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Disruption of TCA Cycle and Glutamate Metabolism Identified by Metabolomics in an In Vitro Model of Amyotrophic Lateral Sclerosis

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Abstract

This study aims to develop a cellular metabolomics model that reproduces the pathophysiological conditions found in amyotrophic lateral sclerosis in order to improve knowledge of disease physiology. We used a co-culture model combining the motor neuron-like cell line NSC-34 and the astrocyte clone C8-D1A, with each over-expressing wild-type or G93C mutant human SOD1, to examine amyotrophic lateral sclerosis (ALS) physiology. We focused on the effects of mutant human SOD1 as well as oxidative stress induced by menadione on intracellular metabolism using a metabolomics approach through gas chromatography coupled with mass spectrometry (GC-MS) analysis. Preliminary non-supervised analysis by Principal Component Analysis (PCA) revealed that cell type, genetic environment, and time of culture influenced the metabolomics profiles. Supervised analysis using orthogonal partial least squares discriminant analysis (OPLS-DA) on data from intracellular metabolomics profiles of SOD1G93C co-cultures produced metabolites involved in glutamate metabolism and the tricarboxylic acid cycle (TCA) cycle. This study revealed the feasibility of using a metabolomics approach in a cellular model of ALS. We identified potential disruption of the TCA cycle and glutamate metabolism under oxidative stress, which is consistent with prior research in the disease. Analysis of metabolic alterations in an in vitro model is a novel approach to investigation of disease physiology.

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Abbreviations

ALS:

Amyotrophic lateral sclerosis

F or SALS:

Familial or sporadic amyotrophic lateral sclerosis

SOD1:

Superoxide dismutase 1 (m, mutant; WT, wild-type)

CSF:

Cerebrospinal fluid

DNA:

Deoxyribonucleic acid

cDNA:

Complementary DNA

EAAT2:

Excitatory amino acid transporter 2

GAPDH:

Glyceraldehyde 3-phosphate dehydrogenase

GC-MS:

Gas chromatography coupled to mass spectrometry

GFP:

Green fluorescent protein

GFAP:

Glial fibrillary acidic protein

GLT1:

Glial glutamate transporter 1

QC:

Quality control

PCA:

Principal Component Analysis

OPLS-DA:

Orthogonal partial least-squares discriminant analysis

RNA:

Ribonucleic acid

ROS:

Reactive oxygen species

TCA cycle:

Tricarboxylic acid cycle

GSH:

Glutathione

NMDA:

N-methyl-D-aspartic acid

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Acknowledgments

We thank the Association ARSLA (Association pour la Recherche sur la Sclérose Latérale Amyotrophique) for its financial support. We thank Christophe Arnoult from GICC Unity Research (Génétique, Immunothérapie, Chimie et Cancer) for its technical support.

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Authors and Affiliations

  1. UMR INSERM U930, Université François-Rabelais de Tours, Equipe « Neurogénétique et neurométabolomique », 37032, Tours, France

    Charlotte Veyrat-Durebex, Philippe Corcia, Audrey Dangoumau, Patrick Vourc’h, Patrick Emond, Frédéric Laumonnier, Lydie Nadal-Desbarats, Christian R Andres & Hélène Blasco

  2. CHRU de Tours, Laboratoire de Biochimie et de biologie moléculaire, 37044, Tours, France

    Charlotte Veyrat-Durebex, Patrick Vourc’h, Christian R Andres & Hélène Blasco

  3. CHRU de Tours, Service de Neurologie, 37044, Tours, France

    Philippe Corcia

  4. INSERM U966, 37032, Tours, France

    Eric Piver

  5. Département de Pharmacologie médicale, INSERM U1171, Université Lille Nord de France, CHRU de Lille, Lille, France

    David Devos & Flore Gouel

  6. PPF-ASB, Université François Rabelais de Tours, Tours, France

    Patrick Emond & Lydie Nadal-Desbarats

  7. Northern Navajo Medical Center, Shiprock, NM, USA

    Paul H Gordon

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  1. Charlotte Veyrat-Durebex
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  2. Philippe Corcia
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Correspondence to Charlotte Veyrat-Durebex.

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Veyrat-Durebex, C., Corcia, P., Piver, E. et al. Disruption of TCA Cycle and Glutamate Metabolism Identified by Metabolomics in an In Vitro Model of Amyotrophic Lateral Sclerosis. Mol Neurobiol 53, 6910–6924 (2016). https://doi.org/10.1007/s12035-015-9567-6

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