Research Article: CSF N-Glycan Profiles to Investigate Biomarkers in Brain Developmental Disorders: Application to Leukodystrophies Related to eIF2B Mutations

Date Published: August 29, 2012

Publisher: Public Library of Science

Author(s): Anne Fogli, Christine Merle, Véronique Roussel, Raphael Schiffmann, Sylvie Ughetto, Manfred Theisen, Odile Boespflug-Tanguy, Francisco José Esteban.


Primary or secondary abnormalities of glycosylation have been reported in various brain diseases. Decreased asialotransferrin to sialotransferrin ratio in cerebrospinal fluid (CSF) is a diagnostic marker of leukodystrophies related to mutations of genes encoding translation initiation factor, EIF2B. We investigated the CSF glycome of eIF2B-mutated patients and age-matched normal individuals in order to further characterize the glycosylation defect for possible use as a biomarker.

We conducted a differential N-glycan analysis using MALDI-TOF/MS of permethylated N-glycans in CSF and plasma of controls and eIF2B-mutated patients. We found in control CSF that tri-antennary/bisecting and high mannose structures were highly represented in samples obtained between 1 to 5 years of age, whereas fucosylated, sialylated structures were predominant at later age. In CSF, but not in plasma, of eIF2B-mutated patient samples, we found increased relative intensity of bi-antennary structures and decreased tri-antennary/bisecting structures in N-glycan profiles. Four of these structures appeared to be biomarker candidates of glycomic profiles of eIF2B-related disorders.

Our results suggest a dynamic development of normal CSF N-glycan profiles from high mannose type structures to complex sialylated structures that could be correlated with postnatal brain maturation. CSF N-glycome analysis shows relevant quantitative changes associated with eIF2B related disorders. This approach could be applied to other neurological disorders involving developmental gliogenesis/synaptogenesis abnormalities.

Partial Text

Genetic defects of N-glycan metabolisms due to abnormal hydrolysis (oligo saccharidosis), transport or storage (sialidosis and sialic acid storage disorders such as Salla disease and infantile sialic acid storage disease) and synthesis as in congenital disorders of glycosylation (CDG) are responsible for severe alterations of the CNS including myelination impairment [1]. Glycosylation is the most abundant post-translational event that yields functional active proteins. Glycan moieties play a major role for cell-cell and cell-matrix recognition during brain development and functions. Glycosylation maintains the axono-myelin-glial compartments in close contact with the blood and CSF compartments. Specific myelin glycoproteins such as MAG (myelin-associated glycoprotein), and MOG (myelin/oligodendrocyte glycoprotein) are respectively at the axonal and matrix interface [2]. Moreover, abnormal sugar chains of the cerebrospinal fluid (CSF) transferrin have been reported in various neurodegenerative disorders [3], [4]. Decreased CSF asialotransferrin to sialotransferrin ratio is considered as a biomarker of the CACH/VWM disorder (childhood ataxia with central nervous system hypomyelination/vanishing white matter) [5], [6] and can be measured using a HPLC method when 1.5 mL CSF is available [7]. This vacuolating form of leukodystrophy is related to mutations in the initiation factor, eIF2B, an ubiquitous factor involved in the global protein synthesis and its regulation under normal and stress conditions [8]–[12]. Its nucleotide guanine exchange activity (GEF) measured in patients transformed lymphocytes is decreased in eIF2B-mutated cells in comparison to controls [13], [14]. Little is known about how eIF2B mutations have an effect mainly on white matter (WM). Recent studies suggested a primitive abnormal maturation of glial cells during development in eIF2B-related disorders, leading to the alteration of the WM structure and to associated neurological dysfunctions [15]–[17]. In the present study, we investigated the CSF glycomic profile of control patients at different ages and compared them to those of eIF2B mutated patients using MALDI-TOF/MS (matrix-assisted laser desorption ionisation – time of flight – mass spectrometry) in order to test its usefulness as a biomarker identification in brain developmental disorders. We first described distinct CSF control N-glycan profiles in two groups of patients’ age at sampling (before and after 5 years of age). We then identified highly indicative changes in CSF N-glycan profiles of eIF2B-related disorders without changes in plasma. Four of these CSF N-glycan structures appeared as biomarker candidates that characterise the glycomic profiles of eIF2B-related disorders.

Using MALDI TOF MS-based analysis of permethylated glycans, we discovered an evolution of the control CSF glycome from N-glycan profiles rich in high mannose type structures to profiles with complex sialylated structures that could be correlated with postnatal brain maturation. We identified significant disease correlated differences in CSF but not in plasma N-glycan profiles of eIF2B-mutated patients suggesting for the first time a global change in N-glycosylation of CSF proteins in this pathology.