A lipidomic approach to the study of human CD4(+) T lymphocytes in multiple sclerosis

Abstract

Background: Lipids play different important roles in central nervous system so that dysregulation of lipid pathways has been implicated in a growing number of neurodegenerative disorders including multiple sclerosis (MS). MS is the most prevalent autoimmune disorder of the central nervous system, with neurological symptoms caused by inflammation and demyelination. In this study, a lipidomic analysis was performed for the rapid profile of CD4(+) T lymphocytes from MS patient and control samples in an untargeted approach.

Methods: A matrix-assisted laser desorption/ionization-time of flight (MALDI-TOF) mass spectrometry based approach was used for the analysis of lipid extracts using 9-aminoacridine as matrix. Lipids were analyzed in negative mode and selected species fragmented using MALDI tandem mass spectrometry for their structural assignments.

Results: The analysis reveals some modifications in the phospholipid pattern of MS CD4(+) T lymphocytes with respect to healthy controls with a significant increase of cardiolipin species in MS samples.

Conclusions: These results demonstrate the feasibility of a MALDI-TOF approach for the analysis of CD4(+) lipid extracts and suggest how alterations in the lipid metabolism characterized lymphocytes of MS patients.

Figure 1

MALDI-TOF mass spectra of lipids from CD4⁺ T lymphocytes obtained in positive and negative mode. Representative MALDI-TOF mass spectra of lipids extracted from CD4⁺ T lymphocytes (upper panels) prepared by the Bligh and Dyer procedure. Spectra were acquired in both negative (a) and positive (b) mode, using the 9-AA matrix by averaging 2.000 consecutive laser shots. MALDI-TOF spectra of 9-AA matrix detected in negative (c) and positive mode (d) is reported. Spectra were acquired using a Ultraflex Extreme MALDI-TOF/TOF (Bruker).

Figure 2

MALDI-TOF mass spectra of CD4⁺ lipids from MS and control subjects obtained in negative mode. Representative negative ion MALDI-TOF spectrum of total lipid extracts from healthy controls (a) and multiple sclerosis patients (b) CD4⁺ T lymphocytes, using the 9-AA matrix by averaging 2.000 consecutive laser shots.

Figure 3

Partial least squares discriminat analysis. Partial least squares discriminat analysis (PLS-DA) score plot from lipidomics data derived from control (black triangle) and MS (red circle) patients. Below, histograms show peaks that were significantly different (P < 0.05) among groups. Data reported as mean ± SD.

Figure 4

MS/MS based characterization of the cardiolipin m/z 1,448,14 ion. Representative MS spectra of cardiolipin species obtained from a MS sample (a), and from cardiolipin standard (b). Following MS/MS of the 1,448,14 cardiolipin generated fragment ions were illustrated (c); cardiolipin structure is reported in the insert.