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First published online 2 March 2004
doi: 10.1242/jcs.01000

Journal of Cell Science 117, 1495-1502 (2004)
Published by The Company of Biologists 2004
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The stroma as a crucial target in rat mammary gland carcinogenesis

Maricel V. Maffini1, Ana M. Soto1,*, Janine M. Calabro1, Angelo A. Ucci2 and Carlos Sonnenschein1

1 Department of Anatomy and Cellular Biology, Tufts University School of Medicine, 136 Harrison Avenue, Boston, MA 02111, USA
2 Department of Pathology, Tufts-New England Medical Center, 750 Washington Street, Boston, MA 02111, USA



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  		Fig. 1. (A) Whole-mount preparation of an intact mammary gland from a 21-day-old rat showing the ductal tree and lymph nodes. (B) Mammary gland fat pad cleared of epithelium at 21 days of age and excised at the end of the experiment, 11 months later. Bar, 4 mm.

 


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  		Fig. 2. Repopulation of the mammary gland. (A) Mammary epithelial cells grown in culture showing the expression of cytokeratin (red) and vimentin (green). Mammary epithelial cells averaged 90% of the total cell population transplanted into cleared fat pads. Counterstaining, Hoescht 33258 (blue). Mammary epithelial cells were injected into cleared fat pads and the recombinants were harvested at 0 (B), 30 (C), 60 (D) and 90 (E) days after cell injection. Bars, 20 µm (A); 2 mm (B-E).

 


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  		Fig. 3. Neoplasms developed in NMU-treated stroma only. (A) Mammary gland whole-mount preparations show abnormal outgrowths in animals whose cleared fat pads were exposed to NMU prior to recombination with vehicle-treated mammary epithelial cells (Group 1) or NMU-treated mammary epithelial cells (Group 2). Neoplastic lesions (arrows) were confirmed histologically. Groups 3 and 4 developed normal-like ductal outgrowths. Bar, 2 mm. (B) Percentage of neoplastic lesions and incidence of carcinomas per experimental group. The number of rats with neoplastic lesions out of the total number of animals in each group is indicated in parenthesis. *See Materials and Methods for further details.

 


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  		Fig. 4. Incidence of neoplasms and latency period. (A) Number of neoplastic lesions in inguinal and thoracic mammary glands (mean±s.e.). (B) Latency of neoplastic lesions in inguinal and thoracic mammary glands expressed in weeks (mean±s.e.).

 


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  		Fig. 5. Sections of recombinant tissues. (A) Section from a recombinant of vehicle-exposed stroma and NMU-exposed mammary epithelial cells. The histoarchitecture resembles a normal mammary gland. (B) Papillary carcinoma from a recombinant of NMU-exposed stroma and vehicle-exposed mammary epithelial cells. Hematoxylin and eosin staining (A,B). (C) Immunohistochemistry for cytokeratin, vimentin and desmin in sections of the tumor shown in B. Counterstaining: Harris' hematoxylin. Bar, 100 µm.

 


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  		Fig. 6. Analysis of the presence of point mutation in the Ha-ras-1 gene using the MAMA. The mutant-specific amplification product is 74 bp whereas the normal product is 128 bp. Lanes 1, 5, 6 and 7: mammary tumors from Group 5. Tumor in lane 1 lacks Ha-ras-1 gene mutation. Lane 2: mammary tumor from Group 1. Note absence of Ha-ras-1 gene mutation. Lanes 3 and 4: mammary tumors from Group 2. Lanes 8 and 9: normal mammary tissue from intact animals taken randomly from the colony. Note: the smaller bands in lanes 1 and 2 correspond to dimers of the primer.

 

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© The Company of Biologists Ltd 2004