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P2X7 receptor activation induces cell death and CD23 shedding in human RPMI 8226 multiple myeloma cells

Journal Article


Abstract


  • Background: The extracellular ATP-gated cation channel, P2X7 receptor, has an emerging role in neoplasia, however progress in the field is limited by a lack of malignant cell lines expressing this receptor.

    Methods: Immunofluorescence labelling and a fixed-time ATP-induced ethidium(+) uptake assay were used to screen a panel of human malignant cell lines for the presence of functional P2X7. The presence of P2X7 was confirmed by RT-PCR, immunoblotting and pharmacological approaches. ATP-induced cell death was measured by colourimetric tetrazolium-based and cytofluorometric assays. ATP-induced CD23 shedding was measured by immunofluorescence labelling and ELISA.

    Results: RPMI 8226 multiple myeloma cells expressed P2X7 mRNA and protein, as well as P2X1, P2X4 and P2X5 mRNA. ATP induced ethidium(+) uptake into these cells with an EC50 of similar to 116 mu M, and this uptake was reduced in the presence of extracellular Ca2+ and Mg2+. The P2X7 agonist 2'- and 3'-0(4-benzoylbenzoyl) ATP, but not UTP, induced ethidium(+) uptake. ATP-induced ethidium(+) uptake was impaired by the P2X7 antagonists, KN-62 and A-438079. ATP induced death and CD23 shedding in RPMI 8226 cells, and both processes were impaired by P2X7 antagonists. The metalloprotease antagonists, BB-94 and GM6001, impaired ATP-induced CD23 shedding but not ethidium(+) uptake.

    Conclusions: P2X7 receptor activation induces cell death and CD23 shedding in RPMI 8226 cells. General significance: RPMI 8226 cells may be useful to study the role of P2X7 in multiple myeloma and B-lymphocytes. (C) 2010 Elsevier B.V. All rights reserved.

Authors


  •   Farrell, Andrew W. (external author)
  •   Gadeock, Safina (external author)
  •   Pupovac, Aleta (external author)
  •   Wang, Bin (external author)
  •   Jalilian, Iman (external author)
  •   Ranson, Marie
  •   Sluyter, Ronald

Publication Date


  • 2010

Citation


  • Farrell, A. W., Gadeock, S., Pupovac, A., Wang, B., Jalilian Hassan Pour, I., Ranson, M. & Sluyter, R. (2010). P2X7 receptor activation induces cell death and CD23 shedding in human RPMI 8226 multiple myeloma cells. BBA - General Subjects, 1800 (11), 1173-1182.

Scopus Eid


  • 2-s2.0-77956289939

Ro Metadata Url


  • http://ro.uow.edu.au/scipapers/5077

Number Of Pages


  • 9

Start Page


  • 1173

End Page


  • 1182

Volume


  • 1800

Issue


  • 11

Abstract


  • Background: The extracellular ATP-gated cation channel, P2X7 receptor, has an emerging role in neoplasia, however progress in the field is limited by a lack of malignant cell lines expressing this receptor.

    Methods: Immunofluorescence labelling and a fixed-time ATP-induced ethidium(+) uptake assay were used to screen a panel of human malignant cell lines for the presence of functional P2X7. The presence of P2X7 was confirmed by RT-PCR, immunoblotting and pharmacological approaches. ATP-induced cell death was measured by colourimetric tetrazolium-based and cytofluorometric assays. ATP-induced CD23 shedding was measured by immunofluorescence labelling and ELISA.

    Results: RPMI 8226 multiple myeloma cells expressed P2X7 mRNA and protein, as well as P2X1, P2X4 and P2X5 mRNA. ATP induced ethidium(+) uptake into these cells with an EC50 of similar to 116 mu M, and this uptake was reduced in the presence of extracellular Ca2+ and Mg2+. The P2X7 agonist 2'- and 3'-0(4-benzoylbenzoyl) ATP, but not UTP, induced ethidium(+) uptake. ATP-induced ethidium(+) uptake was impaired by the P2X7 antagonists, KN-62 and A-438079. ATP induced death and CD23 shedding in RPMI 8226 cells, and both processes were impaired by P2X7 antagonists. The metalloprotease antagonists, BB-94 and GM6001, impaired ATP-induced CD23 shedding but not ethidium(+) uptake.

    Conclusions: P2X7 receptor activation induces cell death and CD23 shedding in RPMI 8226 cells. General significance: RPMI 8226 cells may be useful to study the role of P2X7 in multiple myeloma and B-lymphocytes. (C) 2010 Elsevier B.V. All rights reserved.

Authors


  •   Farrell, Andrew W. (external author)
  •   Gadeock, Safina (external author)
  •   Pupovac, Aleta (external author)
  •   Wang, Bin (external author)
  •   Jalilian, Iman (external author)
  •   Ranson, Marie
  •   Sluyter, Ronald

Publication Date


  • 2010

Citation


  • Farrell, A. W., Gadeock, S., Pupovac, A., Wang, B., Jalilian Hassan Pour, I., Ranson, M. & Sluyter, R. (2010). P2X7 receptor activation induces cell death and CD23 shedding in human RPMI 8226 multiple myeloma cells. BBA - General Subjects, 1800 (11), 1173-1182.

Scopus Eid


  • 2-s2.0-77956289939

Ro Metadata Url


  • http://ro.uow.edu.au/scipapers/5077

Number Of Pages


  • 9

Start Page


  • 1173

End Page


  • 1182

Volume


  • 1800

Issue


  • 11