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Lactate-modulated induction of THBS-1 activates transforming growth factor (TGF)-beta2 and migration of glioma cells in vitro.

Seliger C, Leukel P, Moeckel S, Jachnik B, Lottaz C, Kreutz M, Brawanski A, Proescholdt M, Bogdahn U, Bosserhoff AK, Vollmann-Zwerenz A, Hau P - PLoS ONE (2013)

Bottom Line: Transforming growth factor (TGF)-beta2, which we previously showed to be induced by lactic acid, is a key pathophysiological factor in glioblastoma, leading to increased invasion and severe local immunosuppression after proteolytic cleavage from its latency associated peptide.Lactate levels were reduced by knockdown of LDH-A using specific small interfering RNA (siRNA) and competitive inhibition of LDH-A by sodium oxamate.Knockdown of LDH-A with subsequent decrease of lactate concentration leads to reduced levels of THBS-1 and TGF-beta2 in glioma cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Neurology and Wilhelm Sander-NeuroOncology Unit, University Hospital Regensburg, Regensburg, Germany.

ABSTRACT

Background: An important phenomenon observed in glioma metabolism is increased aerobic glycolysis in tumor cells, which is generally referred to as the Warburg effect. Transforming growth factor (TGF)-beta2, which we previously showed to be induced by lactic acid, is a key pathophysiological factor in glioblastoma, leading to increased invasion and severe local immunosuppression after proteolytic cleavage from its latency associated peptide. In this study we tested the hypothesis, that lactate regulates TGF-beta2 expression and glioma cell migration via induction of Thrombospondin-1 (THBS-1), a TGF-beta activating protein.

Methods: Lactate levels were reduced by knockdown of LDH-A using specific small interfering RNA (siRNA) and competitive inhibition of LDH-A by sodium oxamate. Knockdown of THBS-1 was performed using specific siRNA. Western Blot, qRT-PCR, and ELISA were used to investigate expression levels of LDH-A, LDH-B, TGF-beta2 and THBS-1. Migration of cells was examined by Spheroid, Scratch and Boyden Chamber assays.

Results: Knockdown of LDH-A with subsequent decrease of lactate concentration leads to reduced levels of THBS-1 and TGF-beta2 in glioma cells. Lactate addition increases THBS-1 protein, leading to increased activation of TGF-beta2. Inhibition of THBS-1 reduces TGF-beta2 protein and migration of glioma cells. Addition of synthetic THBS-1 can rescue reduced TGF-beta2 protein levels and glioma cell migration in siLDH-A treated cells.

Conclusion: We define a regulatory cascade between lactate, THBS-1 and TGF-beta2, leading to enhanced migration of glioma cells. Our results demonstrate a specific interaction between tumor metabolism and migration and provide a better understanding of the mechanisms underlying glioma cell invasion.

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Related in: MedlinePlus

Glioma cell migration is enhanced after treatment with sodium lactate.HTZ-349 (A) and U87 (B) glioma cells were treated with 20 mM sodium lactate (pH 7.4) and HCl (pH 7.1) was used as control. 24 hours after treatment cell migration was analyzed using Boyden Chamber assays. The Y-axis indicates number of migrated cells. Treatment with lactate leads to a highly significant increase in glioma cell migration (U87 and HTZ-349, p < 0.001***).
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pone-0078935-g002: Glioma cell migration is enhanced after treatment with sodium lactate.HTZ-349 (A) and U87 (B) glioma cells were treated with 20 mM sodium lactate (pH 7.4) and HCl (pH 7.1) was used as control. 24 hours after treatment cell migration was analyzed using Boyden Chamber assays. The Y-axis indicates number of migrated cells. Treatment with lactate leads to a highly significant increase in glioma cell migration (U87 and HTZ-349, p < 0.001***).

Mentions: Migration is a key feature of glioma malignancy. To evaluate if lactate is able to modulate glioma cell migration, we performed Boyden Chamber assays with U87 and HTZ-349 glioma cells. Treatment of U87 or HTZ-349 glioma cells with 20 mM sodium lactate corresponds to levels measured in brain tumor specimen9 and led to a highly significant increase in glioma cell migration 24 hours after treatment (Figure 2A, B). Hydrochloric acid alone failed to significantly enhance HTZ-349 glioma cell migration (Figure 2A), whereas U87 glioma cells also showed moderately increased migratory capacity after acidification by hydrochloric acid (Figure 2B).


Lactate-modulated induction of THBS-1 activates transforming growth factor (TGF)-beta2 and migration of glioma cells in vitro.

Seliger C, Leukel P, Moeckel S, Jachnik B, Lottaz C, Kreutz M, Brawanski A, Proescholdt M, Bogdahn U, Bosserhoff AK, Vollmann-Zwerenz A, Hau P - PLoS ONE (2013)

Glioma cell migration is enhanced after treatment with sodium lactate.HTZ-349 (A) and U87 (B) glioma cells were treated with 20 mM sodium lactate (pH 7.4) and HCl (pH 7.1) was used as control. 24 hours after treatment cell migration was analyzed using Boyden Chamber assays. The Y-axis indicates number of migrated cells. Treatment with lactate leads to a highly significant increase in glioma cell migration (U87 and HTZ-349, p < 0.001***).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3815307&req=5

pone-0078935-g002: Glioma cell migration is enhanced after treatment with sodium lactate.HTZ-349 (A) and U87 (B) glioma cells were treated with 20 mM sodium lactate (pH 7.4) and HCl (pH 7.1) was used as control. 24 hours after treatment cell migration was analyzed using Boyden Chamber assays. The Y-axis indicates number of migrated cells. Treatment with lactate leads to a highly significant increase in glioma cell migration (U87 and HTZ-349, p < 0.001***).
Mentions: Migration is a key feature of glioma malignancy. To evaluate if lactate is able to modulate glioma cell migration, we performed Boyden Chamber assays with U87 and HTZ-349 glioma cells. Treatment of U87 or HTZ-349 glioma cells with 20 mM sodium lactate corresponds to levels measured in brain tumor specimen9 and led to a highly significant increase in glioma cell migration 24 hours after treatment (Figure 2A, B). Hydrochloric acid alone failed to significantly enhance HTZ-349 glioma cell migration (Figure 2A), whereas U87 glioma cells also showed moderately increased migratory capacity after acidification by hydrochloric acid (Figure 2B).

Bottom Line: Transforming growth factor (TGF)-beta2, which we previously showed to be induced by lactic acid, is a key pathophysiological factor in glioblastoma, leading to increased invasion and severe local immunosuppression after proteolytic cleavage from its latency associated peptide.Lactate levels were reduced by knockdown of LDH-A using specific small interfering RNA (siRNA) and competitive inhibition of LDH-A by sodium oxamate.Knockdown of LDH-A with subsequent decrease of lactate concentration leads to reduced levels of THBS-1 and TGF-beta2 in glioma cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Neurology and Wilhelm Sander-NeuroOncology Unit, University Hospital Regensburg, Regensburg, Germany.

ABSTRACT

Background: An important phenomenon observed in glioma metabolism is increased aerobic glycolysis in tumor cells, which is generally referred to as the Warburg effect. Transforming growth factor (TGF)-beta2, which we previously showed to be induced by lactic acid, is a key pathophysiological factor in glioblastoma, leading to increased invasion and severe local immunosuppression after proteolytic cleavage from its latency associated peptide. In this study we tested the hypothesis, that lactate regulates TGF-beta2 expression and glioma cell migration via induction of Thrombospondin-1 (THBS-1), a TGF-beta activating protein.

Methods: Lactate levels were reduced by knockdown of LDH-A using specific small interfering RNA (siRNA) and competitive inhibition of LDH-A by sodium oxamate. Knockdown of THBS-1 was performed using specific siRNA. Western Blot, qRT-PCR, and ELISA were used to investigate expression levels of LDH-A, LDH-B, TGF-beta2 and THBS-1. Migration of cells was examined by Spheroid, Scratch and Boyden Chamber assays.

Results: Knockdown of LDH-A with subsequent decrease of lactate concentration leads to reduced levels of THBS-1 and TGF-beta2 in glioma cells. Lactate addition increases THBS-1 protein, leading to increased activation of TGF-beta2. Inhibition of THBS-1 reduces TGF-beta2 protein and migration of glioma cells. Addition of synthetic THBS-1 can rescue reduced TGF-beta2 protein levels and glioma cell migration in siLDH-A treated cells.

Conclusion: We define a regulatory cascade between lactate, THBS-1 and TGF-beta2, leading to enhanced migration of glioma cells. Our results demonstrate a specific interaction between tumor metabolism and migration and provide a better understanding of the mechanisms underlying glioma cell invasion.

Show MeSH
Related in: MedlinePlus