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Nicolas G. Bazan, Karin Z. Berry, William C. Gordon, Eric J. Knott, Cornelius E. Regan, Jr, Robert C. Murphy, Bok Kyoo Jun; Spatial Distribution Of Lipid Molecular Species In Photoreceptors And Other Cells By Maldi Imaging Mass Spectrometry Of The Human Retina. Invest. Ophthalmol. Vis. Sci. 2012;53(14):2216.
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© ARVO (1962-2015); The Authors (2016-present)
To identify lipid species and their distribution within the human macula by direct raster-like scans of histological sections while collecting data at each point by imaging matrix-assisted laser desorption/ionization mass spectrometry (MALDI IMS) to generate images of the location of specific lipids.
Sections of horizontal strips containing optic nerve and macular region from human eyes were obtained. Some were H&E stained for orientation. After matrix deposition, a quadrupole-TOF tandem mass spectrometer with an orthogonal MALDI source (QSTAR XL, Applied Biosystems/MDS Sciex) acquired images. MALDI mass spectra were obtained. Plates were scanned at 12.75 mm/min (vertical 50 μm shift, lateral resolution 50μm). Data were processed by Analyst software and images visualized with TissueView. Collisional ion activation occurred after MALDI imaging on the same section. LC-MS/MS analysis was performed on central and peripheral retina to identify very long chain polyunsaturated fatty acyl groups (VLC-PUFAs).
MALDI IMS analysis of histological sections revealed differential distribution of lipid species among eye tissues (e.g., fatty support tissues, TAG(52:2)+Na; optic nerve, SM(18:1/18:0)+Na, PS(18:0/18:1), PE(18:0p/22:6); inner retina, PC(34:1)+Na, PE(18:0p/22:6); photoreceptors, PC(40:6)+Na, PE (18:0/22:6); RPE, Cer(18:1/18:0)-H2O)) and stratification (e.g., inner retina, PC(48:11)+Na; photoreceptors, PC(40:6 & 56:11)+Na). MS analysis revealed more PC-VLC-PUFAs within the central region, with 2-3 times as much 56 & 58 acyl carbons (10-12 double bonds) in phospholipid species.
MALDI IMS revealed lipid stratification among photoreceptor and inner retina layers, and in optic nerve and fatty support tissues. Phosphatidyl choline species, especially those enriched in LC- and VLC-PUFAs, localize to the photoreceptor layer; were more abundant in central retina. We have further used conventional LC-MS/MS- based lipidomic analysis to characterize lipid molecular species in macula as a function of aging and in degenerative diseases. The ability to localize specific lipids to the micrometer level in a retinal section will lead to identification of regional differences in lipid populations in both healthy and diseased retinas.
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