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ENU-induced in vitro neoplastic transformation of rat mammary epithelial cells



ENU-induced in vitro neoplastic transformation of rat mammary epithelial cells



Anticancer Research 11(5): 1783-1792



Normal mammary epithelial cells, originating from female Sprague-Dawley rats, were grown in Dulbecco's Modified Eagles Medium containing 25% horse serum and hormone supplements. Once established as an epithelial cell culture, the cells were treated with N-ethyl-N-nitrosourea (ENU) in various doses (25-500 ug/ml) to study the process of in vitro mammary epithelial cell neoplastic transformation. The ENU-treatment of primary mammary epithelial cell culture resulted in a sequence of phenotypic changes, termed stages I-V. The rat mammary epithelial cells, after a period of approximately 30 days post-ENU exposure, showed a marked proliferation of morphologically altered cells which formed multi-layered colonies. Subsequently, these cells acquired the capacity to form colonies in soft agar and eventually produced a high yield of palpable tumors when inoculated into newborn female isologous hosts or female athymic nude mice. The immediate effect of ENU on these cells was monitored by measurement of cellular DNA content, unscheduled DNA synthesis, cell proliferation and chromosomal aberrations. The ENU effect on cell proliferation and DNA synthesis was dose dependent; doses greater than 100 ug/ml reduced the cell number and DNA synthesis. Cytofluorometric histograms of non-ENU-treated rat mammary epithelial cells showed a near diploid population of cells. The ENU exposed cells subsequently became hyperdiploid (24-72 hours after ENU) and then regained their near diploid pattern at 120 hours after ENU exposure. The ENU-treated cells also showed a second peak of cells with DNA content in the tetraploid and octaploid range at 24-72 hours after ENU exposure. Single chromatid breaks, isochromatid breaks, chromosomal exchanges, multiple chromosomal breaks and double minutes were among the chromosomal aberrations seen in ENU-treated cells. Most of the chromosomal aberrations peaked at 6 hours post-ENU exposure. The ENU-induced model of in vitro meplastic transformation of rat mammary epithelium as described in this communication appears to provide a good model for the systematic study of the early critical cellular events prerequisite to this carcinogenic process.

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Accession: 039886647

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PMID: 1837443



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