Comparative growth of normal and malignant mouse mammary epithelium cultured serum-free on a biomatrix from preadipocytes

Abstract

Growth of normal and malignant mouse mammary epithelial cells (MMEC) on a biomatrix of substrate-attached material from 3T3-L1 preadipocytes was evaluated to devise culture conditions that are suitable for transformation studies but do not involve embedding cells in a gel. The biomatrix was prepared as described by Levine and Stockdale, and serum-free medium contained bovine serum albumin, insulin, progesterone, prolactin, and linoleic acid. Each cell type produced a distinctive pattern of colony architecture in this culture system. Cells from virgin mice (vMMEC) usually formed elaborate, three-dimensional structures resembling ducts and alveoli; cells from pregnant mice (pMMEC) grew as flat monolayers; and tumor cells grew in multilayered clusters. Cell growth was monitored by an assay for succinate dehydrogenase. Similar growth rates were found through Day 8 in cultures of vMMEC and D2 carcinoma cells. Growth of vMMEC slowed thereafter, whereas tumor cells typically continued growing through Day 14 to 18. Increase in cell number during 18 days in culture was 3-, 7-, 9-, and 11-fold for cells from pregnant and virgin mice, BALB/cfC3H and D2 carcinomas, respectively. The percent cells in S phase on Day 2 of culture was 9% for pMMEC, 4 to 11% for BALB/cfC3H tumor cells, 20% for vMMEC, and 24% for D2 tumor cells. Thus, this culture system promotes extended growth of MMEC and offers several advantages over embedding cells in a collagen gel. It may therefore be applicable to in vitro transformation studies with MMEC.