Bioenergetic Failure in Rat Oligodendrocyte Progenitor Cells Treated with Cerebrospinal Fluid Derived from Multiple Sclerosis Patients

Abstract

In relapsing-remitting multiple sclerosis (RRMS) subtype, the patient's brain itself is capable of repairing the damage, remyelinating the axon and recovering the neurological function. Cerebrospinal fluid (CSF) is in close proximity with brain parenchyma and contains a host of proteins and other molecules, which influence the cellular physiology, that may balance damage and repair of neurons and glial cells. The purpose of this study was to determine the pathophysiological mechanisms underpinning myelin repair in distinct clinical forms of MS and neuromyelitis optica (NMO) patients by studying the effect of diseased CSF on glucose metabolism and ATP synthesis. A cellular model with primary cultures of oligodendrocyte progenitor cells (OPCs) from rat cerebrum was employed, and cells were treated with CSF from distinct clinical forms of MS, NMO patients and neurological controls. Prior to comprehending mechanisms underlying myelin repair, we determine the best stably expressed reference genes in our experimental condition to accurately normalize our target mRNA transcripts. The GeNorm and NormFinder algorithms showed that mitochondrial ribosomal protein (Mrpl19), hypoxanthine guanine phosphoribosyl transferase (Hprt), microglobulin β2 (B2m), and transferrin receptor (Tfrc) were identified as the best reference genes in OPCs treated with MS subjects and were used for normalizing gene transcripts. The main findings on microarray gene expression profiling analysis on CSF treated OPCs cells revealed a disturbed carbohydrate metabolism and ATP synthesis in MS and NMO derived CSF treated OPCs. In addition, using STRING program, we investigate whether gene-gene interaction affected the whole network in our experimental conditions. Our findings revealed downregulated expression of genes involved in carbohydrate metabolism, and that glucose metabolism impairment and reduced ATP availability for cellular damage repair clearly differentiate more benign forms from the most aggressive forms and worst prognosis in MS patients.

Keywords: cerebrospinal fluid; gene expression; glucose metabolism; multiple sclerosis; myelin repair; neuromyelitis optica; oligodendrocyte progenitor cells.

FIGURE 1

Normalized expression of (A) ActB; (B) B2m; (C) Hprt; (D) Mrpl19; (E) Tfrc; (F) Gapdh reference genes in OPCs tested in distinct disease courses of multiple sclerosis. G+/M- and G+/M+: Inflammatory forms of relapsing remitting multiple sclerosis; Med: Medullary form; PPMS: Primary progressive multiple sclerosis; NMO: Neuromyelitis Optica; Control: Other non-inflammatory neurological diseases (NIND).

FIGURE 2

Normalized expression of (A) Hk; (B) Gpi; (C) Aldoc; (D) Gapdh; (E) Tpi; (F) Pgk1; (G) Pgam1; (H) Eno1; (I) Eno2; (J) Pk; (K) Pkm2 genes involved in glycolytic pathway in OPCs treated with CSF of MS and NMO patients related to gene expression in OPCs treated with CSF of non-inflammatory neurological controls. OPCs: oligodendrocyte progenitor cells; Control: Neurons treated with CSF of non-inflammatory neurological controls; G+/M- and G+/M+: types of inflammatory MS; Med: Medullary MS; PP: Primary progressive multiple sclerosis; NMO: Neuromyelitis Optica. Four microarrays were used in each MS or NMO type of patients.

FIGURE 3

Normalized expression of (A) Pdha1; (B) Aco2; (C) Idh; (D) Ogdh; (E) Sdh; (F) Mdh2 genes involved in TCA cycle in OPCs treated with CSF of MS and NMO patients related to gene expression in OPCs treated with CSF of non-inflammatory neurological controls. OPCs: oligodendrocyte progenitor cells; Control: Neurons treated with CSF of non-inflammatory neurological controls; G+/M- and G+/M+: types of inflammatory MS; Med: Medullary MS; PP: Primary progressive multiple sclerosis; NMO: Neuromyelitis Optica. Four microarrays were used in each MS or NMO type of patients.

FIGURE 4

Normalized expression of (A) ATP5a1; (B) ATP5b; (C) Cox; (D) MT-ND2; (E) Cyc1 genes involved in oxidative phosphorylation in OPCs treated with CSF of MS and NMO patients related to gene expression in OPCs treated with CSF of non-inflammatory neurological controls. OPCs: oligodendrocyte progenitor cells; Control: Neurons treated with CSF of non-inflammatory neurological controls; G+/M- and G+/M+: types of inflammatory MS; Med: Medullary MS; PP: Primary progressive multiple sclerosis; NMO: Neuromyelitis Optica. Four microarrays were used in each MS or NMO type of patients.

FIGURE 5

Figure illustrating cumulative flux index in glycolysis, TCA cycle and electron transport chain in glucose metabolic pathway.

FIGURE 6

Visualization of gene interaction network generated by STRING v10 in OPCs exposed to CSF from (A–C) G+/M– RRMS; (D–F) G+/M+ RRMS; (G–I) Medullary MS; (J–L) PPMS; (M–O) NMO. Different line colors represent the types of evidence for the association between genes involved in (A,D,G,J,M) glycolysis; (B,E,H,K,N) TCA cycle and (C,F,I,L,O) oxidative phosphorylation in OPCs; Red color signifies upregulated expression, blue color signifies down regulated expression and gray color signifies genes that showed no variation in their expression in our experimental conditions. LOC689343 signifies Pk gene and ENSRNOG00000015290 signifies Tpi gene.