Human lens lipids differ markedly from those of commonly used experimental animals

Abstract

Electrospray ionisation tandem mass spectrometry has allowed the unambiguous identification and quantification of individual lens phospholipids in human and six animal models. Using this approach ca. 100 unique phospholipids have been characterised. Parallel analysis of the same lens extracts by a novel direct-insertion electron-ionization technique found the cholesterol content of human lenses to be significantly higher (ca. 6 times) than lenses from the other animals. The most abundant phospholipids in all the lenses examined were choline-containing phospholipids. In rat, mouse, sheep, cow, pig and chicken, these were present largely as phosphatidylcholines, in contrast 66% of the total phospholipid in Homo sapiens was sphingomyelin, with the most abundant being dihydrosphingomyelins, in particular SM(d18:0/16:0) and SM(d18:0/24:1). The abundant glycerophospholipids within human lenses were found to be predominantly phosphatidylethanolamines and phosphatidylserines with surprisingly high concentrations of ether-linked alkyl chains identified in both classes. This study is the first to identify the phospholipid class (head-group) and assign the constituent fatty acid(s) for each lipid molecule and to quantify individual lens phospholipids using internal standards. These data clearly indicate marked differences in the membrane lipid composition of the human lens compared to commonly used animal models and thus predict a significant variation in the membrane properties of human lens fibre cells compared to those of other animals.

Identification of Abundant Alkyl Ether Glycerophospholipids in the Human Lens by Tandem Mass Spectrometry Techniques

“…The ion at m/z 764 is due to the protonated internal standard GPSer-(17:0/17:0), but the other three ions have previously been tentatively identified as GPSer ethers. 1 This assignment is supported by the negative ion m/z 153 precursor scan (dehydrated glycerophosphate) where the deprotonated ions of these GPSer lipids are not observed (Figure 1c). The m/z 153 ion is produced from diacyl-GPSer, glycerophosphatidic acid, glycerophosphoglycerol, and glycerophosphoinisitol.…”

“…Indeed, the 1-O-alkyl ethers identified in both classes account for 52% and 66%, respectively, of the total GPEtn and GPSer content of the human lens. 1 Evidence for lower levels of a range of other isomers including up to 9 isomers of GPSer(18:1e/18:1) differing only in the positions of double bonds on the two chains was also obtained. There is significant structural homology in the series of GPEtn and GPSerethers identified in this study that even extends to the relative distribution of double bond positional isomers.…”

“…We have previously assigned many of the lipids in this spectrum, including several with masses consistent with GPEtn-ethers. 1 The identity of some of these ions as 1-Oalk-1′-enyl-GPEtn (# in Figure 1a 1a), produced no such characteristic ions upon CID. 1 The structural elucidation of these lipids and a series of related GPSerethers is outlined below.…”

“…Phospholipids were extracted from human lenses as previously described. 1 In brief, lenses were frozen under liquid nitrogen, ground using a mortar and pestle and chloroform/ methanol (2:1 v/v) containing 0.01% BHT added at a ratio of 20:1 solvent to tissue (v/w). A methanolic internal standard solution was prepared comprising dinonadecanoyl phosphatidylcholine [GPCho(19: 15 The internal standard mixture was added to the homogenates at 1.4 mL g(tissue) -1 for later quantification of phospholipids.…”

“…A small fraction of this ion population was therefore tentatively assigned to a third ether-bearing GPEtn. 1 The CID and Figure 4c. For the m/z 730 molecular anion, the n-7 ions appear at about 20% of the abundance of the n-9 ions; this is significantly less than the 50% observed for m/z 728 and is closer to the 15% observed for the m/z 702.…”