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Eicosapentaenoic acid promotes apoptosis in Ramos cells via activation of caspase-3 and -9

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Lipids

Abstract

Eicosapentaenoic acid (EPA; 20∶5n−3) may reduce the cell number in cultured leukemia/lymphoma cells owing to reduced cell proliferation, induction of cell death, or a combination of these processes. EPA has been shown to promote apoptosis in Ramos cells, and our present study was focused on a possible cell cycle arrest and the pathways by which the apoptotic process is induced. Apoptosis may proceed along the intrinsic (mitochondrial) or the extrinsic (death receptor) pathway, which are mediated via different caspases. Caspases are a class of homologous cysteine proteases recognized as pivotal mediators of apoptosis. We investigated whether EPA affects progression of the cell cycle or promotes apoptosis directly. By incorporation of [3H]thymidine and [3H]valine, we showed that DNA, as well as protein synthesis, was reduced after incubation of Ramos cells with EPA for 6h. We monitored cell cycle distribution by 5-bromo-2′-deoxyuridine staining and observed no cell cycle arrest in the EPA-incubated cells. Incubation of cells with EPA caused PS-flipping, as demonstrated by annexin V-binding (flow cytometry), and cleavage of poly(ADP-ribose) polymerase measured by Western blot analysis. Furthermore, we observed increased activity of caspase-3 and-9, but not of caspase-8. Whereas inhibitors of caspase-3 and-9 reduced EPA-induced apoptosis, inhibition of caspase-8 did not. This suggests that EPA may promote apoptosis via the intrinsic pathway in Ramos cells. Thus, the reduction in cell number can be explained by a direct apoptotic effect of EPA rather than via cell cycle arrest.

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Abbreviations

AnV:

annexin V

BrdU:

5-bromo-2′-deoxyuridine

FACS:

fluorescence-activated cell sorter

FCS:

fetal calf serum

FITC:

fluorescein isothiocyanate

HO342:

Hoechst 33342

HRP:

horseradish peroxidase

PARP:

poly(ADP-ribose) polymerase

PI:

propidium iodide

References

  1. Albino, A.P., Juan, G., Traganos, F., Reinhart, L., Connolly, J., Rose, D.P., and Darzynkiewicz, Z. (2000) Cell Cycle Arrest and Apoptosis of Melanoma Cells by Docosahexaenoic Acid: Association with Decreased pRb Phosphorylation, Cancer Res. 60, 4139–4145.

    PubMed  CAS  Google Scholar 

  2. Caygill, C.P., Charlett, A., and Hill, M.J. (1996) Fat, Fish, Fish Oil and Cancer, Br. J. Cancer 74, 159–164.

    PubMed  CAS  Google Scholar 

  3. Finstad, H.S., Myhrstad, M.C., Heimli, H., Lomo, J., Blomhoff, H.K., Kolset, S.O., and Drevon, C.A. (1998) Multiplication and Death-type of Leukemia Cell Lines Exposed to Very Long-Chain Polyunsaturated Fatty Acids, Leukemia 12, 921–929.

    Article  PubMed  CAS  Google Scholar 

  4. Kaizer, L., Boyd, N.F., Kriukov, V., and Tritchler, D. (1989) Fish Consumption and Breast Cancer Risk: An Ecological Study, Nutr. Cancer 12, 61–68.

    Article  PubMed  CAS  Google Scholar 

  5. Petrik, M.B., McEntee, M.F., Johnson, B.T., Obukowicz, M.G., and Whelan, J. (2000) Highly Unsaturated (n−3) Fatty Acids, but Not alpha-Linolenic, Conjugated Linoleic or gamma-Linolenic Acids, Reduce Tumorigenesis in Apc(Min/+) Mice, J. Nutr. 130, 2434–2443.

    PubMed  CAS  Google Scholar 

  6. Rao, C.V., Hirose, Y., Indranie, C., and Reddy, B.S. (2001) Modulation of Experimental Colon Tumorigenesis by Types and Amounts of Dietary Fatty Acids, Cancer Res. 61, 1927–1933.

    PubMed  CAS  Google Scholar 

  7. Heimli, H., Finstad, H.S., and Drevon, C.A. (2001) Necrosis and Apoptosis in Lymphoma Cell Lines Exposed to Eicosapentaenoic Acid and Antioxidants, Lipids 36, 613–621.

    PubMed  CAS  Google Scholar 

  8. Kerr, J.F., Wyllie, A.H., and Currie, A.R. (1972) Apoptosis: A Basic Biological Phenomenon with Wide-Ranging Implications in Tissue Kinetics, Br. J. Cancer 26, 239–257.

    PubMed  CAS  Google Scholar 

  9. Lowe, S.W., and Lin, A.W. (2000) Apoptosis in Cancer, Carcinogenesis 21, 485–495.

    Article  PubMed  CAS  Google Scholar 

  10. Ashkenazi, A., and Dixit, V.M. (1998) Death Receptors: Signaling and Modulation, Science 281, 1305–1308.

    Article  PubMed  CAS  Google Scholar 

  11. Salvesen, G.S., and Dixit, V.M. (1997) Caspases: Intracellular Signaling by Proteolysis, Cell 91, 443–446.

    Article  PubMed  CAS  Google Scholar 

  12. Wallach, D., Boldin, M., Varfolomeev, E., Beyaert, R., Vandenabeele, P., and Fiers, W. (1997) Cell Death Induction by Receptors of the TNF Family: Towards a Molecular Understanding, FEBS Lett. 410, 96–106.

    Article  PubMed  CAS  Google Scholar 

  13. Li, P., Nijhhwan, D., Budihardjo, I., Srinivasula, S.M., Ahmad, M., Alnemri, E.S., and Wang, X. (1997) Cytochrome c and dATP-Dependent Formation of Apaf-1/Caspase-9 Complex Initiates an Apoptotic Protease Cascade, Cell 91, 479–489.

    Article  PubMed  CAS  Google Scholar 

  14. Zou, H., Henzel, W.J., Liu, X., Lutschg, A., and Wang, X. (1997) Apaf-1, a Human Protein Homologous to C. elegans CED-4, Participates in Cytochrome c-Dependent Activation of Caspase-3, Cell 90, 405–413.

    Article  PubMed  CAS  Google Scholar 

  15. Adams, J.M., and Cory, S. (1998) The Bcl-2 Protein Family: Arbiters of Cell Survival, Science 281, 1322–1326.

    Article  PubMed  CAS  Google Scholar 

  16. Green, D.R., and Reed, J.C. (1998) Mitochondria and Apoptosis, Science 281, 1309–1312.

    Article  PubMed  CAS  Google Scholar 

  17. Hengartner, M.O. (2000) The Biochemistry of Apoptosis, Nature 407, 770–776.

    Article  PubMed  CAS  Google Scholar 

  18. Kaufmann, S.H., Desnoyers, S., Ottaviano, Y., Davidson, N.E., and Poirier, G.G. (1993) Specific Proteolytic Clevage of Poly(ADP-ribose) Polymerase: An Early Marker of Chemotherapy-Induced Apoptosis, Cancer Res. 53, 3976–3985.

    PubMed  CAS  Google Scholar 

  19. Lazebnik, Y.A., Kaufmann, S.H., Desnoyers, S., Poirier, G.G., and Earnshaw, W.C. (1994) Cleavage of Poly(ADP-ribose) Polymerase by a Proteinase with Properties like ICE, Nature 371, 346–347.

    Article  PubMed  CAS  Google Scholar 

  20. Yung, T.M., and Satoh, M.S. (2001) Functional Competition Between Poly(ADP-ribose) Polymerase and Its 24- kDa Apoptotic Fragment in DNA Repair and Transcription, J. Biol. Chem. 276, 11279–11286.

    Article  PubMed  CAS  Google Scholar 

  21. Gastman, B.R., Yin, X.M., Johnson, D.E., Wieckowski, E., Wang, G.Q., Watkins, G.C., and Rabinowich, H. (2000) Tumor-Induced Apoptosis of T Cells: Amplification by a Mitochondrial Cascade, Cancer Res. 60, 6811–6817.

    PubMed  CAS  Google Scholar 

  22. Schempp, C.M., Simon-Haarhaus, B., Termeer, C.C., and Simon, J.C. (2001) Hypericin Photo-Induced Apoptosis Involves the Tumor Necrosis Factor-Related Apoptosis-Inducing Ligand (TRAIL) and Activation of Caspase-8, FEBS Lett. 493, 26–30.

    Article  PubMed  CAS  Google Scholar 

  23. Takahashi, A., Hirata, H., Yonehara, S., Imai, Y., Lee, K.K., Moyer, R.W., Turner, P.C., Mesner, P.W., Okazaki, T., Sawai, H. et al. (1997) Affinity Labeling Displays the Stepwise Activation of ICE-Related Proteases by Fas, Staurosporine, and CrmA-Sensitive Caspase-8, Oncogene 14, 2741–2752.

    Article  PubMed  CAS  Google Scholar 

  24. Bratton, D.L., Fadok, V.A., Richter, D.A., Kailey, J.M., Guthrie, L.A., and Henson, P.M. (1997) Appearance of Phosphatidylserine on Apoptotic Cells Requires Calcium-Mediated Nonspecific Flip-Flop and Is Enhanced by Loss of the Aminophospholipid Translocase, J. Biol. Chem. 272, 26159–26165.

    Article  PubMed  CAS  Google Scholar 

  25. Span, L.F., Pennings, A.H., Vierwinden, G., Boezeman, J.B., Raymakers, R.A., and de Witte, T. (2002) The Dynamic Process of Apoptosis Analyzed by Flow Cytometry Using Annexin-V/Propidium Iodide and a Modified in situ End Labeling Technique, Cytometry 47, 24–31.

    Article  PubMed  CAS  Google Scholar 

  26. Zhuang, J., Ren, Y., Snowden, R.T., Zhu, H., Gogvadze, V., Savill, J.S., and Cohen, G.M. (1998) Dissociation of Phagocyte Recognition of Cells Undergoing Apoptosis from Other Features of the Apoptotic Program, J. Biol. Chem. 273, 15628–15632.

    Article  PubMed  CAS  Google Scholar 

  27. Cain, K., Brown, D.G., Langlais, C., and Cohen, G.M. (1999) Caspase Activation Involves the Formation of the Aposome, a Large (approximately 700 kDa) Caspase-Activating Complex, J. Biol. Chem. 274, 22686–22692.

    Article  PubMed  CAS  Google Scholar 

  28. Antonsson, B., and Martinou, J.C. (2000) The Bcl-2 Protein Family, Exp. Cell Res. 256, 50–57.

    Article  PubMed  CAS  Google Scholar 

  29. Gross, A., McDonnell, J.M., and Korsmeyer, S.J. (1999) BCL-2 Family Members and the Mitochondria in Apoptosis, Genes Dev. 13, 1899–1911.

    PubMed  CAS  Google Scholar 

  30. Reed, J.C. (1997) Double Identity for Proteins of the BCL-2 Family, Nature 387, 773–776.

    Article  PubMed  CAS  Google Scholar 

  31. Glazyrin, A.L., Adsay, V.N., Vaitkevicius, V.K., and Sarkar, F.H. (2001) CD95-Related Apoptotic Machinery Is Functional in Pancreatic Cancer Cells, Pancreas 22, 357–365.

    Article  PubMed  CAS  Google Scholar 

  32. Herrera, B., Fernandez, M., Alvarez, A.M., Roncero, C., Benito, M., Gil, J., and Fabregat, I. (2001) Activation of Caspases Occurs Downstream from Radical Oxygen Species Production, Bcl-xL Down-Regulation, and Early Cytochrome C Release in Apoptosis Induced by Transforming Growth Factor beta in Rat Fetal Hepatocytes, Hepatology 34, 548–556.

    Article  PubMed  CAS  Google Scholar 

  33. Kiyokawa, N., Mori, T., Taguchi, T., Saito, M., Mimori, K., Suzuki, T., Sekino, T., Sato, N., Nakajima, H., Katagiri, Y.U., et al. (2001) Activation of the Caspase Cascade During Stx1-Induced Apoptosis in Burkitt's Lymphoma Cells, J. Cell Biochem. 81, 128–142.

    Article  PubMed  CAS  Google Scholar 

  34. Finstad, H.S., Drevon, C.A., Kulseth, M.A., Synstad, A.V., Knudsen, E., and Kolset, S.O. (1998) Cell Proliferation, Apoptosis and Accumulation of Lipid Droplets in U937-1 Cells Incubated with Eicosapentaenoic Acid, Biochem. J. 336, 451–459.

    PubMed  CAS  Google Scholar 

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Authors and Affiliations

  1. Institute for Medical Biochemistry, University of Oslo, Norway

    Soheil Naderi

  2. Institute for Nutrition Research, University of Oslo, Blindern, P.O. Box 1046, 0316, Oslo, Norway

    Hilde Heimli, Camilla Giske, Christian A. Drevon & Kristin Hollung

Authors
  1. Hilde Heimli
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  2. Camilla Giske
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  3. Soheil Naderi
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  4. Christian A. Drevon
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  5. Kristin Hollung
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Correspondence to Christian A. Drevon.

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Heimli, H., Giske, C., Naderi, S. et al. Eicosapentaenoic acid promotes apoptosis in Ramos cells via activation of caspase-3 and -9. Lipids 37, 797–802 (2002). https://doi.org/10.1007/s11745-002-0963-6

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  • DOI: https://doi.org/10.1007/s11745-002-0963-6

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