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

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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.

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Protection of HT22 against (A) H2O2 and (B) Glu by pyruvate (PA), oxaloacetate (OAA), and α-ketoglutarate (AKG). (A) Protective effects of 1mM PA, OAA, or AKG against various concentrations of H2O2. (B) No protective effect of 1 mM PA, OAA, or AKG against high concentrations of Glu was found. White bars indicate H2O2 or Glu alone; and gray bars H2O2 + KCI (PA, OAA, or AKG). Values, presented as a percentage of the control MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide) value (obtained in the absence of glutamate), are given as the mean ± SD (n = 4). * significantly different (p < 0.05) from samples without a KCI. KCI: Krebs cycle intermediate.
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antioxidants-06-00021-f001: Protection of HT22 against (A) H2O2 and (B) Glu by pyruvate (PA), oxaloacetate (OAA), and α-ketoglutarate (AKG). (A) Protective effects of 1mM PA, OAA, or AKG against various concentrations of H2O2. (B) No protective effect of 1 mM PA, OAA, or AKG against high concentrations of Glu was found. White bars indicate H2O2 or Glu alone; and gray bars H2O2 + KCI (PA, OAA, or AKG). Values, presented as a percentage of the control MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide) value (obtained in the absence of glutamate), are given as the mean ± SD (n = 4). * significantly different (p < 0.05) from samples without a KCI. KCI: Krebs cycle intermediate.

Mentions: We used HT22 cells—a neuronal cell line derived from a mouse hippocampus—as a model for oxidative cell damage. Treatment of the cells with 100, 200, or 500 μM H2O2 (Figure 1A) or with 5, 10, or 20 mM Glu (Figure 1B) induced cell death by oxidative stress within 24 h. Pre (1 h)-treatment of the cells with the KCIs at 1 mM protected the cells from the toxic effects of H2O2 (Figure 1A), whereas these KCIs were not—or very little if they were—protective against Glu toxicity (Figure 1B). These KCIs were not toxic to HT22 cells up to 10 mM, but they caused a small but significant reduction in cell survival when used at 20–50 mM (Figure 2). These results suggest that PA, OAA, and AKG could protect neuronal cells against H2O2, but not against Glu.


Krebs Cycle Intermediates Protective against Oxidative Stress by Modulating the Level of Reactive Oxygen Species in Neuronal HT22 Cells
Protection of HT22 against (A) H2O2 and (B) Glu by pyruvate (PA), oxaloacetate (OAA), and α-ketoglutarate (AKG). (A) Protective effects of 1mM PA, OAA, or AKG against various concentrations of H2O2. (B) No protective effect of 1 mM PA, OAA, or AKG against high concentrations of Glu was found. White bars indicate H2O2 or Glu alone; and gray bars H2O2 + KCI (PA, OAA, or AKG). Values, presented as a percentage of the control MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide) value (obtained in the absence of glutamate), are given as the mean ± SD (n = 4). * significantly different (p < 0.05) from samples without a KCI. KCI: Krebs cycle intermediate.
© Copyright Policy - open-access
Related In: Results  -  Collection

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

antioxidants-06-00021-f001: Protection of HT22 against (A) H2O2 and (B) Glu by pyruvate (PA), oxaloacetate (OAA), and α-ketoglutarate (AKG). (A) Protective effects of 1mM PA, OAA, or AKG against various concentrations of H2O2. (B) No protective effect of 1 mM PA, OAA, or AKG against high concentrations of Glu was found. White bars indicate H2O2 or Glu alone; and gray bars H2O2 + KCI (PA, OAA, or AKG). Values, presented as a percentage of the control MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide) value (obtained in the absence of glutamate), are given as the mean ± SD (n = 4). * significantly different (p < 0.05) from samples without a KCI. KCI: Krebs cycle intermediate.
Mentions: We used HT22 cells—a neuronal cell line derived from a mouse hippocampus—as a model for oxidative cell damage. Treatment of the cells with 100, 200, or 500 μM H2O2 (Figure 1A) or with 5, 10, or 20 mM Glu (Figure 1B) induced cell death by oxidative stress within 24 h. Pre (1 h)-treatment of the cells with the KCIs at 1 mM protected the cells from the toxic effects of H2O2 (Figure 1A), whereas these KCIs were not—or very little if they were—protective against Glu toxicity (Figure 1B). These KCIs were not toxic to HT22 cells up to 10 mM, but they caused a small but significant reduction in cell survival when used at 20–50 mM (Figure 2). These results suggest that PA, OAA, and AKG could protect neuronal cells against H2O2, but not against Glu.

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 &alpha;-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&prime;,7&prime;-dichlorofluorescein diacetate fluorescence. In contrast, none of the KCIs&mdash;used at 1 mM&mdash;protected against cell death induced by high concentrations of glutamate&mdash;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.


Related in: MedlinePlus