Calicum microdomains form within neutrophils at the neutrophil–tumor cell synapse: role in antibody-dependent target cell apoptosis

  • Andrea J. Clark
  • Michelle Diamond
  • Megan Elfline
  • Howard R. Petty
Original Article
First Online:
Received:
Accepted:

Abstract

Ca2+ messages are broadly important in cellular signal transduction. In immune cells, Ca2+ signaling is an essential step in many forms of activation. Neutrophil-mediated antibody-dependent cell-mediated cytotoxicity (ADCC) is one form of leukocyte activation that plays an important role in tumor cell killing in vitro and in patient care. Using fluorescence methodologies, we found that neutrophils exhibit Ca2+ signals during ADCC directed against breast fibrosarcoma cells. Importantly, these signals were localized to Ca2+ microdomains at the neutrophil-to-tumor cell interface where they display dynamic features such as movement, fusion, and fission. These signals were blocked by the intracellular Ca2+ buffer BAPTA. At the neutrophil–tumor cell synapse, the neutrophil’s cytoplasm was enriched in STIM1, a crucial mediator of Ca2+ signaling, whereas the Ca2+-binding proteins calbindin and parvalbumin were not affected. Our findings suggest that Ca2+ microdomains are due to an active signaling process. As Ca2+ signals within neutrophils were necessary for specific tumor cell apoptosis, a central role of microdomains in leukocyte-mediated tumor cell destruction is indicated.

Keywords

Breast cancer Apoptosis Antibody Signal transduction Calcium 
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Notes

Acknowledgments

This work was supported by the NCI and by the Wilson Medical Foundation.

Supplementary material

262_2009_735_MOESM1_ESM.pdf (408 kb)
Supplementary figures (PDF 409 kb)

References

  1. 1.
    Adams GP, Weiner LM (2005) Monoclonal antibody therapy of cancer. Nat Biotechnol 23(9):1147–1157CrossRefPubMedGoogle Scholar
  2. 2.
    Vokes EE, Chu E (2006) Anti-EGFR therapies: clinical experience in colorectal, lung, and head and neck cancers. Oncology 20(5 Suppl 2):15–25PubMedGoogle Scholar
  3. 3.
    Di Carlo E, Forni G, Lollini PL, Colombo MP, Modesti A, Musiani P (2001) The intriguing role of polymorphonuclear neutrophils in antitumor reactions. Blood 97(2):339–345CrossRefPubMedGoogle Scholar
  4. 4.
    Desjarlais JR, Lazar GA, Zhukovsky EA, Chu SY (2007) Optimizing engagement of the immune system by anti-tumor antibodies: an engineer’s perspective. Drug Discov Today 12(21–22):898–910PubMedGoogle Scholar
  5. 5.
    Hara I, Takechi Y, Houghton AN (1995) Implicating a role for immune recognition of self in tumor rejection: passive immunization against the brown locus protein. J Exp Med 182(5):1609–1614CrossRefPubMedGoogle Scholar
  6. 6.
    Naftzger C, Takechi Y, Kohda H, Hara I, Vijayasaradhi S, Houghton AN (1996) Immune response to a differentiation antigen induced by altered antigen: a study of tumor rejection and autoimmunity. Proc Natl Acad Sci USA 93(25):14809–14814CrossRefPubMedGoogle Scholar
  7. 7.
    Clynes R, Takechi Y, Moroi Y, Houghton A, Ravetch JV (1998) Fc receptors are required in passive and active immunity to melanoma. Proc Natl Acad Sci USA 95(2):652–656CrossRefPubMedGoogle Scholar
  8. 8.
    Clynes RA, Towers TL, Presta LG, Ravetch JV (2000) Inhibitory Fc receptors modulate in vivo cytoxicity against tumor targets. Nat Med 6(4):443–446CrossRefPubMedGoogle Scholar
  9. 9.
    Bibeau F, Lopez-Crapez E, Di Fiore F et al (2009) Impact of FcγRIIa-FcγRIIIa polymorphisms and KRAS mutations on the clinical outcome of patients with metastatic colorectal cancer treated with cetuximab plus irinotecan. J Clin Oncol 27(7):1122–1129CrossRefPubMedGoogle Scholar
  10. 10.
    Cartron G, Dacheux L, Salles G et al (2002) Therapeutic activity of humanized anti-CD20 monoclonal antibody and polymorphism in IgG Fc receptor FcγRIIIa gene. Blood 99(3):754–758CrossRefPubMedGoogle Scholar
  11. 11.
    Musolino A, Naldi N, Bortesi B et al (2008) Immunoglobulin G fragment C receptor polymorphisms and clinical efficacy of trastuzumab-based therapy in patients with HER-2/neu-positive metastatic breast cancer. J Clin Oncol 26(11):1789–1796CrossRefPubMedGoogle Scholar
  12. 12.
    van de Winkel JGJ, Hogarth PM (1998) The immunoglobulin receptors and their physiological and pathological roles in immunity. Kluwer Academic Publishers, DordrechtGoogle Scholar
  13. 13.
    Stadick H, Stockmeyer B, Kühn R et al (2002) Epidermal growth factor receptor and g250: useful target antigens for antibody mediated cellular cytotoxicity against renal cell carcinoma? J Urol 167(2 Pt 1):707–712PubMedGoogle Scholar
  14. 14.
    Stockmeyer B, Valerius T, Repp R et al (1997) Preclinical studies with FcγR bispecific antibodies and granulocyte colony-stimulating factor-primed neutrophils as effector cells against HER-2/neu overexpressing breast cancer. Cancer Res 57(4):696–701PubMedGoogle Scholar
  15. 15.
    Würflein D, Dechant M, Stockmeyer B et al (1998) Evaluating antibodies for their capacity to induce cell-mediated lysis of malignant B cells. Cancer Res 58(14):3051–3058PubMedGoogle Scholar
  16. 16.
    Challacombe JM, Suhrbier A, Parsons PG et al (2006) Neutrophils are a key component of the antitumor efficacy of topical chemotherapy with ingenol-3-angelate. J Immunol 177(11):8123–8132PubMedGoogle Scholar
  17. 17.
    Santana C, Noris G, Espinoza B, Ortega E (1996) Protein tyrosine phosphorylation in leukocyte activation through receptors for IgG. J Leukoc Biol 60(4):433–440PubMedGoogle Scholar
  18. 18.
    Safiejko-Mroczka B, Bell PB Jr (1996) Bifunctional protein cross-linking reagents improve labeling of cytoskeletal proteins for qualitative and quantitative fluorescence microscopy. J Histochem Cytochem 44(6):641–656PubMedGoogle Scholar
  19. 19.
    Clark AJ, Petty HR (2007) Super-quiet microfluorometry: examples of tumor cell metabolic dynamics. In: Méndez-Vilas A, Díaz J (eds) Modern research and educational topics on microscopy. FORMATEX, Spain, pp 403–408Google Scholar
  20. 20.
    Clark AJ, Petty HR (2008) Observation of calcium microdomains at the uropod of living morphologically polarized human neutrophils using flashlamp-based fluorescence microscopy. Cytometry A 73(7):673–678PubMedGoogle Scholar
  21. 21.
    Feske S (2007) Calcium signalling in lymphocyte activation and disease. Nat Rev Immunol 7(9):690–702CrossRefPubMedGoogle Scholar
  22. 22.
    Dastoor Z, Dreyer JL (2001) Potential role of nuclear translocation of glyceraldehyde-3-phosphate dehydrogenase in apoptosis and oxidative stress. J Cell Sci 114:1643–1653PubMedGoogle Scholar
  23. 23.
    van de Winkel JG, Tax WJ, Jacobs CW, Huizinga TW, Willems PH (1990) Cross-linking of both types of IgG Fc receptors, FcγRI and FcγRII, enhances intracellular free Ca2+ in the monocytic cell line U937. Scand J Immunol 31(3):315–325CrossRefPubMedGoogle Scholar
  24. 24.
    Rizzuto R, Pinton P, Ferrari D et al (2003) Calcium and apoptosis: facts and hypotheses. Oncogene 22(53):8619–8627CrossRefPubMedGoogle Scholar
  25. 25.
    Bakowski D, Parekh AB (2007) Regulation of store-operated calcium channels by the intermediary metabolite pyruvic acid. Curr Biol 17(12):1076–1081CrossRefPubMedGoogle Scholar
  26. 26.
    Melamed-Book N, Kachalsky SG, Kaiserman I, Rahamimoff R (1999) Neuronal calcium sparks and intracellular calcium “noise”. Proc Natl Acad Sci USA 96(26):15217–15221CrossRefPubMedGoogle Scholar
  27. 27.
    van de Winkel JG, Anderson CL (1991) Biology of human immunoglobulin G Fc receptors. J Leukoc Biol 49(5):511–524PubMedGoogle Scholar
  28. 28.
    van de Winkel JG, van Duijnhoven HL, van Ommen R, Capel PJ, Tax WJ (1988) Selective modulation of two human monocyte Fc receptors for IgG by immobilized immune complexes. J Immunol 140(10):3515–3521PubMedGoogle Scholar
  29. 29.
    Kávai M, Gyimesi E, Szücs G, Szegedi G (1991) Binding and endocytosis of erythrocytes sensitized with rabbit IgG via Fc γ receptors of human monocytes. Immunology 74(4):657–660PubMedGoogle Scholar
  30. 30.
    Flesch BK, Vöge K, Henrichs T, Neppert J (2001) Fcγ receptor-mediated immune phagocytosis depends on the class of FcγR and on the immunoglobulin-coated target cell. Vox Sang 81(2):128–133CrossRefPubMedGoogle Scholar
  31. 31.
    Li P, Jiang N, Nagarajan S, Wohlhueter R, Selvaraj P, Zhu C (2007) Affinity and kinetic analysis of Fcγ receptor IIIa (CD16a) binding to IgG ligands. J Biol Chem 282(9):6210–6221CrossRefPubMedGoogle Scholar
  32. 32.
    Petty HR, Francis JW, Anderson CL (1989) Cell surface distribution of Fc receptors II and III on living human neutrophils before and during antibody dependent cellular cytotoxicity. J Cell Physiol 141(3):598–605CrossRefPubMedGoogle Scholar
  33. 33.
    Kwiatkowska K, Frey J, Sobota A (2003) Phosphorylation of FcγRIIA is required for the receptor-induced actin rearrangement and capping: the role of membrane rafts. J Cell Sci 116(Pt 3):537–550CrossRefPubMedGoogle Scholar
  34. 34.
    David A, Fridlich R, Aviram I (2005) The presence of membrane Proteinase 3 in neutrophil lipid rafts and its colocalization with FcγRIIIb and cytochrome b558. Exp Cell Res 308(1):156–165CrossRefPubMedGoogle Scholar
  35. 35.
    Petty HR, Worth RG, Todd RF (2002) Interactions of integrins with their partner proteins in leukocyte membranes. Immunol Res 25(1):75–95CrossRefPubMedGoogle Scholar
  36. 36.
    van Spriel AB, Leusen JH, van Egmond M et al (2001) Mac-1 (CD11b/CD18) is essential for Fc receptor-mediated neutrophil cytotoxicity and immunologic synapse formation. Blood 97(8):2478–2486CrossRefPubMedGoogle Scholar
  37. 37.
    Kushner BH, Cheung NK (1992) Absolute requirement of CD11/CD18 adhesion molecules, FcRII and the phosphatidylinositol-linked FcRIII for monoclonal antibody-mediated neutrophil antihuman tumor cytotoxicity. Blood 79(6):1484–1490PubMedGoogle Scholar
  38. 38.
    Van Spriel A, Van Ojik H, Bakker A et al (2003) Mac-1 (CD11b/CD18) is crucial for effective Fc receptor-mediated immunity to melanoma. Blood 101(1):253–258CrossRefPubMedGoogle Scholar
  39. 39.
    Pani B, Ong H, Liu X et al (2008) Lipid rafts determine clustering of STIM1 in endoplasmic reticulum-plasma membrane junctions and regulation of store-operated Ca2+ entry (SOCE). J Biol Chem 283(25):17333–17340CrossRefPubMedGoogle Scholar
  40. 40.
    Yang S, Zhang JJ, Huang XY (2009) Orai1 and STIM1 are critical for breast tumor cell migration and metastasis. Cancer Cell 15(2):124–134CrossRefPubMedGoogle Scholar
  41. 41.
    Dykstra M, Cherukuri A, Sohn HW, Tzeng SJ, Pierce SK (2003) Location is everything: lipid rafts and immune cell signaling. Annu Rev Immunol 21:457–481CrossRefPubMedGoogle Scholar
  42. 42.
    Dallegri F, Patrone F, Frumento G, Sacchetti C (1984) Antibody-dependent killing of tumor cells by polymorphonuclear leukocytes. Involvement of oxidative and nonoxidative mechanisms. J Natl Cancer Inst 73(2):331–339PubMedGoogle Scholar
  43. 43.
    Hafeman DG, Lucas ZJ (1979) Polymorphonuclear leukocyte-mediated, antibody-dependent, cellular cytotoxicity against tumor cells: dependence on oxygen and the respiratory burst. J Immunol 123(1):55–62PubMedGoogle Scholar
  44. 44.
    Borregaard N, Cowland JB (1997) Granules of the human neutrophilic polymorphonuclear leukocyte. Blood 89(10):3503–3521PubMedGoogle Scholar
  45. 45.
    Lew PD (1990) Receptors and intracellular signaling in human neutrophils. Am Rev Respir Dis 141(3 Pt 2):S127–S131PubMedGoogle Scholar
  46. 46.
    Stockmeyer B, Beyer T, Neuhuber W et al (2003) Polymorphonuclear granulocytes induce antibody-dependent apoptosis in human breast cancer cells. J Immunol 171(10):5124–5129PubMedGoogle Scholar
  47. 47.
    Mazumder S, Plesca D, Almasan A (2008) Caspase-3 activation is a critical determinant of genotoxic stress-induced apoptosis. Methods Mol Biol 414:13–21CrossRefPubMedGoogle Scholar
  48. 48.
    Chuang DM, Hough C, Senatorov VV (2005) Glyceraldehyde-3-phosphate dehydrogenase, apoptosis, and neurodegenerative diseases. Annu Rev Pharmacol Toxicol 45:269–290CrossRefPubMedGoogle Scholar
  49. 49.
    Shashidharan P, Chalmers-Redman RM, Carlile GW et al (1999) Nuclear translocation of GAPDH-GFP fusion protein during apoptosis. Neuroreport 10(5):1149–1153CrossRefPubMedGoogle Scholar
  50. 50.
    Netterwald J (2008) Lights! Camera! Action!. Drug Discov Dev 11(2):18–22Google Scholar
  51. 51.
    Hinkovska-Galcheva V, Clark AJ, Hiraoka M et al (2007) Ceramide kinase mediates store operated calcium signals and enhances phagolysosomal fusion. J Lipid Res 49(3):531–542CrossRefPubMedGoogle Scholar
  52. 52.
    Pan JG, Mak TW (2007) Metabolic targeting as an anticancer strategy: dawn of a new era? Sci STKE 2007(381):pe14PubMedGoogle Scholar
  53. 53.
    Bonnet S, Archer SL, Allalunis-Turner J et al (2007) A mitochondria-K+ channel axis is suppressed in cancer and its normalization promotes apoptosis and inhibits cancer growth. Cancer Cell 11(1):37–51CrossRefPubMedGoogle Scholar
  54. 54.
    Lardner A (2001) The effects of extracellular pH on immune function. J Leukoc Biol 69(4):522–530PubMedGoogle Scholar
  55. 55.
    Zablocki K, Szczepanowska J, Duszynski J (2005) Extracellular pH modifies mitochondrial control of capacitative calcium entry in Jurkat cells. J Biol Chem 280(5):3516–3521CrossRefPubMedGoogle Scholar

Copyright information

© Springer-Verlag 2009

Authors and Affiliations

  • Andrea J. Clark
    • 1
  • Michelle Diamond
    • 1
  • Megan Elfline
    • 1
  • Howard R. Petty
    • 1
    • 2
  1. 1.Department of Ophthalmology and Visual SciencesThe University of Michigan Medical SchoolAnn ArborUSA
  2. 2.Department of Microbiology and ImmunologyThe University of Michigan Medical SchoolAnn ArborUSA

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