Cancer Letters; 1 ( 1 9 7 5 ) 2 5 - - 2 8 © E l s e v i e r / N o r t h - H o l l a n d , A m s t e r d a m - - P r i n t e d in T h e N e t h e r l a n d s
THE CARCINOGENIC EFFECT OF DIMETHYLNITROSAMINE ON THE CHINESE HAMSTER (CRICETULUS GRISEUS)
GERD REZNIK
A bteilung f~tr Experimentelle Pathologie, Medizinische Hochschule, Hannover, 3000 Hannover. Kleefeld, Karl-Wiechert-AUee 9 (G.F.R.) (Received 30 J u n e 1 9 7 5 )
SUMMARY
Three groups of Chinese hamsters (20 males, 20 females/group) received weekly subcutaneous injections for life of dimethylnitrosamine (DMN) at dose levels 0.20, 0.10 and 0.05 of the LDs0. Eighty-two to 100% of the animals developed tum~)rs of vascular origin. The neoplasms were primarily hepatic hemangioendotheliomas and their incidence was unrelated to dose or sex. Although animals in the low dose group received a total dose which was onethird that of the highest dose group, their survival times were only slightly longer.
INTRODUCTION
Liver tumors are readily induced in Syrian golden hamsters by dimethylnitrosamine (DMN) and may develop following several treatments with DMN or even a single administration [3, 9]. Chronic weekly DMN treatment of European hamsters induced malignant hemangioendotheliomas of the liver and kidneys, as well as hepatocellular carcinomas [5]. Liver tumors have also been reported in chronic tests using Syrian hamsters treated subcutaneously (s.c.) with DMN and in rats administered low doses of the c o m p o u n d for long periods of time [4]. Short-term treatment of rats with high, single or continuous, doses of DMN has produced kidney tumors [4, 6]. The present studies were performed to compare the reaction of the Chinese hamster to DMN treatment with those of the above species. MATERIAL AND METHODS
Sixty male and 60 female Chinese hamsters were divided into 3 equal groups and treated s.c. once weekly for life with DMN at levels of 0.20, 0.10, or 0.05 of the LDs0. Twenty male and 20 female control hamsters received injections of 0.9% physiological saline. All hamsters were 8 weeks old at the
26
beginning of the experiment. To determine the LDs0, WeiFs method [10] was used, with 7-days' observation of the animals. Hamsters were housed by sex in groups of 5 in Makrolon cages, Type III, under standard conditions and given a pelleted diet (Hope Farms RMH-TMB) and water ad libitum. Animals were allowed to die spontaneously or were killed when moribund. Surviving controls were sacrificed after 100 weeks. Hamsters were completely autopsied, and skulls decalcified in Decal (Scientific Products, Evanston, Ill.) and cut into transverse 2-mm sections. After they were embedded in 5 pm paraplast, graded sections of all organs were processed with hematoxylin and eosin and Van Gieson's for routine histologic examination. RESULTS AND DISCUSSION
The LDs0 of DMN given s.c. was 17.7 mg/kg b o d y weight (b.w.) for males and females. Survival rates, tumor numbers and types are given in Tables 1 and 2. Male and female hamsters exhibited almost equal survival times (Table 1), TABLE 1 TREATMENT, TOTAL DOSE AND SURVIVAL RATE OF DMN-TREATED CHINESE HAMSTERS Initial no. animals
20 20 20 20 20 20 20 20
Sex
Weekly dose ( m g / k g b.w.)
T o t a l dose ( m g / k g b.w.)
Survival rate SD
M F M F M F M F
3.54 3,54 1.77 1.77 0.89 0.89 no treatment no treatment
.~
SD
105.9 103.7 50.7 58.6 31.9 31.6 0 0
22.7 15.2 6.8 15.9 4.0 4.2 0 0
29.9 29.3 28.7 33.1 36.0 35.8 86.9 81.9
6.4 4.3 3.8 9.0 4.5 4.7 29.1 38.7
TABLE 2 C A R C I N O G E N E S I S O F D I M E T H Y L N I T R O S A M I N E IN C H I N E S E H A M S T E R S No. of h a m s t e r s w i t h t u m o r s of:
Dose (mg/kg b.w.)
Effective no. animals g F
Total tumor incidence (%) M F
Liver M F
Lungs M F
Nasal cavity M F
3.54 1.77 0.89 Control
17 16 19 20
82.4 93.8 100 0
14 15 19 0
0 0 0 0
2 0 0 0
17 20 19 19
100 85 94.7 5.3
aMetastasis o f a h e m a n g i o e n d o t h e l i o m a .
17 17 18 0
0 1a 0 0
1 0 0 1
27
which increased at the lower dose levels. Hamsters from the lowest dosage group received approximately one-third less DMN than did those from the highest group (Table 1). However, the t u m o r incidence did not differ markedly among the various groups and was nearly 100% for all dosage groups (Table 2). The first liver t u m o r occurred at the 26th experimental week in a male from the highest dosage group. All hepatic tumors were of vascular origin (mainly hemangioendotheliomas) (Fig. 1). Macroscopically, they appeared as grey-white white or red-brown nodules, measuring up to 10 mm in diameter. In most cases, t u m o r cells were situated in marginal regions of large hemorrhages, whereas neoplasms composed of densely packed cells were seen only occasionally. In addition to liver tumors, 3 animals in the highest dosage group also exhibited adenocarcinomas of the nasal cavity. A pulmonary metastasis with a structure resembling that of an hepatic hemangioendothelioma was observed in a female treated with 1.77 mg/kg b.w. DMN. It is apparent, in comparing the rates of DMN-induced tumors in European, Syrian and Chinese hamsters, that the Chinese hamsters developed the most liver neoplasms (100%) in all dosage groups, while in the Syrian hamster a high incidence of liver cell carcinomas, cholangiomas and hemangioendotheliomas was also observed [2, 8]. In mice, a predominant vascular c y t o t r o p h y has been reported [1] and a similar reaction to DMN treatment described in the Syrian hamster [7]. However, unlike the situation in the European hamster, there
Fig. 1. Malignant hepatic h e m a n g i o e n d o t h e l i o m a in a male Chinese hamster 35 weeks after beginning s.c. t r e a t m e n t with 0.89 m g / k g b.w. DMN.
28
was no apparent dose-response relationship in these experiments with the Chinese hamster. Furthermore, no sex differences were found in t u m o r rates. Of 120 DMN-treated Chinese hamsters, only one animal, a female, developed a pulmonary metastasis with characteristics of a malignant hepatic hemangioendothelioma. By contrast, Syrian [3] and European hamsters [5] demonsrrated a markedly higher percentage of pulmonary metastases. The uniformity of the tumors developed b y Chinese hamsters (mainly hepatic hemangioendotheliomas) is y e t another difference in the species' response to DMN. Also of interest is that the animals treated with 0.89 mg/kg b.w. received only one-third as much DMN as hamsters administered 3.54 mg/kg b.w., and still presented a greater tumor incidence than those in the high dose group, the t u m o r rate being dependent upon the longer life span of the animals. Neoplasms of other sites (adenocarcinomas of the nasal cavity) occurred in only 3 cases, whereas renal tumors, observed at a relatively high rate in the European hamster [5], were absent in the Chinese hamster. REFERENCES 1 Cardesa, A., Pour, P., Althoff, J., and Mohr, U. (1973): vascular tumors in female Swiss mice after intraperitoneal injection of dimethylnitrosamine. J. Natl. Cancer Inst., 51, 201--208. 2 Haas, H., Mohr, U., and Kriiger, F.W. (1973): Comparative studies with different doses of N-nitrosomorpholine, N-nitrosopiperidine, N-nitrosomethylurea, and dimethylnitrosamine in Syrian golden hamsters. J. Natl. Cancer Inst., 51, 1295--1301. 3 Herrold, K.M. (1967): Histogenesis of malignant liver tumors induced by dimethylnitrosamine. An experimental study in Syrian hamsters. J. Natl. Cancer Inst., 39, ! 0 9 9 - 1111. 4 Magee, P.N., and Barnes, J.M. (1962): Induction of kidney tumors in the rat with dimethylnitrosamine (N-nitrosodimethylamine). J. Path. Bact., 84, 19--31. 5 Mohr, U., Haas, H., and Hilfrich, J. (1974): The carcinogenic effects of dimethylnitrosamine and nitrosomethylurea in European hamsters (Cricetus cricetus L.). Br. J. Cancer, 29, 359--364. 6 Riopelle, J.L., and Jasmin, G. (1969): Nature, classification and nomenclature of kidney tumors induced in the rat by dimethylnitrosamine. J. Natl. Cancer Inst., 42, 643--662. 7 Stenb~ick, F., Ferrero, A., Montesano, R., and Shubik, P. (1973): Synergistic effect of ferric oxide on dimethylnitrosamine carcinogenesis in the Syrian golden hamster. Z. Krebsforsch., 79, 31--38. 8 Tomatis, L., Magee, P.N., and Shubik, P. (1964): Induction of liver tumors in the Syrian golden hamster by feeding dimethylnitrosamine. J. Natl. Cancer Inst., 33, 341-345. 9 Tomatis, L., and Cefis, F. (1967): The effects of multiple and single administration of dimethylnitrosamine to hamsters. Tumori, 43, 447--451. 10 Weil, C.S. (1952): Tables for convenient calculation of effective dose (LDs0 or EDs0 ) and instructions in their use. Biometrics, 8, 249--263.
THE CARCINOGENIC EFFECT OF DIMETHYLNITROSAMINE ON THE CHINESE HAMSTER (CRICETULUS GRISEUS)
GERD REZNIK
A bteilung f~tr Experimentelle Pathologie, Medizinische Hochschule, Hannover, 3000 Hannover. Kleefeld, Karl-Wiechert-AUee 9 (G.F.R.) (Received 30 J u n e 1 9 7 5 )
SUMMARY
Three groups of Chinese hamsters (20 males, 20 females/group) received weekly subcutaneous injections for life of dimethylnitrosamine (DMN) at dose levels 0.20, 0.10 and 0.05 of the LDs0. Eighty-two to 100% of the animals developed tum~)rs of vascular origin. The neoplasms were primarily hepatic hemangioendotheliomas and their incidence was unrelated to dose or sex. Although animals in the low dose group received a total dose which was onethird that of the highest dose group, their survival times were only slightly longer.
INTRODUCTION
Liver tumors are readily induced in Syrian golden hamsters by dimethylnitrosamine (DMN) and may develop following several treatments with DMN or even a single administration [3, 9]. Chronic weekly DMN treatment of European hamsters induced malignant hemangioendotheliomas of the liver and kidneys, as well as hepatocellular carcinomas [5]. Liver tumors have also been reported in chronic tests using Syrian hamsters treated subcutaneously (s.c.) with DMN and in rats administered low doses of the c o m p o u n d for long periods of time [4]. Short-term treatment of rats with high, single or continuous, doses of DMN has produced kidney tumors [4, 6]. The present studies were performed to compare the reaction of the Chinese hamster to DMN treatment with those of the above species. MATERIAL AND METHODS
Sixty male and 60 female Chinese hamsters were divided into 3 equal groups and treated s.c. once weekly for life with DMN at levels of 0.20, 0.10, or 0.05 of the LDs0. Twenty male and 20 female control hamsters received injections of 0.9% physiological saline. All hamsters were 8 weeks old at the
26
beginning of the experiment. To determine the LDs0, WeiFs method [10] was used, with 7-days' observation of the animals. Hamsters were housed by sex in groups of 5 in Makrolon cages, Type III, under standard conditions and given a pelleted diet (Hope Farms RMH-TMB) and water ad libitum. Animals were allowed to die spontaneously or were killed when moribund. Surviving controls were sacrificed after 100 weeks. Hamsters were completely autopsied, and skulls decalcified in Decal (Scientific Products, Evanston, Ill.) and cut into transverse 2-mm sections. After they were embedded in 5 pm paraplast, graded sections of all organs were processed with hematoxylin and eosin and Van Gieson's for routine histologic examination. RESULTS AND DISCUSSION
The LDs0 of DMN given s.c. was 17.7 mg/kg b o d y weight (b.w.) for males and females. Survival rates, tumor numbers and types are given in Tables 1 and 2. Male and female hamsters exhibited almost equal survival times (Table 1), TABLE 1 TREATMENT, TOTAL DOSE AND SURVIVAL RATE OF DMN-TREATED CHINESE HAMSTERS Initial no. animals
20 20 20 20 20 20 20 20
Sex
Weekly dose ( m g / k g b.w.)
T o t a l dose ( m g / k g b.w.)
Survival rate SD
M F M F M F M F
3.54 3,54 1.77 1.77 0.89 0.89 no treatment no treatment
.~
SD
105.9 103.7 50.7 58.6 31.9 31.6 0 0
22.7 15.2 6.8 15.9 4.0 4.2 0 0
29.9 29.3 28.7 33.1 36.0 35.8 86.9 81.9
6.4 4.3 3.8 9.0 4.5 4.7 29.1 38.7
TABLE 2 C A R C I N O G E N E S I S O F D I M E T H Y L N I T R O S A M I N E IN C H I N E S E H A M S T E R S No. of h a m s t e r s w i t h t u m o r s of:
Dose (mg/kg b.w.)
Effective no. animals g F
Total tumor incidence (%) M F
Liver M F
Lungs M F
Nasal cavity M F
3.54 1.77 0.89 Control
17 16 19 20
82.4 93.8 100 0
14 15 19 0
0 0 0 0
2 0 0 0
17 20 19 19
100 85 94.7 5.3
aMetastasis o f a h e m a n g i o e n d o t h e l i o m a .
17 17 18 0
0 1a 0 0
1 0 0 1
27
which increased at the lower dose levels. Hamsters from the lowest dosage group received approximately one-third less DMN than did those from the highest group (Table 1). However, the t u m o r incidence did not differ markedly among the various groups and was nearly 100% for all dosage groups (Table 2). The first liver t u m o r occurred at the 26th experimental week in a male from the highest dosage group. All hepatic tumors were of vascular origin (mainly hemangioendotheliomas) (Fig. 1). Macroscopically, they appeared as grey-white white or red-brown nodules, measuring up to 10 mm in diameter. In most cases, t u m o r cells were situated in marginal regions of large hemorrhages, whereas neoplasms composed of densely packed cells were seen only occasionally. In addition to liver tumors, 3 animals in the highest dosage group also exhibited adenocarcinomas of the nasal cavity. A pulmonary metastasis with a structure resembling that of an hepatic hemangioendothelioma was observed in a female treated with 1.77 mg/kg b.w. DMN. It is apparent, in comparing the rates of DMN-induced tumors in European, Syrian and Chinese hamsters, that the Chinese hamsters developed the most liver neoplasms (100%) in all dosage groups, while in the Syrian hamster a high incidence of liver cell carcinomas, cholangiomas and hemangioendotheliomas was also observed [2, 8]. In mice, a predominant vascular c y t o t r o p h y has been reported [1] and a similar reaction to DMN treatment described in the Syrian hamster [7]. However, unlike the situation in the European hamster, there
Fig. 1. Malignant hepatic h e m a n g i o e n d o t h e l i o m a in a male Chinese hamster 35 weeks after beginning s.c. t r e a t m e n t with 0.89 m g / k g b.w. DMN.
28
was no apparent dose-response relationship in these experiments with the Chinese hamster. Furthermore, no sex differences were found in t u m o r rates. Of 120 DMN-treated Chinese hamsters, only one animal, a female, developed a pulmonary metastasis with characteristics of a malignant hepatic hemangioendothelioma. By contrast, Syrian [3] and European hamsters [5] demonsrrated a markedly higher percentage of pulmonary metastases. The uniformity of the tumors developed b y Chinese hamsters (mainly hepatic hemangioendotheliomas) is y e t another difference in the species' response to DMN. Also of interest is that the animals treated with 0.89 mg/kg b.w. received only one-third as much DMN as hamsters administered 3.54 mg/kg b.w., and still presented a greater tumor incidence than those in the high dose group, the t u m o r rate being dependent upon the longer life span of the animals. Neoplasms of other sites (adenocarcinomas of the nasal cavity) occurred in only 3 cases, whereas renal tumors, observed at a relatively high rate in the European hamster [5], were absent in the Chinese hamster. REFERENCES 1 Cardesa, A., Pour, P., Althoff, J., and Mohr, U. (1973): vascular tumors in female Swiss mice after intraperitoneal injection of dimethylnitrosamine. J. Natl. Cancer Inst., 51, 201--208. 2 Haas, H., Mohr, U., and Kriiger, F.W. (1973): Comparative studies with different doses of N-nitrosomorpholine, N-nitrosopiperidine, N-nitrosomethylurea, and dimethylnitrosamine in Syrian golden hamsters. J. Natl. Cancer Inst., 51, 1295--1301. 3 Herrold, K.M. (1967): Histogenesis of malignant liver tumors induced by dimethylnitrosamine. An experimental study in Syrian hamsters. J. Natl. Cancer Inst., 39, ! 0 9 9 - 1111. 4 Magee, P.N., and Barnes, J.M. (1962): Induction of kidney tumors in the rat with dimethylnitrosamine (N-nitrosodimethylamine). J. Path. Bact., 84, 19--31. 5 Mohr, U., Haas, H., and Hilfrich, J. (1974): The carcinogenic effects of dimethylnitrosamine and nitrosomethylurea in European hamsters (Cricetus cricetus L.). Br. J. Cancer, 29, 359--364. 6 Riopelle, J.L., and Jasmin, G. (1969): Nature, classification and nomenclature of kidney tumors induced in the rat by dimethylnitrosamine. J. Natl. Cancer Inst., 42, 643--662. 7 Stenb~ick, F., Ferrero, A., Montesano, R., and Shubik, P. (1973): Synergistic effect of ferric oxide on dimethylnitrosamine carcinogenesis in the Syrian golden hamster. Z. Krebsforsch., 79, 31--38. 8 Tomatis, L., Magee, P.N., and Shubik, P. (1964): Induction of liver tumors in the Syrian golden hamster by feeding dimethylnitrosamine. J. Natl. Cancer Inst., 33, 341-345. 9 Tomatis, L., and Cefis, F. (1967): The effects of multiple and single administration of dimethylnitrosamine to hamsters. Tumori, 43, 447--451. 10 Weil, C.S. (1952): Tables for convenient calculation of effective dose (LDs0 or EDs0 ) and instructions in their use. Biometrics, 8, 249--263.
