Europ. ,1".Canter Vol. 12, pp. 1017-1019. Pergamon Press 1976. Printed in Great Britain
Letter to the Editor
Reduction by Pituitary Grafts of Mammary Tumor Age. Its Variability in ,a High Mammary Tumor Strain of Mice: Effects on Mammary DNA Synthesis* HIROSHI NAGASAWA, REIKO YANAI and HISASHI TANIGUCHI Pharmacology Division, National Cancer Center Research Institute, Tsukiji 5-1-1, Chuo-ku, Tokyo 104, Japan
as the control. Each mouse w a s checked for palpable m a m m a r y tumors every 7 days Until it died or became moribund. At about 70 days of age, some mice of both groups were given an intraperitoneal injection of 50/~Ci 3H-thymidine (5 Ci/mmole; T h e Radiochemical Centre, Amersham, England) and 2 hours later 3H-thymidine incorporated into m a m m a r y DNA was assayed [9] as the index of m a m m a r y DNA synthesis. M a m m a r y tumor incidences were 95.9% (47/49) and 88-5% (128/145) in the experimental and control groups, respectively, n o significant difference being observed. The normal distribution curves [10] o f m a m m a r y tumor age are shown in Fig. 1. The mice with pituitary grafts had significantly earlier m a m m a r y tumor age than the control (P < 0.01). Moreover, a marked difference in the variation of tumor age between groups was observed as shown by the standard deviation and the fiducial limit. Thirty-six per c e n t of the grafted mice developed m a m m a r y tumors at an age corresponding to the average age, and 70 % showed tumors within one month before and after the average age. Meanwhile, only 20 % of the control mice had a tumor age similar to the average, and the number of mice showing tumors one month before and after the average age was less than 35%. These indicate that the variation in .mammary tumor
EVEN in high m a m m a r y tumor strains of mice maintained under the similar conditions, there may be considerable variation in m a m m a r y tumor age. Pituitary grafts at young age [1] or neonatally [2] as well as after maturity [3, 4] promote m a m m a r y tumorigenesis in mice and prolactin from the grafts has a major part in this process [3, 4]. However, in all of these experiments, the primary attention was paid to m a m m a r y t u m o r incidence, but neither to m a m m a r y tumor age nor to its variation. I n view of the importance of m a m m a r y DNA synthesis around the time when carcinogenic agents act on the glands in m a m m a r y tumorigenesis [5-7], pituitaries grafted into very young animals should maintain high m a m m a r y DNA synthesis, which in turn might result in a less variable as well as earlier m a m m a r y tumor appearance. The present study deals with this problem. The m a m m a r y tumor susceptible SHN strain of virgin mice [8] were used. At weaning (20-23 days o f age), half of the females in each litter received 21isologous pituitary grafts under the right kidney capsule. The other females in the litter were given no treatment and served Accepted 11 June'. 1976 *Supported in part by grants-in-aid for Cancer Research from the Minist':y of Education, Science and Culture (No 901039) and the Society for Promotion of Cancer Research, Japan
1017
1018
Letter to the Editor
age is reduced by about one-half by the isografting of 2 pituitaries at weaning. The rates of mammary DNA synthesis at 70 days of age varied between 120 and 400 dpm/#g DNA and between 13 and 710 dpm/#g DNA, mostly less than 120 dpm/#g DNA in the
declined thereafter, while pituitary grafting constantly maintained high mammary DNA synthesis irrespective of the host age [11]. Therefore, the marked reduction in both mammary tumor age and its variability by pituitary grafting is mostly attributable to the
4C
I
2AP
M=59 N =47 SD=I2 FL = 2 . 4
3C
•
Z
o E e~ "o
E 4Control M'8.9
2c
/
E .E o E
~'
/
/
~
\
N = ,28 \ SO = 2.O
?,
to
g
E >b .E FI
o
1
2
5 Memmory
I0 Tumor
oge,
.el lee
15 months
Fig. I. Normal distribution curves of mammary tumor age. 2AP : grafted with 2 isologous pituitaries each at weaning. M : mean in months, N: number of mile, SD: standard deviation, FL : fiducial limit at P = 0.05.
experimental and control groups, respectively. The averages of 230 and 151 dpm/#g DNA were not significantly different owing to the large variation in the control (Fig. 2). The mammary gland is considered to be exposed to several carcinogenic agents throughout the lifetime of the animal. Thus, in view of the importance of mammary DNA synthesis in mammary tumor induction [5-7], the longer the period of high mammary DNA synthesis, the higher the probability of malignant transformation of mammary cells. In the present experiment, pituitary grafting at weaning markedly increased mammary DNA synthesis at 70 days of age. Mammary DNA synthesis of mammary tumor susceptible strains of mice shows peak around 70 days of age and
Control
r
2AP
Fig. 2. 3H-thymidine incorporation into mamma01 D N A in each group at about 70 days of age. 2AP: grafted with 2 isologouspituitaries at weaning.
continuous promotion of mammary DNA synthesis by prolactin from the grafts. Bern [ 12] inferred that hormone stimulation of mammary gland can create conditions favorable for the action of carcinogenic agents. The data support this hypothesis and strongly suggest that one role of prolactin in mammary tumor induction is to sensitize the gland to carcinogenic agents through its constant stimulation of mammary DNA synthesis. Acknowledgement--We thank Prof. H. A. Bern, Department of Zoology and Cancer Research Laboratory, University of California, Berkeley, California, U.S.A. and Dr. T. Mori, Zoological Institute, University of Tokyo, Tokyo, Japan for reading the manuscript and for valuable suggestions.
REFERENCES 1. 2.
D. MEDINA, K. B. DEOME and L. YOUNG, Tumor-producing capabilities of hyperplastic alveolar nodules in virgin and hormone-stimulated BALB/c f. C3H and C3Hfmice. d. nat. Cancer Inst. 44, 167 (1970). T. MoRx and H. A. BERN, Personal communication.
Letter to the Editor 3. 4. 5. 6. 7. 8.
9. 10. I 1. 12.
L . M . BOOT, Induction by prolactin of mammary tumours in mice. Verh. kon. Ned. Akad. Wet., Afd. Natuurk., Tweed, Reeks, deel LVIII, Suppl. 3. (1969). G. R6PcKE, Interaction of hypophyseal isografts and ovarian hormones in mammary tumour development in mice. Thesis, Mondeel-Offsetdrukkerij, Amsterdam (1975). H. NAOASAWAand R. YANAI, Frequency of mammary cell division in relation t o age: Its significance in the induction of mammary tumors by carcinogen in rats. J. nat. Cancer Inst. 52, 609 (1974). H. NAOASAWA,R. YANAI and H. TANmUCHI, Importance of mammary gland DNA synthesis in carcinogen-induced mammary tumorigenesis in rats. Cancer Res. 36, 2223 (1976). ]?,. YANAI and H. NAOASAWA,Effects of pituitary graft and 2-bromo-a-ergocryptine on mammary DNA synthesis in mice in relation to mammary tumorigenesis. J. nat. Cancer Inst. 56, 1055 (1976). H. NAOASAWA, R. YANAI, H. TANIOUCHI, R. TOKUZEN and the late W. NAXAI-IAaA, Two-way selection of a stock of Swiss albino mice for mammary tumorigenesis: Establishment of two new strains (SHN and SLN). J. nat. Cancer Inst. 57, In press (1976). H. NAOASAWAand R. YANAI, Effects of estrogen and/or pituitary graft on nucleic acid synthesis of carcinogen-induced mammary tumors in rats. d. nat. Cancer Inst. 52, 1219 (1974). G . W . SNEDECOR,Statistical Methods. 5th ed., p. 194. Iowa State Univ. Press., Ames. H. NAOASAWAand R. YANk, Unpublished. H . A . B~RN, Nature of the hormonal influence in mouse mammary cancer. Science 131~ 1039 (1960).
1019
Letter to the Editor
Reduction by Pituitary Grafts of Mammary Tumor Age. Its Variability in ,a High Mammary Tumor Strain of Mice: Effects on Mammary DNA Synthesis* HIROSHI NAGASAWA, REIKO YANAI and HISASHI TANIGUCHI Pharmacology Division, National Cancer Center Research Institute, Tsukiji 5-1-1, Chuo-ku, Tokyo 104, Japan
as the control. Each mouse w a s checked for palpable m a m m a r y tumors every 7 days Until it died or became moribund. At about 70 days of age, some mice of both groups were given an intraperitoneal injection of 50/~Ci 3H-thymidine (5 Ci/mmole; T h e Radiochemical Centre, Amersham, England) and 2 hours later 3H-thymidine incorporated into m a m m a r y DNA was assayed [9] as the index of m a m m a r y DNA synthesis. M a m m a r y tumor incidences were 95.9% (47/49) and 88-5% (128/145) in the experimental and control groups, respectively, n o significant difference being observed. The normal distribution curves [10] o f m a m m a r y tumor age are shown in Fig. 1. The mice with pituitary grafts had significantly earlier m a m m a r y tumor age than the control (P < 0.01). Moreover, a marked difference in the variation of tumor age between groups was observed as shown by the standard deviation and the fiducial limit. Thirty-six per c e n t of the grafted mice developed m a m m a r y tumors at an age corresponding to the average age, and 70 % showed tumors within one month before and after the average age. Meanwhile, only 20 % of the control mice had a tumor age similar to the average, and the number of mice showing tumors one month before and after the average age was less than 35%. These indicate that the variation in .mammary tumor
EVEN in high m a m m a r y tumor strains of mice maintained under the similar conditions, there may be considerable variation in m a m m a r y tumor age. Pituitary grafts at young age [1] or neonatally [2] as well as after maturity [3, 4] promote m a m m a r y tumorigenesis in mice and prolactin from the grafts has a major part in this process [3, 4]. However, in all of these experiments, the primary attention was paid to m a m m a r y t u m o r incidence, but neither to m a m m a r y tumor age nor to its variation. I n view of the importance of m a m m a r y DNA synthesis around the time when carcinogenic agents act on the glands in m a m m a r y tumorigenesis [5-7], pituitaries grafted into very young animals should maintain high m a m m a r y DNA synthesis, which in turn might result in a less variable as well as earlier m a m m a r y tumor appearance. The present study deals with this problem. The m a m m a r y tumor susceptible SHN strain of virgin mice [8] were used. At weaning (20-23 days o f age), half of the females in each litter received 21isologous pituitary grafts under the right kidney capsule. The other females in the litter were given no treatment and served Accepted 11 June'. 1976 *Supported in part by grants-in-aid for Cancer Research from the Minist':y of Education, Science and Culture (No 901039) and the Society for Promotion of Cancer Research, Japan
1017
1018
Letter to the Editor
age is reduced by about one-half by the isografting of 2 pituitaries at weaning. The rates of mammary DNA synthesis at 70 days of age varied between 120 and 400 dpm/#g DNA and between 13 and 710 dpm/#g DNA, mostly less than 120 dpm/#g DNA in the
declined thereafter, while pituitary grafting constantly maintained high mammary DNA synthesis irrespective of the host age [11]. Therefore, the marked reduction in both mammary tumor age and its variability by pituitary grafting is mostly attributable to the
4C
I
2AP
M=59 N =47 SD=I2 FL = 2 . 4
3C
•
Z
o E e~ "o
E 4Control M'8.9
2c
/
E .E o E
~'
/
/
~
\
N = ,28 \ SO = 2.O
?,
to
g
E >b .E FI
o
1
2
5 Memmory
I0 Tumor
oge,
.el lee
15 months
Fig. I. Normal distribution curves of mammary tumor age. 2AP : grafted with 2 isologous pituitaries each at weaning. M : mean in months, N: number of mile, SD: standard deviation, FL : fiducial limit at P = 0.05.
experimental and control groups, respectively. The averages of 230 and 151 dpm/#g DNA were not significantly different owing to the large variation in the control (Fig. 2). The mammary gland is considered to be exposed to several carcinogenic agents throughout the lifetime of the animal. Thus, in view of the importance of mammary DNA synthesis in mammary tumor induction [5-7], the longer the period of high mammary DNA synthesis, the higher the probability of malignant transformation of mammary cells. In the present experiment, pituitary grafting at weaning markedly increased mammary DNA synthesis at 70 days of age. Mammary DNA synthesis of mammary tumor susceptible strains of mice shows peak around 70 days of age and
Control
r
2AP
Fig. 2. 3H-thymidine incorporation into mamma01 D N A in each group at about 70 days of age. 2AP: grafted with 2 isologouspituitaries at weaning.
continuous promotion of mammary DNA synthesis by prolactin from the grafts. Bern [ 12] inferred that hormone stimulation of mammary gland can create conditions favorable for the action of carcinogenic agents. The data support this hypothesis and strongly suggest that one role of prolactin in mammary tumor induction is to sensitize the gland to carcinogenic agents through its constant stimulation of mammary DNA synthesis. Acknowledgement--We thank Prof. H. A. Bern, Department of Zoology and Cancer Research Laboratory, University of California, Berkeley, California, U.S.A. and Dr. T. Mori, Zoological Institute, University of Tokyo, Tokyo, Japan for reading the manuscript and for valuable suggestions.
REFERENCES 1. 2.
D. MEDINA, K. B. DEOME and L. YOUNG, Tumor-producing capabilities of hyperplastic alveolar nodules in virgin and hormone-stimulated BALB/c f. C3H and C3Hfmice. d. nat. Cancer Inst. 44, 167 (1970). T. MoRx and H. A. BERN, Personal communication.
Letter to the Editor 3. 4. 5. 6. 7. 8.
9. 10. I 1. 12.
L . M . BOOT, Induction by prolactin of mammary tumours in mice. Verh. kon. Ned. Akad. Wet., Afd. Natuurk., Tweed, Reeks, deel LVIII, Suppl. 3. (1969). G. R6PcKE, Interaction of hypophyseal isografts and ovarian hormones in mammary tumour development in mice. Thesis, Mondeel-Offsetdrukkerij, Amsterdam (1975). H. NAOASAWAand R. YANAI, Frequency of mammary cell division in relation t o age: Its significance in the induction of mammary tumors by carcinogen in rats. J. nat. Cancer Inst. 52, 609 (1974). H. NAOASAWA,R. YANAI and H. TANmUCHI, Importance of mammary gland DNA synthesis in carcinogen-induced mammary tumorigenesis in rats. Cancer Res. 36, 2223 (1976). ]?,. YANAI and H. NAOASAWA,Effects of pituitary graft and 2-bromo-a-ergocryptine on mammary DNA synthesis in mice in relation to mammary tumorigenesis. J. nat. Cancer Inst. 56, 1055 (1976). H. NAOASAWA, R. YANAI, H. TANIOUCHI, R. TOKUZEN and the late W. NAXAI-IAaA, Two-way selection of a stock of Swiss albino mice for mammary tumorigenesis: Establishment of two new strains (SHN and SLN). J. nat. Cancer Inst. 57, In press (1976). H. NAOASAWAand R. YANAI, Effects of estrogen and/or pituitary graft on nucleic acid synthesis of carcinogen-induced mammary tumors in rats. d. nat. Cancer Inst. 52, 1219 (1974). G . W . SNEDECOR,Statistical Methods. 5th ed., p. 194. Iowa State Univ. Press., Ames. H. NAOASAWAand R. YANk, Unpublished. H . A . B~RN, Nature of the hormonal influence in mouse mammary cancer. Science 131~ 1039 (1960).
1019
