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P2X7 receptor activation mediates organic cation uptake into human myeloid leukaemic KG-1 cells

Journal Article


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Abstract


  • The P2X7 purinergic receptor is an ATP-gated cation channel with an emerging role in neoplasia. In this study we demonstrate that the human KG-1 cell line, a model of acute myelogenous leukaemia, expresses functional P2X7. RT-PCR and immunochemical techniques demonstrated the presence of P2X7 mRNA and protein respectively in KG-l cells, as well as in positive control multiple myeloma RPMI 8226 cells. Flow cytometric measurements demonstrated that ATP induced ethidium + uptake into KG-l cells suspended in sucrose medium (EC 50 of ~3 μM), but not into cells in NaCl medium. In contrast, ATP induced ethidium + uptake into RPMI 8226 cells suspended in either sucrose or NaCl medium (EC 50 of ~3 or ~99 μM, respectively), as well as into RPMI 8226 cells in KCl medium (EC 50 of ~18 μM). BzATP and to a lesser extent ATPγS and αβ-methylene ATP, but not ADP or UTP, also induced ethidium + uptake into KG-1 cells. ATP-induced ethidium + uptake was completely impaired by the P2X7 antagonists, AZ10606120 and A-438079. ATP-induced ethidium + uptake was also impaired by probenecid but not by carbenoxolone, both pannexin-1 antagonists. ATP induced YO-PRO-1 2+ and propidium 2+ uptake into KG-1 cells. Finally, sequencing of full-length P2X7 cDNA identified several single nucleotide polymorphisms (SNPs) in KG-1 cells including H155Y, A348T, T357S and Q460R. RPMI 8226 cells contained A348T, A433V and H521Q SNPs. In conclusion, the KG-1 cell line expresses functional P2X7. This cell line may help elucidate the signalling pathways involved in P2X7-induced survival and invasiveness of myeloid leukaemic cells. © 2012 Springer Science+Business Media B.V.

Authors


  •   Gadeock, Safina (external author)
  •   Pupovac, Aleta (external author)
  •   Sluyter, Vanessa (external author)
  •   Spildrejorde, Mari (external author)
  •   Sluyter, Ronald

Publication Date


  • 2012

Citation


  • Gadeock, S., Pupovac, A., Sluyter, V., Spildrejorde, M. & Sluyter, R. (2012). P2X7 receptor activation mediates organic cation uptake into human myeloid leukaemic KG-1 cells. Purinergic Signalling, 8 (4), 669-676.

Scopus Eid


  • 2-s2.0-84867883658

Ro Full-text Url


  • http://ro.uow.edu.au/cgi/viewcontent.cgi?article=7908&context=scipapers

Ro Metadata Url


  • http://ro.uow.edu.au/scipapers/4565

Number Of Pages


  • 7

Start Page


  • 669

End Page


  • 676

Volume


  • 8

Issue


  • 4

Abstract


  • The P2X7 purinergic receptor is an ATP-gated cation channel with an emerging role in neoplasia. In this study we demonstrate that the human KG-1 cell line, a model of acute myelogenous leukaemia, expresses functional P2X7. RT-PCR and immunochemical techniques demonstrated the presence of P2X7 mRNA and protein respectively in KG-l cells, as well as in positive control multiple myeloma RPMI 8226 cells. Flow cytometric measurements demonstrated that ATP induced ethidium + uptake into KG-l cells suspended in sucrose medium (EC 50 of ~3 μM), but not into cells in NaCl medium. In contrast, ATP induced ethidium + uptake into RPMI 8226 cells suspended in either sucrose or NaCl medium (EC 50 of ~3 or ~99 μM, respectively), as well as into RPMI 8226 cells in KCl medium (EC 50 of ~18 μM). BzATP and to a lesser extent ATPγS and αβ-methylene ATP, but not ADP or UTP, also induced ethidium + uptake into KG-1 cells. ATP-induced ethidium + uptake was completely impaired by the P2X7 antagonists, AZ10606120 and A-438079. ATP-induced ethidium + uptake was also impaired by probenecid but not by carbenoxolone, both pannexin-1 antagonists. ATP induced YO-PRO-1 2+ and propidium 2+ uptake into KG-1 cells. Finally, sequencing of full-length P2X7 cDNA identified several single nucleotide polymorphisms (SNPs) in KG-1 cells including H155Y, A348T, T357S and Q460R. RPMI 8226 cells contained A348T, A433V and H521Q SNPs. In conclusion, the KG-1 cell line expresses functional P2X7. This cell line may help elucidate the signalling pathways involved in P2X7-induced survival and invasiveness of myeloid leukaemic cells. © 2012 Springer Science+Business Media B.V.

Authors


  •   Gadeock, Safina (external author)
  •   Pupovac, Aleta (external author)
  •   Sluyter, Vanessa (external author)
  •   Spildrejorde, Mari (external author)
  •   Sluyter, Ronald

Publication Date


  • 2012

Citation


  • Gadeock, S., Pupovac, A., Sluyter, V., Spildrejorde, M. & Sluyter, R. (2012). P2X7 receptor activation mediates organic cation uptake into human myeloid leukaemic KG-1 cells. Purinergic Signalling, 8 (4), 669-676.

Scopus Eid


  • 2-s2.0-84867883658

Ro Full-text Url


  • http://ro.uow.edu.au/cgi/viewcontent.cgi?article=7908&context=scipapers

Ro Metadata Url


  • http://ro.uow.edu.au/scipapers/4565

Number Of Pages


  • 7

Start Page


  • 669

End Page


  • 676

Volume


  • 8

Issue


  • 4