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Kelly K. Nichols, Jianzhong Chen, Mark Apsega, Jason J. Nichols, Kari B. Green; Identification of Lipid Biomarkers for Dry Eye Disease In Post-Menopausal Women Using Shotgun Electrospray Mass Spectrometry. Invest. Ophthalmol. Vis. Sci. 2012;53(14):619.
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To employ shotgun electrospray ionization mass spectrometry in the analysis of lipid collected from post-menopausal women with and without dry eye to determine quantitative differences in lipid species for the detection of lipid biomarkers.
Lipids were extracted from meibum using 2:1 chloroform:methanol and injected directly with sodium iodide additive as a shotgun approach to an electrospray time-of-flight mass spectrometer (ESI-MS) for analysis (Waters Q-TOF II and Bruker AmaZon ion trap). Meibum samples were collected from patients enrolled in a large study of postmenopausal women with and without dry eye (n = 440). Lipid data from the first 25 subjects from each group is presented here; analysis of the remaining samples is ongoing. Nonlinear Dynamics Progenesis MALDI software was used to quantify differences in lipid profiles to the level of individual lipid peaks in the spectra. Identity of the lipids showing statistical changes in abundance was confirmed by MS/MS.
Shotgun ESI-MS with sodium iodide additive of the samples provides hundreds of lipid peaks for comparison in both positive and negative ion mode, consistent with previous reports. Nonlinear Progenesis analysis of the data reveal over 200 lipids with a fold change greater than 1.25. Statistically significant differences between groups are found in the upper end of the spectra (around 1000 m/z), while observed yet non-significant differences are demonstrated across the spectra, particularly in the m/z range of 600-800 (wax esters); however, significant variability within the experimental groups is demonstrated, indicating larger sample sizes are needed for adequate comparison. In addition, trends showing a higher frequency of mono- and di-acylglycerols vs. tri-acylglycerols are appreciated in dry eye compared to non dry eye spectra. Trends among individual lipid profiles within the sub-groups provide the opportunity for clustering lipid profiles on the basis of clinical outcomes.
The major goal of dry eye disease biomarker discovery, and in this case lipid profiling, is to identify disease-specific lipids from human patients. Use of shot gun electrospray mass spectrometry coupled with Nonlinear Progenesis software analysis allows for comparison of lipid profiles between groups, demonstrating the potential for identifying lipid biomarkers based on up- or down-regulation of lipid sub-classes and/ or molecules.
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