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Evaluation of mitochondrial respiratory function in highly glycolytic glioma cells reveals low ADP phosphorylation in relation to oxidative capacity

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Abstract

High-grade gliomas are aggressive and intensely glycolytic tumors. In the present study, we evaluated the mitochondrial respiratory function of glioma cells (T98G and U-87MG) and fresh human glioblastoma (GBM) tissue. To this end, measurements of oxygen consumption rate (OCR) were performed under various experimental conditions. The OCR of T98G and U-87MG cells was well coupled to ADP phosphorylation based on the ratio of ATP produced per oxygen consumed of ~2.5. In agreement, the basal OCR of GBM tissue was also partially associated with ADP phosphorylation. The basal respiration of intact T98G and U-87MG cells was not limited by the supply of endogenous substrates, as indicated by the increased OCR in response to a protonophore. These cells also displayed a high affinity for oxygen, as evidenced by the values of the partial pressure of oxygen when respiration is half maximal (p 50). In permeabilized glioma cells, ADP-stimulated OCR was only approximately 50% of that obtained in the presence of protonophore, revealing a significant limitation in oxidative phosphorylation (OXPHOS) relative to the activity of the electron transport system (ETS). This characteristic was maintained when the cells were grown under low glucose conditions. Flux control coefficient analyses demonstrated that the impaired OXPHOS was associated with the function of both mitochondrial ATP synthase and the adenine nucleotide translocator, but not the phosphate carrier. Altogether, these data indicate that the availability and metabolism of respiratory substrates and mitochondrial ETS are preserved in T98G and U-87MG glioma cells even though these cells possess a relatively restrained OXPHOS capability.

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Acknowledgements

This work was financially supported by grants from the São Paulo Research Foundation (FAPESP, #11/50400-0 and #13/02618-1) and the Brazilian National Council for Scientific and Technological Development (CNPq, #150546/2015-7 to TRF). ER-S and JSR are recipients of CNPq and CAPES fellowships, respectively.

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Authors and Affiliations

  1. Departamento de Patologia Clínica, Faculdade de Ciências Médicas, Universidade Estadual de Campinas (UNICAMP), Campinas, SP, 13083-887, Brazil

    Erika Rodrigues-Silva, Edilene S. Siqueira-Santos, Juliana S. Ruas, Tiago R. Figueira & Roger F. Castilho

  2. Departamento de Anatomia Patológica, Faculdade de Ciências Médicas, Universidade Estadual de Campinas (UNICAMP), Campinas, SP, Brazil

    Raffaela S. Ignarro & Fábio Rogério

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  1. Erika Rodrigues-Silva
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  2. Edilene S. Siqueira-Santos
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  4. Raffaela S. Ignarro
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  5. Tiago R. Figueira
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Correspondence to Roger F. Castilho.

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Ethical approval

All procedures performed in studies involving human participants were in accordance with the ethical standards of the institutional and/or national research committee and with the 1964 Helsinki declaration and its later amendments or comparable ethical standards. UNICAMP ethics committee approval protocol number was 1164/2011.

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Informed consent was obtained from all individual participants included in the study.

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All applicable international, national, and/or institutional guidelines for the care and use of animals were followed. Experimental procedures involving rats were approved by the local Committee for Ethics in Animal Experimentation (protocol number CEUA 2534-1).

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Rodrigues-Silva, E., Siqueira-Santos, E.S., Ruas, J.S. et al. Evaluation of mitochondrial respiratory function in highly glycolytic glioma cells reveals low ADP phosphorylation in relation to oxidative capacity. J Neurooncol 133, 519–529 (2017). https://doi.org/10.1007/s11060-017-2482-0

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