Chemical–Physical Changes in Cell Membrane Microdomains of Breast Cancer Cells After Omega-3 PUFA Incorporation


Epidemiologic and experimental studies suggest that dietary fatty acids influence the development and progression of breast cancer. However, no clear data are present in literature that could demonstrate how n − 3 PUFA can interfere with breast cancer growth. It is suggested that these fatty acids might change the structure of cell membrane, especially of lipid rafts. During this study we treated MCF-7 and MDA-MB-231 cells with AA, EPA, and DHA to assess if they are incorporated in lipid raft phospholipids and are able to change chemical and physical properties of these structures. Our data demonstrate that PUFA and their metabolites are inserted with different yield in cell membrane microdomains and are able to alter fatty acid composition without decreasing the total percentage of saturated fatty acids that characterize these structures. In particular in MDA-MB-231 cells, that displays the highest content of Chol and saturated fatty acids, we observed the lowest incorporation of DHA, probably for sterical reasons; nevertheless DHA was able to decrease Chol and SM content. Moreover, PUFA are incorporated in breast cancer lipid rafts with different specificity for the phospholipid moiety, in particular PUFA are incorporated in PI, PS, and PC phospholipids that may be relevant to the formation of PUFA metabolites (prostaglandins, prostacyclins, leukotrienes, resolvines, and protectines) of phospholipids deriving second messengers and signal transduction activation. The bio-physical changes after n − 3 PUFA incubation have also been highlighted by atomic force microscopy. In particular, for both cell lines the DHA treatment produced a decrease of the lipid rafts in the order of about 20–30 %. It is worth noticing that after DHA incorporation lipid rafts exhibit two different height ranges. In fact, some lipid rafts have a higher height of 6–6.5 nm. In conclusion n − 3 PUFA are able to modify lipid raft biochemical and biophysical features leading to decrease of breast cancer cell proliferation probably through different mechanisms related to acyl chain length and unsaturation. While EPA may contribute to cell apoptosis mainly through decrease of AA concentration in lipid raft phospholipids, DHA may change the biophysical properties of lipid rafts decreasing the content of cholesterol and probably the distribution of key proteins.

This is a preview of subscription content, access via your institution.

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6





Fatty acids


Polyunsaturated fatty acids


Monounsaturated fatty acids


Saturated fatty acids


Docosahexaenoic acid


Eicosapentaenoic acid


Arachidonic acid














  1. 1.

    Bagga, D., Anders, K. H., Wang, H. J., & Glaspy, J. A. (2002). Long-chain n − 3-to-n − 6 polyunsaturated fatty acid ratios in breast adipose tissue from women with and without breast cancer. Nutrition and Cancer, 42(2), 180–185.

    PubMed  CAS  Article  Google Scholar 

  2. 2.

    Manni, A., Xu, H., Washington, S., Aliaga, C., Cooper, T., Richie, J. P, Jr., et al. (2010). The impact of fish oil on the chemopreventive efficacy of tamoxifen against development of N-methyl-N-nitrosourea-induced rat mammary carcinogenesis. Cancer Prevention Research (Phila), 3(3), 322–330.

    CAS  Article  Google Scholar 

  3. 3.

    Gillet, L., Roger, S., Bougnoux, P., Le Guennec, J. Y., & Besson, P. (2011). Beneficial effects of omega-3 long-chain fatty acids in breast cancer and cardiovascular diseases: Voltage-gated sodium channels as a common feature? Biochimie, 93(1), 4–6.

    PubMed  CAS  Article  Google Scholar 

  4. 4.

    Olbrich, K., Rawicz, W., Needham, D., & Evans, E. (2000). Water permeability and mechanical strength of polyunsaturated lipid bilayers. Biophysical Journal, 79(1), 321–327.

    PubMed  CAS  Article  Google Scholar 

  5. 5.

    Rawicz, W., Olbrich, K. C., McIntosh, T., Needham, D., & Evans, E. (2000). Effect of chain length and unsaturation on elasticity of lipid bilayers. Biophysical Journal, 79(1), 328–339.

    PubMed  CAS  Article  Google Scholar 

  6. 6.

    Brzustowicz, M. R., Cherezov, V., Zerouga, M., Caffrey, M., & Stillwell, W. (2002). Controlling membrane cholesterol content. A role for polyunsaturated (docosahexaenoate) phospholipids. Biochemistry, 41(41), 12509–12519.

    PubMed  CAS  Article  Google Scholar 

  7. 7.

    Botelho, A. V., Gibson, N. J., Thurmond, R. L., Wang, Y., & Brown, M. F. (2002). Conformational energetics of rhodopsin modulated by nonlamellar-forming lipids. Biochemistry, 41(20), 6354–6368.

    PubMed  CAS  Article  Google Scholar 

  8. 8.

    Mitchell, D. C., Niu, S. L., & Litman, B. J. (2003). Enhancement of G protein-coupled signaling by DHA phospholipids. Lipids, 38(4), 437–443.

    PubMed  CAS  Article  Google Scholar 

  9. 9.

    London, E., & Brown, D. A. (2000). Insolubility of lipids in triton X-100: Physical origin and relationship to sphingolipid/cholesterol membrane domains (rafts). Biochimica et Biophysica Acta, 1508(1–2), 182–195.

    PubMed  CAS  Article  Google Scholar 

  10. 10.

    Pike, L. J. (2004). Lipid rafts: Heterogeneity on the high seas. Biochemical Journal, 378(Pt 2), 281–292.

    Google Scholar 

  11. 11.

    Lingwood, D., & Simons, K. (2007). Detergent resistance as a tool in membrane research. Nature Protocols, 2(9), 2159–2165.

    PubMed  CAS  Article  Google Scholar 

  12. 12.

    Simons, K., & Toomre, D. (2001). Lipid rafts and signal transduction. Nature Reviews Molecular Cell Biology, 1(1), 31–39. Erratum in: Nature Reviews Molecular Cell Biology, 2(3), 216.

    Google Scholar 

  13. 13.

    Razani, B., Woodman, S. E., & Lisanti, M. P. (2002). Caveolae: From cell biology to animal physiology. Pharmacological Reviews, 54(3), 431–467.

    PubMed  CAS  Article  Google Scholar 

  14. 14.

    van Meer, G. (2002). Cell biology. The different hues of lipid rafts. Science, 296(5569), 855–857.

    Google Scholar 

  15. 15.

    Fastenberg, M. E., Shogomori, H., Xu, X., Brown, D. A., & London, E. (2003). Exclusion of a transmembrane-type peptide from ordered-lipid domains (rafts) detected by fluorescence quenching: Extension of quenching analysis to account for the effects of domain size and domain boundaries. Biochemistry, 42(42), 12376–12390.

    PubMed  CAS  Article  Google Scholar 

  16. 16.

    Gimpl, G., Burger, K., & Fahrenholz, F. (1997). Cholesterol as modulator of receptor function. Biochemistry, 36(36), 10959–10974.

    PubMed  CAS  Article  Google Scholar 

  17. 17.

    Pike, L. J., & Casey, L. (2002). Cholesterol levels modulate EGF receptor-mediated signaling by altering receptor function and trafficking. Biochemistry, 41(32), 10315–10322.

    PubMed  CAS  Article  Google Scholar 

  18. 18.

    Hanzal-Bayer, M. F., & Hancock, J. F. (2007). Lipid rafts and membrane traffic. FEBS Letters, 581(11), 2098–2104.

    PubMed  CAS  Article  Google Scholar 

  19. 19.

    Jury, E. C., Flores-Borja, F., & Kabouridis, P. S. (2007). Lipid rafts in T cell signalling and disease. Seminars in Cell & Developmental Biology, 18(5), 608–615.

    CAS  Article  Google Scholar 

  20. 20.

    Corsetto, P. A., Montorfano, G., Zava, S., Jovenitti, I. E., Cremona, A., Berra, B., et al. (2011). Effects of n − 3 PUFAs on breast cancer cells through their incorporation in plasma membrane. Lipids in Health and Disease, 10, 73.

    PubMed  CAS  Article  Google Scholar 

  21. 21.

    Fabelo, N., Martin, V., Santpere, G., Marìn, R., Torrent, L., Ferrer, I., et al. (2011). Severe alterations in lipid composition of frontal cortex lipid rafts from Parkinson’s disease and incidental Parkinson’s disease. Molecular Medicine, 17(9–10), 1107–1118.

    PubMed  CAS  Google Scholar 

  22. 22.

    Lowry, O. H., Rosebrough, N. J., Farr, A. L., & Randall, R. J. (1951). Protein measurement with the Folin phenol reagent. Journal of Biological Chemistry, 193(1), 265–275.

    PubMed  CAS  Google Scholar 

  23. 23.

    Jho, D. H., Cole, S. M., Lee, E. M., & Espat, N. J. (2004). Role of omega-3 fatty acid supplementation in inflammation and malignancy. Integrative Cancer Therapies, 3(2), 98–111.

    PubMed  CAS  Article  Google Scholar 

  24. 24.

    Innis, S. M., & Jacobson, K. (2007). Dietary lipids in early development and intestinal inflammatory disease. Nutrition Reviews, 65(12 Pt 2), S188–S193.

    PubMed  Article  Google Scholar 

  25. 25.

    Calder, P. C. (2006). n − 3 polyunsaturated fatty acids, inflammation, and inflammatory diseases. American Journal of Clinical Nutrition, 83(6 Suppl), 1505S–1519S.

    Google Scholar 

  26. 26.

    Menendez, J. A., Lupu, R., & Colomer, R. (2005). Exogenous supplementation with omega-3 polyunsaturated fatty acid docosahexaenoic acid (DHA; 22: 6n − 3) synergistically enhances taxane cytotoxicity and downregulates Her-2/neu (c-erbB-2) oncogene expression in human breast cancer cells. European Journal of Cancer Prevention, 14, 263–270.

    PubMed  CAS  Article  Google Scholar 

  27. 27.

    Calviello, G., Di, N. F., Serini, S., Piccioni, E., Boninsegna, A., & Maggiano, N. (2005). Docosahexaenoic acid enhances the susceptibility of human colorectal cancer cells to 5-fluorouracil. Cancer Chemotherapy and Pharmacology, 55, 12–20.

    PubMed  CAS  Article  Google Scholar 

  28. 28.

    Lindskog, M., Gleissman, H., Ponthan, F., Castro, J., Kogner, P., & Johnsen, J. I. (2006). Neuroblastoma cell death in response to docosahexaenoic acid: Sensitization to chemotherapy and arsenic-induced oxidative stress. International Journal of Cancer, 118, 2584–2593.

    CAS  Article  Google Scholar 

  29. 29.

    Edidin, M. (2001). Membrane cholesterol, protein phosphorylation, and lipid rafts. Science’s STKE, 2001(67), E1.

    Google Scholar 

  30. 30.

    Zajchowski, L. D. (2002). Lipid rafts and little caves. Compartmentalized signalling in membrane microdomains. European Journal of Biochemistry, 269, 737–752.

    PubMed  CAS  Article  Google Scholar 

  31. 31.

    Epand, R. M. (2008). Proteins and cholesterol-rich domains. Biochimica et Biophysica Acta, 1778(7–8), 1576–1582.

    Google Scholar 

  32. 32.

    Maxfield, F., & Tabas, I. (2005). Role of cholesterol and lipid organization in disease. Nature, 438, 612–621.

    PubMed  CAS  Article  Google Scholar 

  33. 33.

    Silvius, J. R. (2003). Role of cholesterol in lipid raft formation: Lessons from lipid model systems. Biochimica et Biophysica Acta, 2003(1610), 174–183.

    Article  Google Scholar 

  34. 34.

    Rizzo, A. M., Montorfano, G., Negroni, M., Adorni, L., Berselli, P., Corsetto, P., et al. (2010). A rapid method for determining arachidonic: Eicosapentaenoic acid ratios in whole blood lipids: Correlation with erythrocyte membrane ratios and validation in a large Italian population of various ages and pathologies. Lipids in Health and Disease, 27, 7–9.

    Article  Google Scholar 

  35. 35.

    Welch, A. A., Shakya-Shrestha, S., Lentjes, M. A., Wareham, N. J., & Khaw, K. T. (2010). Dietary intake and status of n − 3 polyunsaturated fatty acids in a population of fish-eating and non-fish-eating meat-eaters, vegetarians, and vegans and the product-precursor ratio [corrected] of α-linolenic acid to long-chain n − 3 polyunsaturated fatty acids: Results from the EPIC-Norfolk cohort. American Journal of Clinical Nutrition, 92(5), 1040–1051.

    PubMed  CAS  Article  Google Scholar 

  36. 36.

    Rizzo, A. M., Montorfano, G., Fallini, M., Negroni, M., Paleari, D., Berra, B., et al. (2006). A case-control study in cancer patients (PTS): The arachidonic acid/eicosapentaenoic acid (AA/EPA) ratio as a biomarker. Annnals of Oncology, 2006(17), 306.

    Google Scholar 

  37. 37.

    Hrelia, S., Bordoni, A., Biagi, P., Rossi, C. A., Bernardi, L., Horrobin, D. F., et al. (1996). gamma-Linolenic acid supplementation can affect cancer cell proliferation via modification of fatty acid composition. Biochemical and Biophysical Research Communications, 1996(225), 441–447.

    Article  Google Scholar 

  38. 38.

    Pala, V., Krogh, V., Muti, P., Chajès, V., Riboli, E., Micheli, A., et al. (2001) Erythrocyte membrane fatty acids and subsequent breast cancer: A prospective Italian study. Journal of the National Cancer Institute, 93(14), 1088–1095.

    Google Scholar 

  39. 39.

    Mason, P., Liang, B., Li, L., Fremgen, T., Murphy, E., Quinn, A., et al. (2012). SCD1 inhibition causes cancer cell death by depleting mono-unsaturated fatty acids. PLoS ONE, 7(3), e33823.

    PubMed  CAS  Article  Google Scholar 

  40. 40.

    Yang, Z. H., Miyahara, H., Takemura, S., & Hatanaka, A. (2011). Dietary saury oil reduces hyperglycemia and hyperlipidemia in diabetic KKAy mice and in diet-induced obese C57BL/6J mice by altering gene expression. Lipids, 46(5), 425–434.

    PubMed  CAS  Article  Google Scholar 

  41. 41.

    Muhlhausler, B. S., Cook-Johnson, R., James, M., Miljkovic, D., Duthoit, E., & Gibson, R. (2010). Opposing effects of omega-3 and omega-6 long chain polyunsaturated Fatty acids on the expression of lipogenic genes in omental and retroperitoneal adipose depots in the rat. Journal of Nutrition and Metabolism, Article ID 927836, pp. 9. doi:10.1155/2010/927836.

  42. 42.

    Tonnetti, L., Verí, M. C., Bonvini, E., & D’Adamio, L. (1999). A role for neutral sphingomyelinase-mediated ceramide production in T cell receptor-induced apoptosis and mitogen-activated protein kinase-mediated signal transduction. Journal of Experimental Medicine, 189(10), 1581–1589.

    PubMed  CAS  Article  Google Scholar 

  43. 43.

    Jayadev, S., Liu, B., Bielawska, A. E., Lee, J. Y., Nazaire, F., Pushkareva, M Yu., et al. (1995). Role for ceramide in cell cycle arrest. Journal of Biological Chemistry, 270(5), 2047–2052.

    PubMed  CAS  Article  Google Scholar 

  44. 44.

    Veldman, R. J., Maestre, N., Aduib, O. M., Medin, J. A., Salvayre, R., & Levade, T. (2001). A neutral sphingomyelinase resides in sphingolipid-enriched microdomains and is inhibited by the caveolin-scaffolding domain: Potential implications in tumour necrosis factor signaling. Biochemical Journal, 355(Pt 3), 859–868.

    PubMed  CAS  Google Scholar 

  45. 45.

    Wu, M., Harvey, K. A., Ruzmetov, N., Welch, Z. R., Sech, L., Jackson, K., et al. (2005). Omega-3 polyunsaturated fatty acids attenuate breast cancer growth through activation of a neutral sphingomyelinase-mediated pathway. International Journal of Cancer, 117(3), 340–348.

    CAS  Article  Google Scholar 

  46. 46.

    Wang, T. Y., & Silvius, J. R. (2001). Cholesterol does not induce segregation of liquid-ordered domains in bilayers modeling the inner leaflet of the plasma membrane. Biophysical Journal, 81(5), 2762–2773.

    PubMed  CAS  Article  Google Scholar 

  47. 47.

    Barman, S., & Nayak, D. P. (2007). Lipid raft disruption by cholesterol depletion enhances influenza. Journal of Virology, 81, 12169–12178.

    Google Scholar 

  48. 48.

    Freeman, M. R., & Solomon, K. R. (2004). Cholesterol and prostate cancer. Journal of Cellular Biochemistry, 91, 54–69.

    PubMed  CAS  Article  Google Scholar 

  49. 49.

    Kolanjiappan, K., Ramachandran, C. R., & Manoharan, S. (2003). Biochemical changes in tumor tissues of oral cancer patients. Clinical Biochemistry, 36, 61–65.

    PubMed  CAS  Article  Google Scholar 

  50. 50.

    Bennis, F., Favre, G., Le, G. F., & Soula, G. (1993). Importance of mevalonate-derived products in the control of HMG-CoA reductase activity and growth of human lung adenocarcinoma cell line A549. International Journal of Cancer, 55, 640–645.

    CAS  Article  Google Scholar 

  51. 51.

    El-Sohemy, A., & Archer, M. C. (2000). Inhibition of N-methyl-N-nitrosourea-and 7,12-dimethylbenz[a] anthracene-induced rat mammary tumorigenesis by dietary cholesterol is independent of Ha-Ras mutations. Carcinogenesis, 21, 827–831.

    PubMed  CAS  Article  Google Scholar 

  52. 52.

    Li, Y. C., Park, M. J., Ye, S. K., Kim, C. W., & Kim, Y. N. (2006). Elevated levels of cholesterol-rich lipid rafts in cancer cells are correlated with apoptosis sensitivity induced by cholesterol-depleting agents. American Journal of Pathology, 168, 1107–1118.

    PubMed  CAS  Article  Google Scholar 

  53. 53.

    Huster, D., Arnold, K., & Gawrisch, K. (1998). Influence of docosahexaenoic acid and cholesterol on lateral lipid organization in phospholipid mixtures. Biochemistry, 37, 17299–17308.

    PubMed  CAS  Article  Google Scholar 

  54. 54.

    Mitchell, D. C., & Litman, B. J. (1998). Effect of cholesterol on molecular order and dynamics in highly polyunsaturated phospholipid bilayers. Biophysical Journal, 75, 896–908.

    PubMed  CAS  Article  Google Scholar 

  55. 55.

    Niu, S. L., & Litman, B. J. (2002). Determination of membrane cholesterol partition coefficient using a lipid vesicle cyclodextrin binary system: Effect of phospholipid acyl chain unsaturation and headgroup composition. Biophysical Journal, 83, 3408–3415.

    PubMed  CAS  Article  Google Scholar 

  56. 56.

    Zerouga, M., Jenski, L. J., & Stillwell, W. (1995). Comparison of phosphatidylcholines containing one or two docosahexaenoic acyl chains on properties of phospholipid monolayers and bilayers. Biochimica et Biophysica Acta, 1236, 266–272.

    PubMed  Article  Google Scholar 

  57. 57.

    Kariel, N., Davidson, E., & Keough, K. M. (1991). Cholesterol does not remove the gel-liquid crystalline phase transition of phosphatidylcholines containing two polyenoic acyl chains. Biochimica et Biophysica Acta, 1062, 70–76.

    PubMed  CAS  Article  Google Scholar 

  58. 58.

    Needham, D., & Nunn, R. S. (1990). Elastic deformation and failure of lipid bilayer membranes containing cholesterol. Biophysical Journal, 58, 997–1009.

    PubMed  CAS  Article  Google Scholar 

  59. 59.

    Mitchell, D. C., & Litman, B. J. (1998). Molecular order and dynamics in bilayers consisting of highly polyunsaturated phospholipids. Biophysical Journal, 74, 879–891.

    PubMed  CAS  Article  Google Scholar 

  60. 60.

    Brzustowicz, M. R., Cherezov, V., Caffrey, M., Stillwell, W., & Wassall, S. R. (2002). Molecular organization of cholesterol in polyunsaturated membranes: Microdomain formation. Biophysical Journal, 82, 285–298.

    PubMed  CAS  Article  Google Scholar 

  61. 61.

    Brzustowicz, M. R., Cherezov, V., Zerouga, M., Caffrey, M., & Stillwell, W. (2002). Controlling membrane cholesterol content. A role for polyunsaturated (docosahexaenoate) phospholipids. Biochemistry, 41, 12509–12519.

    PubMed  CAS  Article  Google Scholar 

  62. 62.

    Pasenkiewicz-Gierula, M., Subczynski, W. K., & Kusumi, A. (1990). Rotational diffusion of a steroid molecule in phosphatidylcholine-cholesterol membranes: Fluid-phase microimmiscibility in unsaturated phosphatidylcholine-cholesterol membranes. Biochemistry, 29, 4059–4069.

    PubMed  CAS  Article  Google Scholar 

  63. 63.

    Pike, L. J., Han, X., Chung, K. N., & Gross, R. W. (2002). Lipid rafts are enriched in arachidonic acid and plasmenylethanolamine and their composition is independent of caveolin-1 expression: A quantitative electrospray ionization/mass spectrometric analysis. Biochemistry, 41(6), 2075–2088.

    PubMed  CAS  Article  Google Scholar 

  64. 64.

    Eldho, N. V., Feller, S. E., Tristram-Nagle, S., Polozov, I. V., & Gawrisch, K. (2003). Polyunsaturated docosahexaenoic vs docosapentaenoic acid-differences in lipid matrix properties from the loss of one double bond. Journal of the American Chemical Society, 125(21), 6409–6421.

    PubMed  CAS  Article  Google Scholar 

  65. 65.

    Shaikh, S. R. (2012). Biophysical and biochemical mechanisms by which dietary N − 3 polyunsaturated fatty acids from fish oil disrupt membrane lipid rafts. Journal of Nutritional Biochemistry, 23(2), 101–105.

    PubMed  CAS  Article  Google Scholar 

  66. 66.

    DeGraffenried, L. A., Friedrichs, W. E., Fulcher, L., Fernandes, G., Silva, J. M., Peralba, J. M., et al. (2003). Eicosapentaenoic acid restores tamoxifen sensitivity in breast cancer cells with high Akt activity. Annals of Oncology, 2003(14), 1051–1056.

    Article  Google Scholar 

  67. 67.

    Truan, J. S., Chen, J. M., & Thompson, L. U. (2010). Flaxseed oil reduces the growth of human breast tumors (MCF-7) at high levels of circulating estrogen. Molecular Nutrition & Food Research, 2010(54), 1414–1421.

    Article  Google Scholar 

  68. 68.

    Engelke, M., Tykhonova, S., Zorn-Kruppa, M., & Diehl, H. (2002). Tamoxifen induces changes in the lipid composition of the retinal pigment epithelium cell line D407. Pharmacology and Toxicology, 2002(91), 13–21.

    Google Scholar 

  69. 69.

    Hou, T. Y., Monk, J. M., Fan, Y. Y., Barhoumi, R., Chen, Y. Q., Rivera, G. M., et al. (2012). N − 3 polyunsaturated fatty acids suppress phosphatidylinositol 4,5-bisphosphate-dependent actin remodelling during CD4+ T-cell activation. Biochemical Journal, 2012(443), 27–37.

    Article  Google Scholar 

Download references


We are grateful to anonymous reviewers for their constructive suggestions. Financial support to Dr Angela M. Rizzo came from Italian Space Agency (ASI).

Conflict of interest


Author information



Corresponding author

Correspondence to Angela M. Rizzo.

Additional information

This paper is dedicated to the memory of Prof. Bruno Berra.

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Corsetto, P.A., Cremona, A., Montorfano, G. et al. Chemical–Physical Changes in Cell Membrane Microdomains of Breast Cancer Cells After Omega-3 PUFA Incorporation. Cell Biochem Biophys 64, 45–59 (2012).

Download citation


  • EPA
  • DHA
  • AA
  • PUFA
  • Breast cancer
  • Lipid rafts
  • Phospholipids
  • Membrane