Limits...
Competitive HIF Prolyl Hydroxylase Inhibitors Show Protection against Oxidative Stress by a Mechanism Partially Dependent on Glycolysis.

Bergström AL, Fog K, Sager TN, Bruun AT, Thirstrup K - ISRN Neurosci (2013)

Bottom Line: In the present study, we compared competitive and noncompetitive HPH-inhibitor compounds in two different cell types (SH-SY5Y and PC12).Both competitive and non-competitive HPH inhibitors protected the cells against 6-OHDA induced oxidative stress.In addition, the protective effect of a specific HPH inhibitor was partially preserved when the cells were serum starved and exposed to 2-deoxyglucose, an inhibitor of glycolysis, indicating that other processes than restoring energy supply could be important for the HIF-mediated cytoprotection.

View Article: PubMed Central - PubMed

Affiliation: Department of Neurodegeneration, Lundbeck A/S, Ottiliavej 9, 2500 Valby, Denmark.

ABSTRACT
The hypoxia inducible factor 1 (HIF-1) is a central transcription factor involved in the cellular and molecular adaptation to hypoxia and low glucose supply. The level of HIF-1 is to a large degree regulated by the HIF prolyl hydroxylase enzymes (HPHs) belonging to the Fe(II) and 2-oxoglutarate-dependent dioxygenase superfamily. In the present study, we compared competitive and noncompetitive HPH-inhibitor compounds in two different cell types (SH-SY5Y and PC12). Although the competitive HPH-inhibitor compounds were found to be pharmacologically more potent than the non-competitive compounds at inhibiting HPH2 and HPH1, this was not translated into the cellular effects of the compounds, where the non-competitive inhibitors were actually more potent than the competitive in stabilizing and translocatingHIF1 α to the nucleus (quantified with Cellomics ArrayScan technology). This could be explained by the high cellular concentrations of the cofactor 2-oxoglutarate (2-OG) as the competitive inhibitors act by binding to the 2-OG site of the HPH enzymes. Both competitive and non-competitive HPH inhibitors protected the cells against 6-OHDA induced oxidative stress. In addition, the protective effect of a specific HPH inhibitor was partially preserved when the cells were serum starved and exposed to 2-deoxyglucose, an inhibitor of glycolysis, indicating that other processes than restoring energy supply could be important for the HIF-mediated cytoprotection.

No MeSH data available.


Related in: MedlinePlus

Neuroprotective effect of HPH inhibitors. (a) ATP-levels in SH-SY5Y cells grown overnight in starvation media, pretreated for 3 hours with 25 μM of FG41 (a), DFO (b), CpdA (c), or JNJ (d) followed by 6-OHDA treatment for 3 hours. (e) Mitochondrial membrane potential in SH-SY5Y cells treated with Cpd A (10 or 25 μM) for 3 hours followed by 3 hours treatment with 50 μM 6-OHDA. Asterisks indicate levels significantly increased (t-test) as compared to control.
© Copyright Policy - open-access
Related In: Results  -  Collection


getmorefigures.php?uid=PMC4061615&req=5

fig4: Neuroprotective effect of HPH inhibitors. (a) ATP-levels in SH-SY5Y cells grown overnight in starvation media, pretreated for 3 hours with 25 μM of FG41 (a), DFO (b), CpdA (c), or JNJ (d) followed by 6-OHDA treatment for 3 hours. (e) Mitochondrial membrane potential in SH-SY5Y cells treated with Cpd A (10 or 25 μM) for 3 hours followed by 3 hours treatment with 50 μM 6-OHDA. Asterisks indicate levels significantly increased (t-test) as compared to control.

Mentions: Having established that both the competitive and the non-competitive HPH-inhibitor compounds are able to induce an HIF-1α response in the SH-SY5Y cells, we investigated the potential of these compounds to protect against oxidative stress. To model the cellular dysfunction of dopaminergic neurons in Parkinson's disease, where oxidative stress and mitochondrial dysfunction are cellular hallmarks [10], we treated SH-SY5Y cells with 6-OHDA. Treatment with 6-OHDA generates H2O2 and this oxidative stress leads to collapse of the mitochondrial membrane potential (ΔΨmit) and a reduction in cellular ATP production [11]. We found that 3 hours of 6-OHDA treatment was enough to induce a concentration-dependent collapse of both ΔΨmit and ATP-production, Figure 4, without reducing the cell numbers (not shown).


Competitive HIF Prolyl Hydroxylase Inhibitors Show Protection against Oxidative Stress by a Mechanism Partially Dependent on Glycolysis.

Bergström AL, Fog K, Sager TN, Bruun AT, Thirstrup K - ISRN Neurosci (2013)

Neuroprotective effect of HPH inhibitors. (a) ATP-levels in SH-SY5Y cells grown overnight in starvation media, pretreated for 3 hours with 25 μM of FG41 (a), DFO (b), CpdA (c), or JNJ (d) followed by 6-OHDA treatment for 3 hours. (e) Mitochondrial membrane potential in SH-SY5Y cells treated with Cpd A (10 or 25 μM) for 3 hours followed by 3 hours treatment with 50 μM 6-OHDA. Asterisks indicate levels significantly increased (t-test) as compared to control.
© Copyright Policy - open-access
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4061615&req=5

fig4: Neuroprotective effect of HPH inhibitors. (a) ATP-levels in SH-SY5Y cells grown overnight in starvation media, pretreated for 3 hours with 25 μM of FG41 (a), DFO (b), CpdA (c), or JNJ (d) followed by 6-OHDA treatment for 3 hours. (e) Mitochondrial membrane potential in SH-SY5Y cells treated with Cpd A (10 or 25 μM) for 3 hours followed by 3 hours treatment with 50 μM 6-OHDA. Asterisks indicate levels significantly increased (t-test) as compared to control.
Mentions: Having established that both the competitive and the non-competitive HPH-inhibitor compounds are able to induce an HIF-1α response in the SH-SY5Y cells, we investigated the potential of these compounds to protect against oxidative stress. To model the cellular dysfunction of dopaminergic neurons in Parkinson's disease, where oxidative stress and mitochondrial dysfunction are cellular hallmarks [10], we treated SH-SY5Y cells with 6-OHDA. Treatment with 6-OHDA generates H2O2 and this oxidative stress leads to collapse of the mitochondrial membrane potential (ΔΨmit) and a reduction in cellular ATP production [11]. We found that 3 hours of 6-OHDA treatment was enough to induce a concentration-dependent collapse of both ΔΨmit and ATP-production, Figure 4, without reducing the cell numbers (not shown).

Bottom Line: In the present study, we compared competitive and noncompetitive HPH-inhibitor compounds in two different cell types (SH-SY5Y and PC12).Both competitive and non-competitive HPH inhibitors protected the cells against 6-OHDA induced oxidative stress.In addition, the protective effect of a specific HPH inhibitor was partially preserved when the cells were serum starved and exposed to 2-deoxyglucose, an inhibitor of glycolysis, indicating that other processes than restoring energy supply could be important for the HIF-mediated cytoprotection.

View Article: PubMed Central - PubMed

Affiliation: Department of Neurodegeneration, Lundbeck A/S, Ottiliavej 9, 2500 Valby, Denmark.

ABSTRACT
The hypoxia inducible factor 1 (HIF-1) is a central transcription factor involved in the cellular and molecular adaptation to hypoxia and low glucose supply. The level of HIF-1 is to a large degree regulated by the HIF prolyl hydroxylase enzymes (HPHs) belonging to the Fe(II) and 2-oxoglutarate-dependent dioxygenase superfamily. In the present study, we compared competitive and noncompetitive HPH-inhibitor compounds in two different cell types (SH-SY5Y and PC12). Although the competitive HPH-inhibitor compounds were found to be pharmacologically more potent than the non-competitive compounds at inhibiting HPH2 and HPH1, this was not translated into the cellular effects of the compounds, where the non-competitive inhibitors were actually more potent than the competitive in stabilizing and translocatingHIF1 α to the nucleus (quantified with Cellomics ArrayScan technology). This could be explained by the high cellular concentrations of the cofactor 2-oxoglutarate (2-OG) as the competitive inhibitors act by binding to the 2-OG site of the HPH enzymes. Both competitive and non-competitive HPH inhibitors protected the cells against 6-OHDA induced oxidative stress. In addition, the protective effect of a specific HPH inhibitor was partially preserved when the cells were serum starved and exposed to 2-deoxyglucose, an inhibitor of glycolysis, indicating that other processes than restoring energy supply could be important for the HIF-mediated cytoprotection.

No MeSH data available.


Related in: MedlinePlus