Time-of-flight secondary ion mass spectrometry to assess spatial distribution of A2E and its oxidized forms within lipofuscin granules isolated from human retinal pigment epithelium

Abstract

Lipofuscin granules accumulate in the cells of retinal pigment epithelium with age, particularly in patients with hereditary diseases. These granules are heterogeneous, being composed of mixtures of proteins and lipids, including more than 21 different fluorescent compounds. Bisretinoids and their photo-oxidation and photodegradation products represent the main source of lipofuscin fluorescence and exhibit phototoxic properties. This study used time-of-flight secondary ion mass spectrometry (ToF-SIMS) with in-depth probing to assess the depth distribution of N-retinylidene-N-retinylethanolamine (A2E) and its singly and doubly oxidized forms (A2E-ox and A2E-2ox, respectively) within lipofuscin granules and in their surface layer (lipid membrane). ToF-SIMS showed that A2E and its oxidized forms were uniformly distributed throughout lipofuscin granules but were not present at the membrane surface layer. This finding is important for understanding the process involved in the formation of lipofuscin granules and in their toxicity.

Mass spectrometry imaging for clinical research – latest developments, applications, and current limitations

“…MSI investigations in ophthalmology were recently summarized. 231 All of the studies show the importance of obtaining spatially defined molecular information from ocular tissues to better characterise several vision-threatening diseases, including age-related macular degeneration, 232,233 Stargardt's disease, 234 glaucoma, 235 and cataracts. 236 But MSI has also been used to study the biology of the healthy eye, such as the metabolic changes associated with light-and dark-treated retina.…”

Retinal Pigment Epithelium Pigment Granules: Norms, Age Relations and Pathology

“…LGs are heterogeneous and consist of a mixture of lipids, minor amounts of proteins modified and oxidized by end products of glycation and lipid peroxidation, and bisretinoid fluorophores that absorb light in the blue region of the spectrum and determine the autofluorescent properties of the granules [41,[47][48][49][50][51]. Structurally, LGs are yellow-orange granules surrounded by a lipid membrane and have an average diameter of about 1.0 micron [52][53][54][55].…”

Real-Time Molecular Patterns to support Intraoperative Decision-Making

“…MSI investigations in ophthalmology were recently summarized (264). All of the studies show the importance of obtaining spatially defined molecular information from ocular tissues to better characterise several vision-threatening diseases, including age-related macular degeneration (265,266), Stargardt disease (267), glaucoma (268), and cataracts (269). But MSI has also been used to study the biology of the healthy eye, such as the metabolic changes associated to light-and dark-treated retina (270).…”