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P2X7 receptor activation leads to increased cell death in a radiosensitive human glioma cell line

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Abstract

Gliomas are the most lethal tumors of central nervous system. ATP is an important signaling molecule in CNS and it is a selective P2X7 purinergic receptor ligand at high concentrations. Herein, we investigated whether the activation of P2X7R might be implicated in death of a radiosensitive human glioma lineage. The effects of P2X7R agonists (ATP and BzATP) and irradiation (2 Gy) on glioma cells were analyzed by MTT assay and annexin-V/PI determination, whereas mRNA and protein P2X7R expression was assessed by qRT-PCR and flow cytometry, respectively. P2X7R pore formation was functionality examined by analyzing ethidium bromide uptake. The human glioma cells U-138 MG and U-251 MG were resistant to death when treated with either ATP (5 mM) or BzATP (100 μM), but the radiosensitive M059J glioma cells displayed a significant decrease of cell viability (32.4 ± 4.1 % and 25.6 ± 3.3 %, respectively). The M059J lineage expresses significantly higher mRNA P2X7R levels when compared to the U-138 MG and U-251 cell lines (0.40 ± 0.00; 0.28 ± 0.01, and 0.31 ± 0.01, respectively), and irradiation upregulated P2X7R expression (0.55 ± 0.08) in this lineage. Noteworthy, P2X7R protein doubled after irradiation on M059J lineage, and increased in 50 % and 42.6 % when comparing M059J-irradiated to irradiated U-138 MG and U-251 MG cells, respectively. Ethidium bromide uptake was significantly increased in 104 % and 77.8 % when comparing M059J to U-138 MG and U-251MG, respectively. Finally, the selective P2X7R antagonist A740003 significantly decreased the cell death caused by irradiation. We provide novel evidence indicating that M059J human glioma cell line is ATP-P2X7R sensitive, pointing out the relevance of the purinergic P2X7R on glioma radiosensitivity.

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Acknowledgments

This work was supported by Fundação de Amparo à Pesquisa do Estado do Rio Grande do Sul (FAPERGS) [Proc. 1016960], Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) and Financiadora de Estudos e Projetos (FINEP) [(PUCRSINFRA) # 01.11.0014-00]. We thank for Dr. Carlos Henrique Barrios for the gift of the cell line, Dr. Robson Coutinho-Silva for the gift of compounds and Julia Cisilotto for help with the cell counting.

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Authors and Affiliations

  1. Laboratório de Farmacologia Aplicada, PUCRS, Av. Ipiranga, 6681, Prédio 12C, Sala 148, Partenon, Porto Alegre, RS, Brazil, 90619-900

    Marina Petersen Gehring, Rafael Fernandes Zanin & Fernanda Bueno Morrone

  2. Instituto Nacional de Ciência e Tecnologia Translacional em Medicina (INCT-TM), PUCRS, Av. Ipiranga, 6681, Prédio 12C, Sala 172, Partenon, Porto Alegre, RS, Brazil, 90619-900

    Talita Carneiro Brandão Pereira & Maurício Reis Bogo

  3. Serviço de Radioterapia, Hospital São Lucas, Av. Ipiranga, 6690, Azenha, Porto Alegre, RS, Brazil, 90160-090

    Magali Carvalho Borges & Aroldo Braga Filho

  4. Departamento de Bioquímica, ICBS, UFRGS, Rua Ramiro Barcelos, 2600—Anexo. Santa Cecília, Porto Alegre, RS, Brazil, 90035-000

    Ana Maria Oliveira Battastini

  5. Laboratório de Sinalização e Plasticidade Celular, Departamento de Biofísica, Instituto de Biociências, UFRGS, Av. Bento Gonçalves, 9500, Prédio 43431, Sala 107, Agronomia, Porto Alegre, RS, Brazil, 91501-970

    Guido Lenz

  6. Instituto de Toxicologia e Farmacologia, PUCRS, Av. Ipiranga, 6681, Prédio 12C, Sala 101, Partenon, Porto Alegre, RS, Brazil, 90619-900

    Maria Martha Campos

  7. Laboratório de Biologia Genômica e Molecular, PUCRS, Av. Ipiranga, 6681, Prédio 12C, Sala 172, Partenon, Porto Alegre, RS, Brazil, 90160-090

    Talita Carneiro Brandão Pereira & Maurício Reis Bogo

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  1. Marina Petersen Gehring
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  2. Talita Carneiro Brandão Pereira
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  3. Rafael Fernandes Zanin
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  8. Guido Lenz
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  9. Maria Martha Campos
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  10. Fernanda Bueno Morrone
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Correspondence to Fernanda Bueno Morrone.

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Gehring, M.P., Pereira, T.C.B., Zanin, R.F. et al. P2X7 receptor activation leads to increased cell death in a radiosensitive human glioma cell line. Purinergic Signalling 8, 729–739 (2012). https://doi.org/10.1007/s11302-012-9319-2

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  • DOI: https://doi.org/10.1007/s11302-012-9319-2

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