. The principal cause of death of the females was hepatocellular carcinoma (14 of 20), while only 6 of 20 male rats exhibited that tumor. A few of the male rats had squamous cell carcinomas of the nasal epithelium (4 of 20), tumors which were not observed in the females. NMHPA was a much weaker carcinogen. Many of these rats survived for 2 yr and most had many age-related cancers. Nevertheless, 10 of the NMHPA-treated males and 2 females had adenocarcinoma of the lung, which was absent in the controls and also induced a significant number of neoplastic nodules in the livers of rats of both sexes. NMHEATs was also a weak carcinogen. However, besides many age-related tumors, it induced some hepatocellular carcinomas as well as hemangiosarcomas of the liver. NMHEATs was at least partially hydrolyzed to NMHEA, which was detected in the blood plasma of treated rats. A hypothesis has been advanced that NMHEA is activated to a proximate carcinogen by sulfate conjugation of the hydroxyl group; the present data do not contradict this hypothesis. The relatively lower carcinogenic potency of NMHPA, the different tumor spectrum induced by this chemical, and particularly the differences in chemical behavior suggest that its mode of activation is not the same as that for NMHEA.

Naturally occurring nonneoplastic histopathological lesions in the female SENCAR mouse.

Lesions that were considered naturally occurring were surveyed in female SENCAR mice used in short-term bioassays. These lesions were encountered in selected target organs and in organs and tissues with gross lesions encountered at necropsy. The genitourinary system was the most frequent site for lesions; cystic ovaries, cystic endometrial hyperplasia, and glomerulonephritis were commonly encountered in this system.

A combined carcinogen bioassay utilizing both the lung adenoma and skin papilloma protocols.

To test the feasibility of employing a combined lung adenoma/skin papilloma assay for broader detection of chemical carcinogenesis than that realized with either bioassay done separately, four strains of mice, SENCAR, BALB/c, A/J, and ICR-Swiss, were administered carcinogens either by the oral or intraperitoneal (IP) routes. The carcinogens administered were ethyl carbamate (EC), benzo(a)pyrene [B(a)P], N-[4-(5-nitro-2-furyl)thiazolyl]formamide (FANFT), and acrylamide (ACR). Starting 2 weeks later, 1 to 5 micrograms (depending on strain) of 12-O-tetradecanoylphorbol-13-acetate (TPA) in 0.2 mL acetone/mouse was applied three times weekly to the shaved back for 20 weeks. All strains displayed increases in the yield of lung adenomas in response to EC at 32 weeks. B(a)P increased lung adenomas in only the SENCAR and A/J strain. Only the SENCAR and ICR-Swiss mice gave positive responses in the skin. In the SENCAR mice, positive response was seen with all four chemicals, however, FANFT gave an inconsistent response. The ICR-Swiss mice responded with an increased skin papilloma yield only to EC. In a separate experiment involving only SENCAR mice, animals were treated with a single oral dose of diethylnitrosamine (DEN) followed by triweekly application of 1.0 microgram TPA. This treatment resulted in 51/57 animals developing lung adenomas vs. 5/57 in the control animals. No treatment-related skin tumors resulted with DEN. Histopathologically confirmed lesions indicate that the spectrum of chemicals detected in the SENCAR mouse may be broadened using a combined bioassay that examines both lung and skin responses.

Gross and microscopic lesions in the female SENCAR mouse skin and lung in tumor initiation and promotion studies.

The skin and lung tissues from SENCAR mice used as part of the Environmental Protection Agency's (EPA's) Carcinogenesis Testing Matrix were examined. This study included SENCAR mice used in three different short-term bioassay protocols in which the skin papilloma assay was used to identify initiators, promoters, and complete carcinogens. Also included were the pathology findings from SENCAR mice used in the combined bioassay in which the skin assay and the lung adenoma assay were conducted simultaneously. The gross and microscopic features of treatment-associated and spontaneous lesions of the skin and lung of the SENCAR mouse used in these studies are defined and the lesions most commonly observed are described. Generally, gross observations and microscopic findings in both the skin and lung tissues were poorly correlated. Although there are several definite criteria on which gross interpretations of the various skin and lung lesions can be made, with the exception of pedunculated squamous cell papillomas and the classic squamous cell carcinomas, the various lesion types had a wide variety of clinical presentations that severely compromised the accuracy of gross diagnosis. Further, in the case of benign skin neoplasms, malignant transformation of these tumors most often occurred at the base of the lesion and was initially hidden from gross observation. As a result, approximately 50% of the neoplasms interpreted clinically as benign tumors (papillomas and keratoacanthomas) were actually malignant neoplasms. Moreover, many lesions determined grossly to be nontumorous were in fact found to be neoplastic when examined microscopically. The SENCAR mouse was found to be more responsive in the lung adenoma assay than other strains examined with exception of the Strain A. Although accurate interpretation of the lung lesions in the SENCAR was compromised by nonneoplastic treatment-associated and/or spontaneous lesions, the feasibility of using the SENCAR skin and lung as target tissues in two-stage combined carcinogenesis studies merits further consideration.

Comparative carcinogenesis by hydroxylated nitrosopropylamines in Syrian hamsters.

The relationship between the chemical structure of nitrosamines and their carcinogenic activity has been examined in Syrian golden hamsters in parallel with similar studies in rats to aid in explaining the sharp interspecies differences in response to these compounds. The relationship between the beta-oxidized N-propyl-nitrosamine structure and the induction of tumors of the pancreatic duct in Syrian golden hamsters was investigated by administration of a number of asymmetric acyclic nitrosamines containing that structure to female hamsters for 29-50 weeks. N-Nitroso-2-oxopropyl-2-hydroxyethylamine (OPE), N-nitroso-2-hydroxypropyl-2-hydroxyethylamine (NIEA), and N-nitroso-2,3-dihydroxypropyl-2-oxopropylamine (DHPOP) induced pancreatic tumors. OPE also induced a high incidence of liver neoplasms, and a number of animals given NIEA and N-nitrosoallyl-2-oxopropylamine (NAOP) also had liver neoplasms. N-Nitroso-2,3-dihydroxypropyl-2-hydroxyethylamine was very weakly carcinogenic. N-Nitroso-2,3-dihydroxypropyl-2-hydroxypropylamine and DHPOP induced a high incidence of neoplasms of the forestomach (mainly papillomas). N-Nitrosoallyl-2,3-dihydroxypropylamine, N-nitrosoallyl-2-hydroxypropylamine, and NAOP induced primarily neoplasms of the nasal mucosa but no neoplasms of the pancreatic ducts in hamsters.

Effect of subsequent treatment of chloroform or phenobarbital on the incidence of liver and lung tumors initiated by ethylnitrosourea in 15 day old mice.

The effect of subsequent administration of chloroform or phenobarbital on the incidence of ethylnitrosourea (ENU) initiated liver and lung tumors was investigated. Fifteen day old Swiss mice were administered ENU, and at weaning they started to receive either 1800 p.p.m. chloroform or 500 p.p.m. sodium phenobarbital in their drinking water. The mice continued to receive either chloroform or phenobarbital until 51 weeks of age. They were sacrificed at 52 weeks of age. ENU at 5 and 20 mg/kg, caused a dose-dependent increase in liver and lung tumors. The male mice were more sensitive to the induction of liver tumors, while no sex preference was observed for the induction of lung tumors. In male mice chloroform inhibited, while in female and male mice phenobarbital promoted spontaneous and ENU-induced liver tumors. Subsequent treatment with either chloroform or phenobarbital did not affect the incidence of ENU-induced lung tumors. In conclusion, when administered in the drinking water, chloroform inhibited while phenobarbital promoted hepatocarcinogenesis in mice.

Carcinogenic effect of nitrosoalkylureas and nitrosoalkylcarbamates in Syrian hamsters.

Three nitrosoalkylureas, two nitrosotrialkylureas, and three nitrosoalkylcarbamates were given to Syrian golden hamsters by gavage at approximately equimolar doses. Measured by the time to death with tumors as an index, nitrosoethylurea was the most potent carcinogen, followed by nitroso-2-hydroxyethylurea, which was less effective in males than in females. The least effective compounds, by this measure, were nitrosooxazolidone and nitroso-5-methyloxazolidone. The remaining compounds, nitroso-N-ethylurethan, nitroso-2-hydroxypropylurea, nitrosomethyldiethylurea, and nitrosotriethylurea appeared to be of similar potency. All of the compounds induced papillomas or carcinomas of the nonglandular stomach in high incidence, except in the groups given nitrosohydroxyethylurea or nitrosooxazolidone; exceptionally, only 35% of the latter group had tumors, compared with 70% or more in the other groups. All of the nitrosoalkylureas induced a high incidence of hemangiosarcomas of the spleen, but the nitrosoalkylcarbamates did not. The quite uniform response of the hamster to these compounds contrasts with the great variety of organs and cell types in which they induce tumors in the rat.

Carcinogenesis by oxygenated nitrosomethylpropylamines in Syrian hamsters.

Three oxygenated propylnitrosomethylamines were administered to female Syrian hamsters at doses similar to those which had induced high incidences of esophageal neoplasms in rats. Nitrosomethyl-2-oxopropylamine (NMOP) given at the rate of 2 mg/animal/week, whether as one application of 2 mg or two applications of 1 mg, led to early death of the animals, mostly with liver neoplasms; administration of 1 mg/animal/week led to longer survival, but most animals died with both liver neoplasms and neoplasms of the nasal mucosa. Only one hamster treated with NMOP had a neoplasm of the pancreatic duct. Of the 14 hamsters treated with the higher dose of nitrosomethyl-2-hydroxypropylamine (NMHP) and surviving beyond 6 weeks, most had liver neoplasms and nine had neoplasms of the pancreatic ducts. At the lower dose of NMHP, most hamsters developed neoplasms of the nasal mucosa, as did those receiving the same dose of NMOP, and seven animals had hemangioendothelial tumors of the liver, but only one animal had a carcinoma of the pancreatic duct. Nitrosomethyldihydroxypropylamine (NMDHP) was a much weaker carcinogen than the other two compounds and induced mainly neoplasms of the nasal mucosa, with little shortening of life.

Carcinogenesis by nitrosomorpholines, nitrosooxazolidines and nitrosoazetidine given by gavage to Syrian golden hamsters.

Five cyclic nitrosamines, four containing oxygen in the ring, were administered by gavage to groups of 20 male Syrian golden hamsters. After administration of very similar doses, nitrosomorpholine, nitroso-2-methylmorpholine and nitroso-5-methyl-1,3-oxazolidine caused the animals to die with tumors after similar times, but nitrosomorpholine induced mainly tumors of the nasal cavity (and a few of the trachea), whereas the 2-methyl derivative induced tumors of the nasal cavity and liver. While nitroso-1,3-oxazolidine and its 5-methyl derivative both induced liver tumors (but no tumors in the nasal cavity) those induced by the former compound took much longer to kill the animals. Nitrosoazetidine, a liver carcinogen in rats, but which had been reported to be inactive in hamsters, did induce tumors of the liver in 30% of hamsters after a much larger dose than the other cyclic nitrosamines.

Comparison of the carcinogenic effectiveness of N-nitrosobis(2-hydroxypropyl)amine, N-nitrosobis(2-oxopropyl)amine, N-nitroso(2-hydroxypropyl)(2-oxopropyl)amine, and N-nitroso-2,6-dimethylmorpholine in Syrian hamsters.

For examination of metabolic interrelationships in carcinogenesis between N-nitroso-2,6-dimethylmorpholine, N-nitrosobis(2-oxopropyl)amine (CAS: 60599-38-4), N-nitrosobis(2-hydroxypropyl)amine (CAS: 53609-64-6), and N-nitroso(2-hydroxypropyl) (2-oxopropyl)amine, each was given to a separate group of 20 female Syrian golden hamsters by gavage. All four compounds induced tumors of the pancreatic duct and lung tumors, but the incidences varied from one compound to another. In addition, N-nitrosobis(2-oxopropyl)amine and N-nitroso(2-hydroxypropyl)(2-oxopropyl)amine induced many hepatocellular and cholangiocellular neoplasms, which the other two compounds did not. On the basis of short time to death with tumors and the relatively low total dose administered, N-nitrosobis(2-oxopropyl) amine appeared to be the most potent carcinogen in the hamster among the four. N-nitroso(2-hydroxypropyl)(2-oxopropyl)amine was next in potency but was considerably weaker than N-nitrosobis(2-oxopropyl)amine. N-Nitroso-2,6-dimethylmorpholine, which was similar in potency to N-nitroso(2-hydroxypropyl) (2-oxopropyl)amine, however, did not induce a significant incidence of liver tumors of any type; and N-nitrosobis(2-hydroxypropyl) amine was considerably less potent than the other three compounds. These results did not support the opinion of N-nitroso(2-hydroxypropyl) (2-oxopropyl)amine as the proximate carcinogenic metabolite of all three compounds in the Syrian hamster but instead suggested that these compounds might have acted through formation of different and yet unknown carcinogenic intermediates.

Effect of adrenalectomy on cadmium- and turpentine-induced hepatic synthesis of metallothionein and alpha 2-macrofetoprotein in the rat.

Recent reports have strongly implicated glucocorticoids in the induction of hepatic metallothionein synthesis and hypozincemia which occurs in certain pathophysiologic conditions. Studies were performed in rats to determine the effect of adrenalectomy and glucocorticoid treatment on the hepatic accumulation of metallothionein subsequent to the administration of cadmium and turpentine, two diverse substances known to induce hypozincemia and hepatic synthesis of metallothionein as well as alpha 2-macrofetoprotein in intact rats. By 24 h, both substances induced significant hypozincemia, hepatic metallothionein accumulation, and a severe tissue inflammatory response in adrenalectomized rats. Adrenalectomy only prevented the increase in plasma alpha 2-macrofetoprotein concentration. Results indicate that hepatic synthesis of alpha 2-macrofetoprotein, but not metallothionein, is mediated by adrenal hormones. Thus, glucocorticoids do not play a "vital" role in hepatic metallothionein accumulation or hypozincemia induced by inflammatory stress, as previously postulated.

Dose-response of guinea pigs experimentally infected with aerosols of Legionella pneumophila.

Experiments were carried out to determine the infectivity, clinical course of disease, and lethality of aerosols of Legionella pneumophila for guinea pigs. The median infectious dose was less than 129 organisms; the 50% lethal dose was 1.4 x 10(5) organisms. In addition, the intraperitoneal 50% lethal dose was 3.0 x 10(6) cells, a value indicating that the organisms were less virulent by the intraperitoneal route than by aerosol. Nonfatal disease always included fever and weight loss. These signs were accompanied by sporadic bacteremia and dyspnea. Leukocyte counts were uninformative. In general, the severity of fever and extent of serologic (microagglutination titer) response were dose-related. The guinea pig may be used as a model for Legionnaires' disease, but the only dependable clinical criteria of infection after airborne challenge are weight loss, fever, and seroconversion.

Nonhuman primate model for the study of respiratory Klebsiella pneumoniae infection.

Squirrel monkeys were inoculated by the intratracheal inoculation of 700 Klebsiella pneumoniae organisms and developed lobar pneumonia in about 24 h. Characteristic clinical findings were fever, anorexia, and coughing. Laboratory findings included leukocytosis or leukopenia (with the latter more prominent in ultimately fatal infections), bacteremia, and shedding of bacteria into the pharynx. Infected monkeys showed increased plasma lysozyme activity as well as increased plasma ceruloplasmin, haptoglobin and alpha1-antitrypsin. The mortality rate was 60%, and the mean time of death was 50.5 h. Pathologically, the disease spread by means of Kohn's pores and other pathways that generally did not involve airways as a means of dissemination until about 30 h. Squirrel monkeys seem to be better models for human respiratory K. pneumoniae infection than rats or mice.