Binding of 7,12-Dimethylbenz[a]anthracene to BALB/c Mouse Mammary Gland DNA in Organ Culture

The second thoracic mammary glands of immature female BALB/c mice were cultivated to alveolar development in a chemically defined medium with insulin plus prolactin plus aldosterone plus hydrocortisone (IPAH medium). The glands were treated with labeled (3H or 14C) or unlabeled 7,12-dimethylbenz[a]anthracene (DMBA) in dimethyl sulfoxide (DMSO) for 24 hours between days 3 and 4 of culture. For binding studies, isolated mammary gland DNA was treated with RNase and pronase and the radioactivity in DNA was measured. The DNA of glands treated with [14C]DMBA was also analyzed in a CsCl density gradient. The DNA of glands treated with labeled DMBA was found to contain bound radioactivity. UV absorption spectral analysis also showed the presence of the DMBA nucleus at the 310-nm region in the DNA of DMBA-treated glands. Nearly 100% of the initial [3H]DMBA-DNA radioactivity was retained up to 5 months, and at 8 months 72% of the radioactivity was present in the DNA. For morphologic studies, observation of the glands after treatment with equimolar concentrations (0.004 m) of DMBA, dibenz[a,h]anthracene, benz[a]anthracene, or anthracene in IPAH medium revealed that DMBA was most potent in inducing nodulelike alveolar lesions (NLAL) in the mammary glands in culture. Thus the binding of DMBA to DNA corresponds to its high noduligenic action in the mammary glands in the present organ culture model. The results appear to resemble DMBA-DNA interaction associated with the carcinogenic action of DMBA. The biologic characteristics of the NLAL are being assessed by mammary fat pad transplantation.