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Krebs Cycle Intermediates Protective against Oxidative Stress by Modulating the Level of Reactive Oxygen Species in Neuronal HT22 Cells

View Article: PubMed Central - PubMed

ABSTRACT

Krebs cycle intermediates (KCIs) are reported to function as energy substrates in mitochondria and to exert antioxidants effects on the brain. The present study was designed to identify which KCIs are effective neuroprotective compounds against oxidative stress in neuronal cells. Here we found that pyruvate, oxaloacetate, and α-ketoglutarate, but not lactate, citrate, iso-citrate, succinate, fumarate, or malate, protected HT22 cells against hydrogen peroxide-mediated toxicity. These three intermediates reduced the production of hydrogen peroxide-activated reactive oxygen species, measured in terms of 2′,7′-dichlorofluorescein diacetate fluorescence. In contrast, none of the KCIs—used at 1 mM—protected against cell death induced by high concentrations of glutamate—another type of oxidative stress-induced neuronal cell death. Because these protective KCIs did not have any toxic effects (at least up to 10 mM), they have potential use for therapeutic intervention against chronic neurodegenerative diseases.

No MeSH data available.


(A) Compounds involved in the Krebs cycle and (B) common chemical structure of neuroprotective KCIs (PA, OAA, and AKG). Note that PA, OAA, and AKG have the common chemical structure of the α-keto acid group—indicated by the dotted circle—with which molecules can directly react with H2O2.
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antioxidants-06-00021-f006: (A) Compounds involved in the Krebs cycle and (B) common chemical structure of neuroprotective KCIs (PA, OAA, and AKG). Note that PA, OAA, and AKG have the common chemical structure of the α-keto acid group—indicated by the dotted circle—with which molecules can directly react with H2O2.

Mentions: Here, we found that the α-keto acid group-containing KCIs (PA, OAA, and AKG) could protect neurons against H2O2, possibly through direct interaction with H2O2, although we did not provide direct evidence for this chemical reaction itself. Because DCFH-DA is hydrolyzed by cytosolic esterase and is activated by ROS in the cytoplasm, the inhibition of ROS increase by PA, OAA, and AKG could be due to an H2O2-scavenging effect of them inside the cells. The conclusion of this study is illustrated in Figure 6A and B. PA, OAA, and AKG were neuroprotective, but not the other KCIs. For example, LA and MA— putative neuronal energy substrates that may produce protective KCIs through their metabolism [1,2]—did not protect the cells (Figure 4 and Figure 5). AKG may have other protective effects than α-keto acids such as PA and OAA. AKG at 1mM protected the cells against H2O2 (Figure 1) without causing a significant reduction in ROS levels (Figure 3). Because AKG is reported to activate the degradation of HIF-1α subunit and reduce the expression of downstream enzymes [6,7], reduction of this pathway may have been involved in the protective effects of AKG. FA may be another exceptional KCI. FA at 10 mM protected the cells against Glu-mediated cytotoxicity (Figure 5), although the other KCIs did not do so. Because FA is reported to activate the Nrf2 pathway and induce phase-2 enzymes [18], the activation of this pathway may have been involved in the FA-induced protection.


Krebs Cycle Intermediates Protective against Oxidative Stress by Modulating the Level of Reactive Oxygen Species in Neuronal HT22 Cells
(A) Compounds involved in the Krebs cycle and (B) common chemical structure of neuroprotective KCIs (PA, OAA, and AKG). Note that PA, OAA, and AKG have the common chemical structure of the α-keto acid group—indicated by the dotted circle—with which molecules can directly react with H2O2.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC5384184&req=5

antioxidants-06-00021-f006: (A) Compounds involved in the Krebs cycle and (B) common chemical structure of neuroprotective KCIs (PA, OAA, and AKG). Note that PA, OAA, and AKG have the common chemical structure of the α-keto acid group—indicated by the dotted circle—with which molecules can directly react with H2O2.
Mentions: Here, we found that the α-keto acid group-containing KCIs (PA, OAA, and AKG) could protect neurons against H2O2, possibly through direct interaction with H2O2, although we did not provide direct evidence for this chemical reaction itself. Because DCFH-DA is hydrolyzed by cytosolic esterase and is activated by ROS in the cytoplasm, the inhibition of ROS increase by PA, OAA, and AKG could be due to an H2O2-scavenging effect of them inside the cells. The conclusion of this study is illustrated in Figure 6A and B. PA, OAA, and AKG were neuroprotective, but not the other KCIs. For example, LA and MA— putative neuronal energy substrates that may produce protective KCIs through their metabolism [1,2]—did not protect the cells (Figure 4 and Figure 5). AKG may have other protective effects than α-keto acids such as PA and OAA. AKG at 1mM protected the cells against H2O2 (Figure 1) without causing a significant reduction in ROS levels (Figure 3). Because AKG is reported to activate the degradation of HIF-1α subunit and reduce the expression of downstream enzymes [6,7], reduction of this pathway may have been involved in the protective effects of AKG. FA may be another exceptional KCI. FA at 10 mM protected the cells against Glu-mediated cytotoxicity (Figure 5), although the other KCIs did not do so. Because FA is reported to activate the Nrf2 pathway and induce phase-2 enzymes [18], the activation of this pathway may have been involved in the FA-induced protection.

View Article: PubMed Central - PubMed

ABSTRACT

Krebs cycle intermediates (KCIs) are reported to function as energy substrates in mitochondria and to exert antioxidants effects on the brain. The present study was designed to identify which KCIs are effective neuroprotective compounds against oxidative stress in neuronal cells. Here we found that pyruvate, oxaloacetate, and α-ketoglutarate, but not lactate, citrate, iso-citrate, succinate, fumarate, or malate, protected HT22 cells against hydrogen peroxide-mediated toxicity. These three intermediates reduced the production of hydrogen peroxide-activated reactive oxygen species, measured in terms of 2′,7′-dichlorofluorescein diacetate fluorescence. In contrast, none of the KCIs—used at 1 mM—protected against cell death induced by high concentrations of glutamate—another type of oxidative stress-induced neuronal cell death. Because these protective KCIs did not have any toxic effects (at least up to 10 mM), they have potential use for therapeutic intervention against chronic neurodegenerative diseases.

No MeSH data available.