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Lactic Acidosis in the Presence of Glucose Diminishes Warburg Effect in Lung Adenocarcinoma Cells

Lactic acidosis (3 to 40 mM, pH < 6.9) is a condition found in solid tumors because tumor cells have a high rate of glucose consumption and lactate production even in the presence of oxygen; nevertheless, the microenvironment might still provide a sufficient glucose supply. Lactic acidosis has be...

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Autores principales: Prado-Garcia, Heriberto, Campa-Higareda, Andrea, Romero-Garcia, Susana
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Frontiers Media S.A. 2020
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7303336/
https://www.ncbi.nlm.nih.gov/pubmed/32596143
http://dx.doi.org/10.3389/fonc.2020.00807
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author Prado-Garcia, Heriberto
Campa-Higareda, Andrea
Romero-Garcia, Susana
author_facet Prado-Garcia, Heriberto
Campa-Higareda, Andrea
Romero-Garcia, Susana
author_sort Prado-Garcia, Heriberto
collection PubMed
description Lactic acidosis (3 to 40 mM, pH < 6.9) is a condition found in solid tumors because tumor cells have a high rate of glucose consumption and lactate production even in the presence of oxygen; nevertheless, the microenvironment might still provide a sufficient glucose supply. Lactic acidosis has been proposed to shift metabolism from aerobic glycolysis toward oxidative phosphorylation (OXPHOS). We tested if lung tumor cells cultured under lactic acidosis shift their metabolism from glycolysis to OXPHOS by consuming extracellular lactate, increasing growth rate. We analyzed lung adenocarcinoma (A-549, A-427) cell lines and non-transformed fibroblast cells (MRC-5), which were cultured using RPMI-1640 medium initially containing lactate (2 mM) and glucose (10 mM), at pH 7.2 or 6.2 and oxygen tension 21% O(2) (normoxia) or 2% O(2) (hypoxia). We obtained growth curves, as well as glucose consumption and lactate production rates (measured during exponential growth) for each cell line. HIF-1α (Hypoxia-inducible factor 1 α), CS (citrate synthase) and AMPK (AMP-activated protein kinase) transcript levels were analyzed using RT-qPCR. By flow cytometry, we determined: (a) expression of glucose transporters (GLUT)1 and 4; (b) lactate transporters (MCT)1 and 4; (c) cell cycle profile, and (d) protein levels of HIF-1α, total and phosphorylated AMPK (pAMPK). Mitochondrial functionality was evaluated by measuring O(2) consumption in tumor cells using polarography and a Clark-type electrode. Tumor and non-transformed cells used both aerobic glycolysis and OXPHOS for obtaining energy. As of 48 h of culture, lactate levels ranged from (4.5–14 mM), thus forming a lactic environment. Lactic acidosis diminished GLUT1/GLUT4 expression and glucose consumption in A-549, but not in A-427 cells, and induced differential expression of HIF-1α, AMPK, and CS transcripts. A-427 cells increased pAMPK and HIF-1α levels and shifted their metabolism increasing OXPHOS; thus supporting cell growth. Conversely, A-549 cells increased HIF-1α protein levels, but did not activate AMPK and diminished OXPHOS. A-549 cells survived by arresting cells in G1-phase. Our findings show that lactic acidosis diminishes Warburg effect in tumor cells, but this change does not necessarily promote a shift to OXPHOS. Hence, lung adenocarcinomas show a differential metabolic response even when they are under the same microenvironmental conditions.
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spelling pubmed-73033362020-06-26 Lactic Acidosis in the Presence of Glucose Diminishes Warburg Effect in Lung Adenocarcinoma Cells Prado-Garcia, Heriberto Campa-Higareda, Andrea Romero-Garcia, Susana Front Oncol Oncology Lactic acidosis (3 to 40 mM, pH < 6.9) is a condition found in solid tumors because tumor cells have a high rate of glucose consumption and lactate production even in the presence of oxygen; nevertheless, the microenvironment might still provide a sufficient glucose supply. Lactic acidosis has been proposed to shift metabolism from aerobic glycolysis toward oxidative phosphorylation (OXPHOS). We tested if lung tumor cells cultured under lactic acidosis shift their metabolism from glycolysis to OXPHOS by consuming extracellular lactate, increasing growth rate. We analyzed lung adenocarcinoma (A-549, A-427) cell lines and non-transformed fibroblast cells (MRC-5), which were cultured using RPMI-1640 medium initially containing lactate (2 mM) and glucose (10 mM), at pH 7.2 or 6.2 and oxygen tension 21% O(2) (normoxia) or 2% O(2) (hypoxia). We obtained growth curves, as well as glucose consumption and lactate production rates (measured during exponential growth) for each cell line. HIF-1α (Hypoxia-inducible factor 1 α), CS (citrate synthase) and AMPK (AMP-activated protein kinase) transcript levels were analyzed using RT-qPCR. By flow cytometry, we determined: (a) expression of glucose transporters (GLUT)1 and 4; (b) lactate transporters (MCT)1 and 4; (c) cell cycle profile, and (d) protein levels of HIF-1α, total and phosphorylated AMPK (pAMPK). Mitochondrial functionality was evaluated by measuring O(2) consumption in tumor cells using polarography and a Clark-type electrode. Tumor and non-transformed cells used both aerobic glycolysis and OXPHOS for obtaining energy. As of 48 h of culture, lactate levels ranged from (4.5–14 mM), thus forming a lactic environment. Lactic acidosis diminished GLUT1/GLUT4 expression and glucose consumption in A-549, but not in A-427 cells, and induced differential expression of HIF-1α, AMPK, and CS transcripts. A-427 cells increased pAMPK and HIF-1α levels and shifted their metabolism increasing OXPHOS; thus supporting cell growth. Conversely, A-549 cells increased HIF-1α protein levels, but did not activate AMPK and diminished OXPHOS. A-549 cells survived by arresting cells in G1-phase. Our findings show that lactic acidosis diminishes Warburg effect in tumor cells, but this change does not necessarily promote a shift to OXPHOS. Hence, lung adenocarcinomas show a differential metabolic response even when they are under the same microenvironmental conditions. Frontiers Media S.A. 2020-06-12 /pmc/articles/PMC7303336/ /pubmed/32596143 http://dx.doi.org/10.3389/fonc.2020.00807 Text en Copyright © 2020 Prado-Garcia, Campa-Higareda and Romero-Garcia. http://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.
spellingShingle Oncology
Prado-Garcia, Heriberto
Campa-Higareda, Andrea
Romero-Garcia, Susana
Lactic Acidosis in the Presence of Glucose Diminishes Warburg Effect in Lung Adenocarcinoma Cells
title Lactic Acidosis in the Presence of Glucose Diminishes Warburg Effect in Lung Adenocarcinoma Cells
title_full Lactic Acidosis in the Presence of Glucose Diminishes Warburg Effect in Lung Adenocarcinoma Cells
title_fullStr Lactic Acidosis in the Presence of Glucose Diminishes Warburg Effect in Lung Adenocarcinoma Cells
title_full_unstemmed Lactic Acidosis in the Presence of Glucose Diminishes Warburg Effect in Lung Adenocarcinoma Cells
title_short Lactic Acidosis in the Presence of Glucose Diminishes Warburg Effect in Lung Adenocarcinoma Cells
title_sort lactic acidosis in the presence of glucose diminishes warburg effect in lung adenocarcinoma cells
topic Oncology
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7303336/
https://www.ncbi.nlm.nih.gov/pubmed/32596143
http://dx.doi.org/10.3389/fonc.2020.00807
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