Hormone-Induced Protection of Mammary Tumorigenesisin Genetically Engineered Mouse Models - Genetic Engineering: Recent Developments in Applications - page 271

was provided at 2 to 4 weeks after transplantation. Groups 3 and 4 contained
mice that received the hormones for 2 weeks in one group (the other group was
untreated age-matched controls), followed by a rest period of 4 weeks, followed
by transplantation of the duct fragments into the cleared fat pads of 11-week-old
mice. The mice were palpated weekly until 60 weeks after transplantation.
Whole-mount preparations were made from four mammary glands from each
of the groups in experiments 1 to 3 at 4 weeks after the removal of the Silastic
tubing implants. In all experiments there was no significant difference between
the treated and untreated outgrowths in the percentage of mammary fat pad filled
by the implants. All palpable tumors were fixed in 4% paraformaldehyde for 2
to 4 hours, embedded in paraffin and processed for staining with hematoxylin/
eosin. As the transplanted mammary epithelium develops primarily mammary
adenocarcinomas but also hemangiosarcomas (at 10% incidence, presumably be-
cause of endothelium that is also transplanted as part of the duct fragment), we
assessed all tumors histologically to confirm the cell type of origin. In the text
throughout, tumor refers to mammary adenocarcinoma. Mammary adenocar-
cinoma incidences were evaluated statistically with Fisher's exact test. The BrdU
immunohistochemistry was performed as described [11].
Activated MMTV-Neu Model
The design of the experiments using the MMTV-neu model was slightly different.
FVB transgenic mice were treated for 3 weeks, starting at 7 weeks of age, with 100
µ g of estradiol in Silastic capsules. Another set of transgenic animals were treated
with 100 µ g of estradiol and 15 mg of progesterone, also in Silastic capsules. The
doses of estradiol used result in pregnancy levels of estradiol in the circulation
[19]. The control animals received empty Silastic capsules for the same duration.
Mice were palpated once every week for 8 months to monitor for mammary can-
cer development. Histopathological examination was performed to confirm the
carcinomatous nature of the palpable tumors. Mammary cancer incidence was
evaluated statistically with the χ 2 test.
Hormone Preparations
All mice received either empty Silastic tubing or the hormones in Silastic tub-
ing. For both the p53-null model and the neu model, the tubings were prepared
with the same protocol. The hormones were packed in individual Silastic capsules
(Dow Corning; size 0.078 inch (2 mm) internal diameter, 0.125 inch (3.2 mm)
outside diameter, 2 cm in length). Estradiol-17 β (Sigma, St Louis, MO, USA)
was packed in the Silastic capsules in a cellulose matrix. Progesterone (15 mg;