Identification and Quantitation of Changes in the Platelet Activating Factor Family of Glycerophospholipids over the Course of Neuronal Differentiation by High-Performance Liquid Chromatography Electrospray Ionization Tandem Mass Spectrometry

Abstract

Glycerophospholipids are important structural lipids in membranes with changes associated with progressive neurodegenerative disorders such as Alzheimer disease. Synthesis of the platelet activating factor (PAF) glycerophospholipid subclass is implicated in the control of neuronal differentiation and death. In this article, we combine nanoflow HPLC and mass spectrometry to screen, identify, and quantitate changes in glycerophospholipid subspecies, specifically PAF family members, over the course of neuronal differentiation. Furthermore, precursor ion scans for fragments characteristic of PAF phosphocholine family members and the standard additions of PAF subspecies were combined to perform absolute quantitation of PAF lipids in undifferentiated and differentiated PC12 cells. Surprisingly, a marked asymmetry was detected in the two predominant PAF species (C16:0, C18:0) over the course of differentiation. These results describe a new technique for the sensitive analysis of lipids combining nanoflow HPLC, ESI-MS, and precursor ion scan. Limits of detection of as little as 2 pg of PAF and LPC were obtained, and analysis of the lipidome of as little as 70,000 cells was performed on this system. Furthermore, application to the PC12 model identified a quantifiable difference between PAF molecular species produced over the course of neuronal differentiation.

Heterogeneity in the sn-1 carbon chain of platelet-activating factor glycerophospholipids determines pro- or anti-apoptotic signaling in primary neurons

“…C 16 -PAF and C 18 -PAF are the predominant PAF species present in uninjured tissue (39,40) but differentially accumulate over development of or following injury. Recent studies from our laboratory have shown that C 16 -PAF concentrations increase during neuronal differentiation (41), whereas local C 18 -PAF concentrations have only been shown to increase in the injured central nervous system (32). Here, we show that both C 16 -PAF and C 18 -PAF trigger neuronal death in PAFR-negative cells, but through isoform-specific apoptotic or neurodegenerative signaling pathways, respectively.…”

The Human Serum Metabolome

“…However, the determination of the exact position of the double bond and the relative number of the positional isomers poses a major challenge in lipidomics [95], [98]. Other challenges also exist for this field including: 1) the observation that different classes of lipids may significantly affect instrument response of [99]; 2) the different patterns that identical lipid species may show when analyzed by various types of mass spectrometers or by the same mass spectrometer in different experimental modes [100] and 3) the limited availability of internal standards with the exact structure of the lipids of interest [101]. Even with these limitations, it is still clear that targeted lipid analysis of serum will always yield an enormous abundance or metabolites.…”

Amyloid-β42signals tau hyperphosphorylation and compromises neuronal viability by disrupting alkylacylglycerophosphocholine metabolism

“…1A). PAF isoforms were profiled by high-performance liquid chromatography electrospray ionization mass spectrometry (LC-ESI-MS) (12). Lipids with mass-to-charge ratios (m/z) of 450-600 were analyzed in positive ion mode by MS scan for a protonated molecule at expected m/z followed by precursor ion scan for a diagnostic phosphocholine product ion at m/z 184.…”