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P2X7 receptor activation induces cell death and microparticle release in murine erythroleukemia cells

Journal Article


Abstract


  • Extracellular ATP induces cation fluxes in and impairs the growth of murine erythroleukemia (MEL) cells in a manner characteristic of the purinergic P2X7 receptor, however the presence of P2X7 in these cells is unknown. This study investigated whether MEL cells express functional P2X7. RT-PCR, immunoblotting and immunofluorescence staining demonstrated the presence of P2X7 in MEL cells. Cytofluorometric measurements demonstrated that ATP induced ethidium+ uptake into MEL cells in a concentration-dependent fashion and with an EC50 of ∼ 154 μM. The most potent P2X7 agonist 2′- and 3′-0(4-benzoylbenzoyl) ATP, but not ADP or UTP, induced ethidium+ uptake. ATP-induced ethidium+ and YO-PRO-12+ uptake were impaired by the P2X7 antagonist, A-438079. A colourmetric assay demonstrated that ATP impaired MEL cell growth. A cytofluorometric assay showed that ATP induced MEL cell death and that this process was impaired by A-438079. Finally, cytofluorometric measurements of Annexin-V binding and bio-maleimide staining demonstrated that ATP could induce rapid phosphatidylserine exposure and microparticle release in MEL cells respectively, both of which were impaired by A-438079. These results demonstrate that MEL cells express functional P2X7, and indicate that activation of this receptor may be important in the death and release of microparticles from red blood cells in vivo. © 2010 Elsevier B.V. All rights reserved.

Authors


  •   Constantinescu, Patrick
  •   Wang, Bin (external author)
  •   Kovacevic, Kati (external author)
  •   Jalilian, Iman (external author)
  •   Dr Giel J Bosman, Giel J (external author)
  •   Wiley, James S. (external author)
  •   Sluyter, Ronald

Publication Date


  • 2010

Citation


  • Constantinescu, P., Wang, B., Kovacevic, K., Jalilian Hassan Pour, I., Bosman, G. J.C.G.M., Wiley, J. S. & Sluyter, R. (2010). P2X7 receptor activation induces cell death and microparticle release in murine erythroleukemia cells. BBA - Biomembranes, 1798 (9), 1797-1804.

Scopus Eid


  • 2-s2.0-77954660508

Ro Metadata Url


  • http://ro.uow.edu.au/scipapers/5061

Number Of Pages


  • 7

Start Page


  • 1797

End Page


  • 1804

Volume


  • 1798

Issue


  • 9

Abstract


  • Extracellular ATP induces cation fluxes in and impairs the growth of murine erythroleukemia (MEL) cells in a manner characteristic of the purinergic P2X7 receptor, however the presence of P2X7 in these cells is unknown. This study investigated whether MEL cells express functional P2X7. RT-PCR, immunoblotting and immunofluorescence staining demonstrated the presence of P2X7 in MEL cells. Cytofluorometric measurements demonstrated that ATP induced ethidium+ uptake into MEL cells in a concentration-dependent fashion and with an EC50 of ∼ 154 μM. The most potent P2X7 agonist 2′- and 3′-0(4-benzoylbenzoyl) ATP, but not ADP or UTP, induced ethidium+ uptake. ATP-induced ethidium+ and YO-PRO-12+ uptake were impaired by the P2X7 antagonist, A-438079. A colourmetric assay demonstrated that ATP impaired MEL cell growth. A cytofluorometric assay showed that ATP induced MEL cell death and that this process was impaired by A-438079. Finally, cytofluorometric measurements of Annexin-V binding and bio-maleimide staining demonstrated that ATP could induce rapid phosphatidylserine exposure and microparticle release in MEL cells respectively, both of which were impaired by A-438079. These results demonstrate that MEL cells express functional P2X7, and indicate that activation of this receptor may be important in the death and release of microparticles from red blood cells in vivo. © 2010 Elsevier B.V. All rights reserved.

Authors


  •   Constantinescu, Patrick
  •   Wang, Bin (external author)
  •   Kovacevic, Kati (external author)
  •   Jalilian, Iman (external author)
  •   Dr Giel J Bosman, Giel J (external author)
  •   Wiley, James S. (external author)
  •   Sluyter, Ronald

Publication Date


  • 2010

Citation


  • Constantinescu, P., Wang, B., Kovacevic, K., Jalilian Hassan Pour, I., Bosman, G. J.C.G.M., Wiley, J. S. & Sluyter, R. (2010). P2X7 receptor activation induces cell death and microparticle release in murine erythroleukemia cells. BBA - Biomembranes, 1798 (9), 1797-1804.

Scopus Eid


  • 2-s2.0-77954660508

Ro Metadata Url


  • http://ro.uow.edu.au/scipapers/5061

Number Of Pages


  • 7

Start Page


  • 1797

End Page


  • 1804

Volume


  • 1798

Issue


  • 9