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The spatial organization of proton and lactate transport in a rat brain tumor.

Grillon E, Farion R, Fablet K, De Waard M, Tse CM, Donowitz M, Rémy C, Coles JA - PLoS ONE (2011)

Bottom Line: In contrast, MCT4 and the carbonic anhydrase CAIX, which are associated with hypoxia, were not significantly upregulated in the rim.The spatial distribution of MCT4 was highly correlated with that of CAIX, suggesting that their expression is regulated by the same factors.Since protons extruded by NHE1 diffuse away through extracellular clefts, NHE1 requires a continuous source of intracellular protons.

View Article: PubMed Central - PubMed

Affiliation: Unit 836, Institut National de la Santé et de la Recherche Médicale, La Tronche, Isère, France.

ABSTRACT
Tumors create a heterogeneous acidic microenvironment which assists their growth and which must be taken into account in the design of drugs and their delivery. In addition, the acidic extracellular pH (pHe) is itself exploited in several experimental techniques for drug delivery. The way the acidity is created is not clear. We report here the spatial organization of key proton-handling proteins in C6 gliomas in rat brain. The mean profiles across the tumor rim of the Na+/H+ exchanger NHE1, and the lactate-H+ cotransporter MCT1, both showed peaks. NHE1, which is important for extension and migration of cells in vitro, showed a peak 1.55 times higher than in extratumoural tissue at 0.33 mm from the edge. MCT1 had a broader peak, further into the tumor (maximum 1.76 fold at 1.0 mm from the edge). In contrast, MCT4 and the carbonic anhydrase CAIX, which are associated with hypoxia, were not significantly upregulated in the rim. The spatial distribution of MCT4 was highly correlated with that of CAIX, suggesting that their expression is regulated by the same factors. Since protons extruded by NHE1 diffuse away through extracellular clefts, NHE1 requires a continuous source of intracellular protons. From the stoichiometries of metabolic pathways that produce or consume H+, and the greater availability of glucose compared to oxygen in most parts of a tumor, we support the classic view that most of the net proton efflux from C6 gliomas originates in glycolytic formation of lactate and H+ inside the tumor, but add that some lactate is taken up into cells in the rim on MCT1, and some lactate diffuses away, leaving its associated protons available to re-enter cells for extrusion on NHE1. Therapeutic inhibition of NHE1, MCT1 or CAIX is predicted to affect different parts of a tumor.

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Roles of MCT1 and NHE1 in C6 gliomas.(A) Stoichiometry of the main pathways of glucose metabolism. Note that the protons associated with lactate production are exported by cotransport with lactate. The consequences of CO2 production are potentially more variable than shown. (B) Scheme of fluxes of glucose, oxygen, lactate and H+ in the glioma rim, suggested by the profiles of NHE1 and MCT1. Some of the lactate exported from deeper, hypoxic regions is taken up on MCT1 in the rim of the tumor and oxidized to CO2. Ground colors indicate pHe: red is acid, green is alkaline.
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pone-0017416-g004: Roles of MCT1 and NHE1 in C6 gliomas.(A) Stoichiometry of the main pathways of glucose metabolism. Note that the protons associated with lactate production are exported by cotransport with lactate. The consequences of CO2 production are potentially more variable than shown. (B) Scheme of fluxes of glucose, oxygen, lactate and H+ in the glioma rim, suggested by the profiles of NHE1 and MCT1. Some of the lactate exported from deeper, hypoxic regions is taken up on MCT1 in the rim of the tumor and oxidized to CO2. Ground colors indicate pHe: red is acid, green is alkaline.

Mentions: Where do the protons extruded by NHE1 in the rim of the tumor come from? Since an extracellular accumulation of H+ will dissipate by diffusion of H+ through extracellular space [10], [28], a tumor, as a whole, can only maintain an acidic pHe by constantly extruding protons (or some equivalent process, such as constantly taking up HCO3−). Any cyclic process whereby protons enter the cell from the extracellular space and are then extruded at almost the same place would not change pHe: the protons must be generated within the cell (or come from other cells – see below). A major source of intracellular H+ ions is glycolytic production of lactate. Lactic acid and its precursor, pyruvic acid, are >99% unprotonated at physiological pHs [69]; the stoichiometry of glycolysis for these unprotonated forms is shown in Fig. 4A and it is seen that, in the steady state (when the concentrations of NADH, NAD+, ADP, ATP etc. remain constant), lactate ions and H+ ions are produced in equal numbers. Extracellular lactate diffuses away through extracellular clefts, so an outwardly directed gradient of [lactate−] is created which drives the equimolar cotransport of H+ [31]. Since each lactate− crosses the cell membrane in association with a proton, glycolysis does not subject the cell to continuous loading with internally-generated protons, and is not a direct source of protons for other transporters such as NHE1.


The spatial organization of proton and lactate transport in a rat brain tumor.

Grillon E, Farion R, Fablet K, De Waard M, Tse CM, Donowitz M, Rémy C, Coles JA - PLoS ONE (2011)

Roles of MCT1 and NHE1 in C6 gliomas.(A) Stoichiometry of the main pathways of glucose metabolism. Note that the protons associated with lactate production are exported by cotransport with lactate. The consequences of CO2 production are potentially more variable than shown. (B) Scheme of fluxes of glucose, oxygen, lactate and H+ in the glioma rim, suggested by the profiles of NHE1 and MCT1. Some of the lactate exported from deeper, hypoxic regions is taken up on MCT1 in the rim of the tumor and oxidized to CO2. Ground colors indicate pHe: red is acid, green is alkaline.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3044751&req=5

pone-0017416-g004: Roles of MCT1 and NHE1 in C6 gliomas.(A) Stoichiometry of the main pathways of glucose metabolism. Note that the protons associated with lactate production are exported by cotransport with lactate. The consequences of CO2 production are potentially more variable than shown. (B) Scheme of fluxes of glucose, oxygen, lactate and H+ in the glioma rim, suggested by the profiles of NHE1 and MCT1. Some of the lactate exported from deeper, hypoxic regions is taken up on MCT1 in the rim of the tumor and oxidized to CO2. Ground colors indicate pHe: red is acid, green is alkaline.
Mentions: Where do the protons extruded by NHE1 in the rim of the tumor come from? Since an extracellular accumulation of H+ will dissipate by diffusion of H+ through extracellular space [10], [28], a tumor, as a whole, can only maintain an acidic pHe by constantly extruding protons (or some equivalent process, such as constantly taking up HCO3−). Any cyclic process whereby protons enter the cell from the extracellular space and are then extruded at almost the same place would not change pHe: the protons must be generated within the cell (or come from other cells – see below). A major source of intracellular H+ ions is glycolytic production of lactate. Lactic acid and its precursor, pyruvic acid, are >99% unprotonated at physiological pHs [69]; the stoichiometry of glycolysis for these unprotonated forms is shown in Fig. 4A and it is seen that, in the steady state (when the concentrations of NADH, NAD+, ADP, ATP etc. remain constant), lactate ions and H+ ions are produced in equal numbers. Extracellular lactate diffuses away through extracellular clefts, so an outwardly directed gradient of [lactate−] is created which drives the equimolar cotransport of H+ [31]. Since each lactate− crosses the cell membrane in association with a proton, glycolysis does not subject the cell to continuous loading with internally-generated protons, and is not a direct source of protons for other transporters such as NHE1.

Bottom Line: In contrast, MCT4 and the carbonic anhydrase CAIX, which are associated with hypoxia, were not significantly upregulated in the rim.The spatial distribution of MCT4 was highly correlated with that of CAIX, suggesting that their expression is regulated by the same factors.Since protons extruded by NHE1 diffuse away through extracellular clefts, NHE1 requires a continuous source of intracellular protons.

View Article: PubMed Central - PubMed

Affiliation: Unit 836, Institut National de la Santé et de la Recherche Médicale, La Tronche, Isère, France.

ABSTRACT
Tumors create a heterogeneous acidic microenvironment which assists their growth and which must be taken into account in the design of drugs and their delivery. In addition, the acidic extracellular pH (pHe) is itself exploited in several experimental techniques for drug delivery. The way the acidity is created is not clear. We report here the spatial organization of key proton-handling proteins in C6 gliomas in rat brain. The mean profiles across the tumor rim of the Na+/H+ exchanger NHE1, and the lactate-H+ cotransporter MCT1, both showed peaks. NHE1, which is important for extension and migration of cells in vitro, showed a peak 1.55 times higher than in extratumoural tissue at 0.33 mm from the edge. MCT1 had a broader peak, further into the tumor (maximum 1.76 fold at 1.0 mm from the edge). In contrast, MCT4 and the carbonic anhydrase CAIX, which are associated with hypoxia, were not significantly upregulated in the rim. The spatial distribution of MCT4 was highly correlated with that of CAIX, suggesting that their expression is regulated by the same factors. Since protons extruded by NHE1 diffuse away through extracellular clefts, NHE1 requires a continuous source of intracellular protons. From the stoichiometries of metabolic pathways that produce or consume H+, and the greater availability of glucose compared to oxygen in most parts of a tumor, we support the classic view that most of the net proton efflux from C6 gliomas originates in glycolytic formation of lactate and H+ inside the tumor, but add that some lactate is taken up into cells in the rim on MCT1, and some lactate diffuses away, leaving its associated protons available to re-enter cells for extrusion on NHE1. Therapeutic inhibition of NHE1, MCT1 or CAIX is predicted to affect different parts of a tumor.

Show MeSH
Related in: MedlinePlus