FATTY-ACID MODULATION OF EPIDERMAL GROWTH FACTOR-INDUCED MOUSE MAMMARY EPITHELIAL-CELL PROLIFERATION IN-VITRO

Mammary epithelial cells were isolated from mid-pregnant BALB/c mice, grown in primary culture within collagen gels, and maintained with ser um-free medium containing 10 ng/ml epidermal growth factor (EGF) as th e mitogen. Supplementation of culture medium with the saturated fatty acid, Na-stearate (18:0), significantly attenuated, whereas treatment with the unsaturated fatty acid, Na-arachidonate (20:4), significantly enhanced mammary epithelial cell proliferation, as compared to untrea ted controls. Treatment with various doses of either 18:0 or 20:4 was also found to result in a direct dose-dependent enrichment of mammary epithelial cell membrane fatty acid composition and a concurrent decre ase in the relative levels of other membrane fatty acids, as determine d by gas chromatography. Administration of the prostaglandin synthesis inhibitor, indomethacin, significantly inhibited EGF-induced cell gro wth in all treatment groups, but did not alter the relative inhibitory (18:0) or stimulatory (20:4) effects of fatty acid treatment. EGF-ind uced PRC translocation into the membrane fraction of mammary epithelia l cells was enhanced in 20:4 and attenuated in 18:0 treatment groups, as compared to controls. Western blot analysis of phospholipid-depende nt protein kinase C isoenzymes showed that PKC, was the predominant is oenzyme present in mouse mammary epithelial cells grown in primary cul ture, and the molecular weight of this PKC isoenzyme was determined to be 85 kDa. These results suggest that supplementation of culture medi a with specific fatty acids is associated with significant alterations in mammary epithelial cell membrane fatty acid composition, PKC activ ation, and mitogenic responsiveness. Since EGF can induce both PRC act ivation and cell proliferation, and because PKC activation requires me mbrane-derived phospholipids and diacylglycerol, these data suggest th at specific fatty acid modulation of mammary epithelial cell mitogenes is is mediated through alterations in PRC, activation. (C) 1994 Academ ic Press, Inc.