Potential and Caveats of Lipidomics for Cardiovascular Disease
Authors: Raimund Pechlaner, Stefan Kiechl, Manuel Mayr
Journal: Circulation (2016)

ince the seminal publications from the Framingham study in the mid-60s, 1 the measurement of lipid levels, mainly of total cholesterol, total triglycerides, and low-density lipoprotein and high-density lipoprotein cholesterol, is routine clinical practice for cardiovascular disease (CVD) and lipid-lowering therapy.A more detailed assessment of the lipid composition, that is, the molecular species that constitute the lipid classes, is not widely used, mainly because of the caveats of assessing the lipidome.The human lipidome is estimated to include thousands of molecular lipid species with functional diversity.The molecular lipid species within a lipid class share a modular composition with fatty acids being attached to a common backbone.Although a characteristic head group within the backbone defines the lipid class, the diversity of molecular lipid species derives from the conjugated fatty acids. 2 The conjugated fatty acids can differ in their carbon chain length, the number, position, and configuration (cis or trans) of their double bonds, and the position and type (acyl, alkyl, or vinyl) of linkage to the backbone.Lipidomics refers to the comprehensive profiling of lipids, facilitated by recent advances in mass spectrometry (MS). 2 The identified molecular lipid species are designated by abbreviations that combine the description of the lipid class with the characteristics of the conjugated fatty acids. 3For example, a diglyceride (DG) with 2 fatty acids of 12 and 22 carbon atoms and 0 and 5 double bonds, respectively, both linked by an acyl linkage, can be described either as DG(12:0/22:5), or as DG(34:5).Most MS studies report the numbers of carbon atoms and double bonds either for the individual fatty acids or in total, but do not assign the precise positions of the fatty acids at the backbone or of the double bonds within the individual fatty acids.Sample extraction is key for lipidomic analysis.Traditionally, a 2-phase method has been used.The most popular one is the Folch extraction method with chloroform/methanol. 4 Methanol precipitates the proteins, whereas chloroform ensures effective extraction of a broad range of lipid classes from the precipitated lipoproteins.The 2-phase extraction, however, is laborious and not very amenable to automation.Single-phase methods are less time demanding, but concerns have been raised that the lipid extraction might be less uniform and less efficient. 5iquid chromatography is applicable to a broad range of lipid classes and, thus, the principal separation method used in lipidomics.Chromatographic separation of the lipid extract enhances sensitivity and specificity by reducing sample complexity, but can also add technical variability. 6Before MS analysis, the lipids have to be ionized by electrospray ionization.Then, the mass-to-charge ratio of the lipid species can be determined by MS.Next, fragmentation is commonly induced by tandem MS to enable the structural elucidation of the lipid species.For quantitation, stable isotope-labeled standards are spiked into the samples before lipid extraction to account for technical variation.The addition of authentic standards (targeted MS analysis) Potential and Caveats of lipidomics for Cardiovascular disease