Biochemical and biophysical approaches to study the incorporation of omega-3 LCPUFA on breast cancer cells lipid rafts

In vivo and in vitro studies suggest that n-3 PUFAs can be cancer chemopreventive, chemosuppressive agents and auxiliary agents for cancer therapy. n-3 PUFAs could alter the growth of tumour cells by influencing cell replication, interfering with components of the cell cycle, or increasing cell death, either by inducing necrosis or apoptosis. However, the mechanism by which n-3 PUFAs inhibit breast cancer cells growth, is not yet well understood. It was suggested that these fatty acids might change the fluidity and structure of cell membrane, especially of lipid rafts. We have studied the incorporation of PUFAs in cancer cell membrane and then in lipid rafts by HPLC/GC. We have observed that EPA and DHA are incorporated in cell membrane and in lipid rafts with different specificity for the phospholipids moiety. Worth of note is the observation that only the treatment with DHA induces a reduction of cholesterol content in lipid rafts, indicating a possible change in raft organization. Changes in the lipid rafts structure were also analyzed with atomic force microscopy (AFM) both before and after n-3 PUFA treatment. The AFM analysis of breast cancer lipid rafts indicates a reduction of microdomains number, after DHA incorporation. Moreover PUFA treated rafts are dimensionally different than control rafts. Taken together, our results indicate that n-3 PUFA \u2018feeding\u2019 might induce modifications of lipid rafts structure increasing the degree of fatty acid unsaturation, and these changes of lipid rafts might modify signal transduction and cell-cell interactions. In fact our data have also demonstrated in MDA-MB-231 cells that EPA does not change the level of EGFR in MDA-MB-231 cells treated with EPA or EPA/EGF, but reduces the level of the active form (pEGFR) in cells treated with EPA/EGF. On the contrary DHA slightly decreases the EGFR expression in MDAMB- 231 cells, and especially determines the disappearance of pEGFR in cells treated with DHA and DHA/EGF