Transplacental Effects of Diethylnitrosamine in Syrian Hamsters as Related to Different Days of Administration During Pregnancy 1, 2 Ulrich Mohr, Hildegard Reznik-Schuller, Gerd Reznik, and Jorn Hilfrich SUMMARY-Female Syrian hamsters were given a single sc dose of 45 mg diethylnitrosamine (DEN)/kg body weight on 1 of the 15 days of pregnancy. In the offspring of females treated on 1 of the first 11 days of pregnancy. no respiratory tract tumors were found. The offspring of mothers given DEN on 1 of the last 4 days (12-15) of pregnancy developed respiratory tract neoplasms at a rate of up to 95%. A lower incidence of tumors in other organs seemed independent of the day of DEN treatment.-J Natl Cancer Inst 55: 681-683, 1975.
Diethylnitrosamine (DEN) has a strong carcinogenic effect on the offspring of Syrian hamsters when it is administered to their mothers during pregnancy (1-4). In contrast to various other carcinogens, which mainly induced neurogenic tumors transplacentally (5-8), DEN caused respiratory tract neoplasms in both mothers and their offspring (1-4). To supplement these investigations, our study was designed to elicit the precise day or days of pregnancy on which a single dose of DEN induces the highest tumor incidence in the offspring of Syrian hamsters. Our results should be compared with corressponding investigations of the fetal differentiation of the tracheal epithelium.
3, •
of pregnancy with the solvent alone served as a vehicle control. (DEN was synthesized by Dr. F. W. Kruger, Krebsforsuchungszentrum, Heidelberg, West Germany; it had a purity of 99.5%.) The day after mating was counted as day 1. The litters were weaned on the 25th day after birth and caged in Makrolon cages, Type III (900 cm 2), separated according to sex and litter. The mother, held separately, and the offspring were observed until spontaneous death. Thereafter, complete autopsies were performed; all organs were fixed in 4% buffered formalin, skulls were decalcified with Decal (Scientific Products, Evanston, Ill.) and all tissues were embedded in Paraplast. The larynx, cranial, and middle and caudal parts of the trachea, as well as its bifurcation, were embedded separately and cut into graded sections. Histologic sections were routinely stained with hematoxylin and eosin and those of the respiratory system were also stained with alcian blue and periodic acid-Schiff. Only those offspring and their respective mothers surviving the weaning period were evaluated. Cannibalism of litters by their treated mothers was high, so that several groups were small. RESULTS
MATERIALS AND METHODS
Neither mothers nor offspring in the control group developed tumors of the respiratory system (tables 1, 2) or any other organ. Survival of both treated and control females was shorter than that of males (table 3). The DEN-treated mothers developed respiratory tract tumors at a rate of up to 100% (table 1); the earliest neoplasm, a tracheal squamous cell papilloma, was found the I I th
One hundred and five female Syrian hamsters (Centraal Proe£dierenbedrij£ TNO, Zeist, Holland) were hand-mated and individually caged in Makrolon cages Type II (polyacryl, 400 cm2; Becker and Co. GmbH, Castrop-Rauxel, West Germany)_ The animals were kept under standard laboratory conditions (room temperature 22±2°C; relative humidity 55±5'%; air exchange 8 timesjhr) and given a pellet diet (Hope Farms RMHTMB, Woerden, Holland) and water ad libitum. Fifteen groups, each consisting of 6 pregnant females, were given a single sc injection of 45 mg DEN jkg body weight in solution with physiologic saline on I of the 15 days of pregnancy. An equivalent group treated on day 15
Received March 11, 1975; accepted May 17, 1975. Supported by the Deutsche Forschungsgemeinschaft. Abteilung fur Experimentelle Pathologie, Medizinische Hoch· schule Hannover, 3000 Hannover·Kleefe1d, Karl-Wiechert-Allee 9, West Germany. 4 We are grateful to Naoma Crisp·Lindgren for her assistance with the manuscript. 1
2 3
TABLE I.-Tumors in mothers treated with DEN on different days of pregnancy and in control mothers treated with NaCl Treatment (mg/DEN) 45 45 45 45 45 45 45 45 45 45 45 45 45 45 45
1 ml NaCla
Day of treatment
Larynx
Trachea
Lungs
Total No. of animals
Tumor-bearing animals
1 2 3 4 5 6 7
0 2 2 1 0 0 0 0 1 1 2 1 1 0 0 0
3 3
0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0
3
3
3 6 4 1 5 2 3 2 3 3 4 5 4 5 6
3 6 4 1
8 9 10 11 12 13 14 15 15
5
3 1
3 1 0 1 2 3 2
3 3 3 0
3 1 0 2 3 3
3 ~
3 3
0
• Control; no DEN administered. JOURNAL OF THE NATIONAL CANCER INSTITUTE, VOL. 55, NO.3, SEPTEMBER 1975
681
682
MOHR, REZNIK·SCHULLER., REZNIK, AND HILFR.ICH
TABLE 2.-Respiratory tract tumors in offspring of mothers treated on different days of pregnancy with a single dose of DEN or N aCI (control group) Treatment
Day of treatment
Nasal cavity
Larynx
Trachea
Lungs
1-11 12 13 14 15 15
0 1 1 0 3 0
0
0 1 2 8 10 0
0 0 0 1 3
DEN .... __ DEN_ .... _ DEN ______ DEN ______ DEN .. ____ NaCL. __ ._
2 5 8 5 0
Total No. of animals
Tumor·bearing animals
174
0 4 8 17 19 0
10
20 24 20 36
0
TABLE 3.-Survival times in weeks of mothers treated on different days of pregnancy with a single dose of DEN, of their offspring, and of the control group Offspring Mothers Day of treatment
Treatment
DEN _. _________ DEN ___________ DEN _____ . _____ DEN ___________ DEN ___________ DEN __ ._________ DEN ___________ DEN ___________ DEN ___________ DEN ___________ DEN _________ ._ DEN ___________ DEN ___________ DEN ___________ DEN ___________ NaCL __________
1 2 3 4 5 6 7 8 9 10
12.4-31.6 16.9-53.1 12.9-72.5 13.6-41.7 0 12.6-63.1 44.8-46.2 5.1-31.5 19.5-70.5 24.5-57.5 21.9-54.8 4.3-50.7 10.8-62.2 14.9-38.6 24.0-41.1 43.8-79.4
22.0 35.0 41.2 27.8 44.0 37.8 45.5 18.3 44.5 41.0 38.3 27.5 33.0 26.8 32.6 65.6
11
12 13 14 15 15
week after DEN injection. As shown in table 1 and textfigure 1, in the mothers most neoplasms developed in the trachea; fewer tumors were in the larynx. Histolog· ically, they were squamous cell papillomas and papillary polyps. Only in one instance was a bronchogenic aden· oma found, and no tumors were observed in the nasal cavities. Aside from the respiratory tract tumors, no neoplasms of other sites were detected. The offspring of mothers receiving DEN on 1 of the first 11 prenatal days developed no respiratory tract tumors (table 2, text·fig. 2). However, a neoplastic reo sponse of the respiratory system to prenatal DEN admin· istration was observed in the young of mothers treated on I of the last 4 prenatal days (table 2, text·fig. 2). As shown in table 2 and text·figure 2, the incidence of respiratory tract tumors increased, the later the carcin·
I!l] Mothers •
% 100
Females
Males
Range
Mean
Mean
Range
Mean
Range
86.7 76.5 90.8 104.6 0 116.6 84.0 90.1 83.2 86.4 52.4 73.0 83.8 83.8 86.3 120.5
54.0-100.3 22.4-130.6 63.3-118.3 79.4-129.8 0 82.3-140.9 36.0-132.0 61.1-119.1 33.8-132.6 66.3-106.5 28.6-76.6 64.5-81.5 53.8-113.8 51.8-115.8 57.3-115.3 90.5-145.6
65.8 66.5 87.6 64.1 57.5 62.8 49.6 56.5 67.1 73.0 54.1 67.1 66.1 48.7 45.0 100.2
37.1-94.5 46.8-86.1 56.1-119.1 33.4-94.8 44.0-71.0 36.8-88.8 11.6-77.6 25.6-87.4 39.4-94.8 55.4-90.6 25.8-82.4 46.5-87.7 48.1-84.1 18.1-79.3 24.5--65.4 86.3-115.5
ogen was administered, and finally reached a frequency of 95% in the animals treated on the 15th day of gesta· tion (text-fig. 2). Most neoplasms were laryngeal and tracheal and occurred with an almost equal frequency (table 2, text-fig. I). Histologically, these tumors were squamous cell papil. lomas and papillary polyps which often partially or com· pletely obstructed the lumen of the organs and were always ~ultiple. One male treated on the 12th and I female treated on the 13th prenatal day developed squamous cell papillomas of the maxilloturbinals and nasoturbinals, respectively (table 2). Of the group given DEN on the 15th prenatal day, I male had a squamous cell carcinoma originating from the nasoturbinals, whereas 2 females developed adenocarcinomas of the endoturbinals. Bronchogenic adenomas were found in a male treated Tumours "10
Offspring
100 90
90
80
70
60
60
50
70
50
40
40
30
30
20
20
10
10 23456789101112131415
Nasal Cavity
larynx
Trachea
(.-Comparison of tumor distribution in the respira. tory tract of tumor-bearing mothers and their offspring.
TEXT-FlGURE
Day Of Pregnancy
lungs
2.-Respiratory tract tumors in offspring after DEN treatment of their mothers on different days of pregnancy.
TEXT-FIGURE
TRANSPLACENTAL DEN EFFECT IN HAMSTERS
on the 14th prenatal day and in 2 males and I female given DEN on the 15th gestational day (table 2). The nonrespiratory-tract tumors in the different groups were. histologically. adrenal cortical adenomas in those animals treated on I of the first 8 prenatal days (4 males. I female). whereas all adrenal gland neoplasms found in animals given DEN on the I lth to 13th prenatal days were pheochromocytomas (4 males. 1 female). Four males had squamous cell papillomas of the forestomach. whereas females showed no neoplastic growth in this organ. In the livers of 2 males. adenomas of the bile ducts were found. and I female developed a liver-cell carcinoma. Adenomas of the thyroid gland were diagnosed in 4 hamsters; only I was female. Other lesions included a uterine squamous cell carcinoma and. in males. I fibrosarcoma of the skin. I pancreatic islet cell carcinoma. I testicular fibrosarcoma. and I renal fibrosarcoma. DISCUSSION
Our results show that the time of administration is decisive for the transplacental carcinogenic effect of DEN in the Syrian hamster. Only the offspring of those mothers treated on I of the last 4 days of pregnancy developed respiratory tract tumors. Thus one can conclude that the fetal respiratory epithelium must reach a certain degree of differentiation before becoming susceptible to the carcinogenic effort of DEN. This supposition is supported by the basic investigations of Emura and Mohr (9). They reported the tracheal epithelium of the Syrian hamster to be composed of a single layer of columnar cells until the 12th prenatal day; its differentiation into three distinct cell types began thereafter. One could hypothesize two possible explanations for the inability of undifferentiated epithelial cells to respond with neoplasms to DEN. I) The early embryonic cells. due to a metabolic insufficiency. might fail to transform the DEN to its proximate carcinogenic metabolites. 2) The ceIls might lack a receptor system for the absorption and binding of the carcinogen. The first explanation coincides with the assumption of Druckrey (10). whose experimental results concerning the transplacental effects in rats of various "indirect" alkylating carcinogens suggested that the fetal enzyme system. necessary for the activation of the carcinogen. becomes operative only in the later prenatal phase. However. since the activation of the administered chemical to the proximate carcinogen already takes place in the mother. it is possible that the activated form of the carcinogen might reach the fetus. which supports the second theory. But probably it is too reactive and shortlived for this to happen (10). These results do not permit the conclusion that the transplacental carcinogenic effect of the DEN in the Syrian hamster is generally restricted to the last 4 prenatal days. The neoplastic response of the suprarenal glands. forestomach. liver. and thyroid in groups treated
683
before the 12th day indicates that the susceptibility of other organs to the carcinogen differs from that found decisive for tumor induction in the respiratory system. Similar variations in the sensitivity of different organs to the transplacental carcinogenic effect of various chemicals also have been described in rats (10-12). The possibility that certain amounts of the carcinogen have also been transferred via the milk can be excluded. because thin-layer chromatographic studies with He_ DEN demonstrated that the carcinogen had already disappeared from both mother's milk and blood 120 minutes after injection (13). The present studies confirm and expand the results of previous investigations of the transplacental carcinogenicity of DEN in the Syrian hamster (1-4). They prove that the 12th-15th days of gestation are the decisive period during which the fetus is most susceptible to postnatal development of respiratory tract tumors when their mothers were treated with DEN during pregnancy. REFERENCES (1) MOHR U, ALTHOFF J, WRBA H: I. Diaplacentare Wirkung des Carcinogens Diaethylnitrosamin beim Goldhamster. Z Krebsforsch 66:536--540, 1965 (2) MOHR U, AUTHALER A, ALTHOFF J: 2. Der Nachweis von Diaethylnitrosamin in Goldhamsterembryonen nach Behandlung des Muttertieres. Naturwissenschaften 52: 188-189, 1965 (3) MOHR U, ALTHOFF J, AUTHALAR A: Diaplacental effect of the carcinogen diethylnitrosamine in the golden hamster. Cancer Res 26:2349-2352, 1966 (4) MOHR U: Effects of diethylnitrosamine on fetal and suckling Syrian golden hamsters. Transplacental carcinogenesis. IARC Scientific Pub 4:65-70, 1973 (5) IVANKOVIC S, DRUCKREY H, PREUSSMANN R: Erzeugung neurogener Tumoren bei den Nachkommen nach einmaliger Injektion von Aethylnitrosoharnstoff an schwangere Ratten. Naturwissenschaften 53:410-411, 1966 (6) IVANKOVIC S, DRUCKREY H: Transplacentare Erzeugung maligner Tumoren des Nervensystems. Z Krebsforsch 71 :320360, 1968 (7) IVANKOVIC S, PRF.USSMANN R: Transplacentare Erzeugung rnaIigner Tumoren nach oraler Gabe von Xthylharnstoff und Nitrit an Ratten. Naturwissenschaften 57:460-461, 1970 (8) DRUCKERY H, IVANKOVIC S, PIU'USSMANN R, et al: Transplacental induction of neurogenic malignomas by 1,2-diethylhydrazine, azo- and azoxyethane in rats. Experientia 24:561562,1968 (9) EMURA M, MOHR U: Morphological studies on the development of tracheal epithelium in Syrian golden hamsters. Z Versuchstierkd, 1975. In press (10) DRUCKREY H: Chemical structure and action in transplacental carcinogenesis and teratogenesis. Transplacental carcinogenesis. IARC Scientific Publ 4:45-58. 1973 (11) DRUCKREY H. SCHAGEN B. IVANKOVIC S: Erzeugung neurogener Malignome durch einmalige Gabe von Xthylnitrosoharnstoff (ANH) an neugeborene und junge BD IX-Ratten. Z Krebsforsch 74:141-161.1970 (12) NAPALKOV NP: Some general considerations on the problem of transplacental carcjnogenesis. Transplacental carcinogenesis. IARC Scientific Publ 4:1-13. 1973 (13) SPIELHOFF R. BRESCH H. HONIG M, et al: Milk as a transport agent for diethylnitrosamine in Syrian golden hamsters. J Natl Cancer Inst 53:281-282, 1974
Diethylnitrosamine (DEN) has a strong carcinogenic effect on the offspring of Syrian hamsters when it is administered to their mothers during pregnancy (1-4). In contrast to various other carcinogens, which mainly induced neurogenic tumors transplacentally (5-8), DEN caused respiratory tract neoplasms in both mothers and their offspring (1-4). To supplement these investigations, our study was designed to elicit the precise day or days of pregnancy on which a single dose of DEN induces the highest tumor incidence in the offspring of Syrian hamsters. Our results should be compared with corressponding investigations of the fetal differentiation of the tracheal epithelium.
3, •
of pregnancy with the solvent alone served as a vehicle control. (DEN was synthesized by Dr. F. W. Kruger, Krebsforsuchungszentrum, Heidelberg, West Germany; it had a purity of 99.5%.) The day after mating was counted as day 1. The litters were weaned on the 25th day after birth and caged in Makrolon cages, Type III (900 cm 2), separated according to sex and litter. The mother, held separately, and the offspring were observed until spontaneous death. Thereafter, complete autopsies were performed; all organs were fixed in 4% buffered formalin, skulls were decalcified with Decal (Scientific Products, Evanston, Ill.) and all tissues were embedded in Paraplast. The larynx, cranial, and middle and caudal parts of the trachea, as well as its bifurcation, were embedded separately and cut into graded sections. Histologic sections were routinely stained with hematoxylin and eosin and those of the respiratory system were also stained with alcian blue and periodic acid-Schiff. Only those offspring and their respective mothers surviving the weaning period were evaluated. Cannibalism of litters by their treated mothers was high, so that several groups were small. RESULTS
MATERIALS AND METHODS
Neither mothers nor offspring in the control group developed tumors of the respiratory system (tables 1, 2) or any other organ. Survival of both treated and control females was shorter than that of males (table 3). The DEN-treated mothers developed respiratory tract tumors at a rate of up to 100% (table 1); the earliest neoplasm, a tracheal squamous cell papilloma, was found the I I th
One hundred and five female Syrian hamsters (Centraal Proe£dierenbedrij£ TNO, Zeist, Holland) were hand-mated and individually caged in Makrolon cages Type II (polyacryl, 400 cm2; Becker and Co. GmbH, Castrop-Rauxel, West Germany)_ The animals were kept under standard laboratory conditions (room temperature 22±2°C; relative humidity 55±5'%; air exchange 8 timesjhr) and given a pellet diet (Hope Farms RMHTMB, Woerden, Holland) and water ad libitum. Fifteen groups, each consisting of 6 pregnant females, were given a single sc injection of 45 mg DEN jkg body weight in solution with physiologic saline on I of the 15 days of pregnancy. An equivalent group treated on day 15
Received March 11, 1975; accepted May 17, 1975. Supported by the Deutsche Forschungsgemeinschaft. Abteilung fur Experimentelle Pathologie, Medizinische Hoch· schule Hannover, 3000 Hannover·Kleefe1d, Karl-Wiechert-Allee 9, West Germany. 4 We are grateful to Naoma Crisp·Lindgren for her assistance with the manuscript. 1
2 3
TABLE I.-Tumors in mothers treated with DEN on different days of pregnancy and in control mothers treated with NaCl Treatment (mg/DEN) 45 45 45 45 45 45 45 45 45 45 45 45 45 45 45
1 ml NaCla
Day of treatment
Larynx
Trachea
Lungs
Total No. of animals
Tumor-bearing animals
1 2 3 4 5 6 7
0 2 2 1 0 0 0 0 1 1 2 1 1 0 0 0
3 3
0 0 0 0 0 0 0 0 0 0 0 0 0 1 0 0
3
3
3 6 4 1 5 2 3 2 3 3 4 5 4 5 6
3 6 4 1
8 9 10 11 12 13 14 15 15
5
3 1
3 1 0 1 2 3 2
3 3 3 0
3 1 0 2 3 3
3 ~
3 3
0
• Control; no DEN administered. JOURNAL OF THE NATIONAL CANCER INSTITUTE, VOL. 55, NO.3, SEPTEMBER 1975
681
682
MOHR, REZNIK·SCHULLER., REZNIK, AND HILFR.ICH
TABLE 2.-Respiratory tract tumors in offspring of mothers treated on different days of pregnancy with a single dose of DEN or N aCI (control group) Treatment
Day of treatment
Nasal cavity
Larynx
Trachea
Lungs
1-11 12 13 14 15 15
0 1 1 0 3 0
0
0 1 2 8 10 0
0 0 0 1 3
DEN .... __ DEN_ .... _ DEN ______ DEN ______ DEN .. ____ NaCL. __ ._
2 5 8 5 0
Total No. of animals
Tumor·bearing animals
174
0 4 8 17 19 0
10
20 24 20 36
0
TABLE 3.-Survival times in weeks of mothers treated on different days of pregnancy with a single dose of DEN, of their offspring, and of the control group Offspring Mothers Day of treatment
Treatment
DEN _. _________ DEN ___________ DEN _____ . _____ DEN ___________ DEN ___________ DEN __ ._________ DEN ___________ DEN ___________ DEN ___________ DEN ___________ DEN _________ ._ DEN ___________ DEN ___________ DEN ___________ DEN ___________ NaCL __________
1 2 3 4 5 6 7 8 9 10
12.4-31.6 16.9-53.1 12.9-72.5 13.6-41.7 0 12.6-63.1 44.8-46.2 5.1-31.5 19.5-70.5 24.5-57.5 21.9-54.8 4.3-50.7 10.8-62.2 14.9-38.6 24.0-41.1 43.8-79.4
22.0 35.0 41.2 27.8 44.0 37.8 45.5 18.3 44.5 41.0 38.3 27.5 33.0 26.8 32.6 65.6
11
12 13 14 15 15
week after DEN injection. As shown in table 1 and textfigure 1, in the mothers most neoplasms developed in the trachea; fewer tumors were in the larynx. Histolog· ically, they were squamous cell papillomas and papillary polyps. Only in one instance was a bronchogenic aden· oma found, and no tumors were observed in the nasal cavities. Aside from the respiratory tract tumors, no neoplasms of other sites were detected. The offspring of mothers receiving DEN on 1 of the first 11 prenatal days developed no respiratory tract tumors (table 2, text·fig. 2). However, a neoplastic reo sponse of the respiratory system to prenatal DEN admin· istration was observed in the young of mothers treated on I of the last 4 prenatal days (table 2, text·fig. 2). As shown in table 2 and text·figure 2, the incidence of respiratory tract tumors increased, the later the carcin·
I!l] Mothers •
% 100
Females
Males
Range
Mean
Mean
Range
Mean
Range
86.7 76.5 90.8 104.6 0 116.6 84.0 90.1 83.2 86.4 52.4 73.0 83.8 83.8 86.3 120.5
54.0-100.3 22.4-130.6 63.3-118.3 79.4-129.8 0 82.3-140.9 36.0-132.0 61.1-119.1 33.8-132.6 66.3-106.5 28.6-76.6 64.5-81.5 53.8-113.8 51.8-115.8 57.3-115.3 90.5-145.6
65.8 66.5 87.6 64.1 57.5 62.8 49.6 56.5 67.1 73.0 54.1 67.1 66.1 48.7 45.0 100.2
37.1-94.5 46.8-86.1 56.1-119.1 33.4-94.8 44.0-71.0 36.8-88.8 11.6-77.6 25.6-87.4 39.4-94.8 55.4-90.6 25.8-82.4 46.5-87.7 48.1-84.1 18.1-79.3 24.5--65.4 86.3-115.5
ogen was administered, and finally reached a frequency of 95% in the animals treated on the 15th day of gesta· tion (text-fig. 2). Most neoplasms were laryngeal and tracheal and occurred with an almost equal frequency (table 2, text-fig. I). Histologically, these tumors were squamous cell papil. lomas and papillary polyps which often partially or com· pletely obstructed the lumen of the organs and were always ~ultiple. One male treated on the 12th and I female treated on the 13th prenatal day developed squamous cell papillomas of the maxilloturbinals and nasoturbinals, respectively (table 2). Of the group given DEN on the 15th prenatal day, I male had a squamous cell carcinoma originating from the nasoturbinals, whereas 2 females developed adenocarcinomas of the endoturbinals. Bronchogenic adenomas were found in a male treated Tumours "10
Offspring
100 90
90
80
70
60
60
50
70
50
40
40
30
30
20
20
10
10 23456789101112131415
Nasal Cavity
larynx
Trachea
(.-Comparison of tumor distribution in the respira. tory tract of tumor-bearing mothers and their offspring.
TEXT-FlGURE
Day Of Pregnancy
lungs
2.-Respiratory tract tumors in offspring after DEN treatment of their mothers on different days of pregnancy.
TEXT-FIGURE
TRANSPLACENTAL DEN EFFECT IN HAMSTERS
on the 14th prenatal day and in 2 males and I female given DEN on the 15th gestational day (table 2). The nonrespiratory-tract tumors in the different groups were. histologically. adrenal cortical adenomas in those animals treated on I of the first 8 prenatal days (4 males. I female). whereas all adrenal gland neoplasms found in animals given DEN on the I lth to 13th prenatal days were pheochromocytomas (4 males. 1 female). Four males had squamous cell papillomas of the forestomach. whereas females showed no neoplastic growth in this organ. In the livers of 2 males. adenomas of the bile ducts were found. and I female developed a liver-cell carcinoma. Adenomas of the thyroid gland were diagnosed in 4 hamsters; only I was female. Other lesions included a uterine squamous cell carcinoma and. in males. I fibrosarcoma of the skin. I pancreatic islet cell carcinoma. I testicular fibrosarcoma. and I renal fibrosarcoma. DISCUSSION
Our results show that the time of administration is decisive for the transplacental carcinogenic effect of DEN in the Syrian hamster. Only the offspring of those mothers treated on I of the last 4 days of pregnancy developed respiratory tract tumors. Thus one can conclude that the fetal respiratory epithelium must reach a certain degree of differentiation before becoming susceptible to the carcinogenic effort of DEN. This supposition is supported by the basic investigations of Emura and Mohr (9). They reported the tracheal epithelium of the Syrian hamster to be composed of a single layer of columnar cells until the 12th prenatal day; its differentiation into three distinct cell types began thereafter. One could hypothesize two possible explanations for the inability of undifferentiated epithelial cells to respond with neoplasms to DEN. I) The early embryonic cells. due to a metabolic insufficiency. might fail to transform the DEN to its proximate carcinogenic metabolites. 2) The ceIls might lack a receptor system for the absorption and binding of the carcinogen. The first explanation coincides with the assumption of Druckrey (10). whose experimental results concerning the transplacental effects in rats of various "indirect" alkylating carcinogens suggested that the fetal enzyme system. necessary for the activation of the carcinogen. becomes operative only in the later prenatal phase. However. since the activation of the administered chemical to the proximate carcinogen already takes place in the mother. it is possible that the activated form of the carcinogen might reach the fetus. which supports the second theory. But probably it is too reactive and shortlived for this to happen (10). These results do not permit the conclusion that the transplacental carcinogenic effect of the DEN in the Syrian hamster is generally restricted to the last 4 prenatal days. The neoplastic response of the suprarenal glands. forestomach. liver. and thyroid in groups treated
683
before the 12th day indicates that the susceptibility of other organs to the carcinogen differs from that found decisive for tumor induction in the respiratory system. Similar variations in the sensitivity of different organs to the transplacental carcinogenic effect of various chemicals also have been described in rats (10-12). The possibility that certain amounts of the carcinogen have also been transferred via the milk can be excluded. because thin-layer chromatographic studies with He_ DEN demonstrated that the carcinogen had already disappeared from both mother's milk and blood 120 minutes after injection (13). The present studies confirm and expand the results of previous investigations of the transplacental carcinogenicity of DEN in the Syrian hamster (1-4). They prove that the 12th-15th days of gestation are the decisive period during which the fetus is most susceptible to postnatal development of respiratory tract tumors when their mothers were treated with DEN during pregnancy. REFERENCES (1) MOHR U, ALTHOFF J, WRBA H: I. Diaplacentare Wirkung des Carcinogens Diaethylnitrosamin beim Goldhamster. Z Krebsforsch 66:536--540, 1965 (2) MOHR U, AUTHALER A, ALTHOFF J: 2. Der Nachweis von Diaethylnitrosamin in Goldhamsterembryonen nach Behandlung des Muttertieres. Naturwissenschaften 52: 188-189, 1965 (3) MOHR U, ALTHOFF J, AUTHALAR A: Diaplacental effect of the carcinogen diethylnitrosamine in the golden hamster. Cancer Res 26:2349-2352, 1966 (4) MOHR U: Effects of diethylnitrosamine on fetal and suckling Syrian golden hamsters. Transplacental carcinogenesis. IARC Scientific Pub 4:65-70, 1973 (5) IVANKOVIC S, DRUCKREY H, PREUSSMANN R: Erzeugung neurogener Tumoren bei den Nachkommen nach einmaliger Injektion von Aethylnitrosoharnstoff an schwangere Ratten. Naturwissenschaften 53:410-411, 1966 (6) IVANKOVIC S, DRUCKREY H: Transplacentare Erzeugung maligner Tumoren des Nervensystems. Z Krebsforsch 71 :320360, 1968 (7) IVANKOVIC S, PRF.USSMANN R: Transplacentare Erzeugung rnaIigner Tumoren nach oraler Gabe von Xthylharnstoff und Nitrit an Ratten. Naturwissenschaften 57:460-461, 1970 (8) DRUCKERY H, IVANKOVIC S, PIU'USSMANN R, et al: Transplacental induction of neurogenic malignomas by 1,2-diethylhydrazine, azo- and azoxyethane in rats. Experientia 24:561562,1968 (9) EMURA M, MOHR U: Morphological studies on the development of tracheal epithelium in Syrian golden hamsters. Z Versuchstierkd, 1975. In press (10) DRUCKREY H: Chemical structure and action in transplacental carcinogenesis and teratogenesis. Transplacental carcinogenesis. IARC Scientific Publ 4:45-58. 1973 (11) DRUCKREY H. SCHAGEN B. IVANKOVIC S: Erzeugung neurogener Malignome durch einmalige Gabe von Xthylnitrosoharnstoff (ANH) an neugeborene und junge BD IX-Ratten. Z Krebsforsch 74:141-161.1970 (12) NAPALKOV NP: Some general considerations on the problem of transplacental carcjnogenesis. Transplacental carcinogenesis. IARC Scientific Publ 4:1-13. 1973 (13) SPIELHOFF R. BRESCH H. HONIG M, et al: Milk as a transport agent for diethylnitrosamine in Syrian golden hamsters. J Natl Cancer Inst 53:281-282, 1974
