Cholesterol and Glycosphingolipids of Human Trabecular Meshwork and Aqueous Humor: Comparative Profiles from Control and Glaucomatous Donors

Purpose: To determine the differential profiles of cholesterol and glycosphingolipid species and their quantitative differences between control and glaucomatous aqueous humor (AQH) and the trabecular meshwork (TM) derived from human donors. Methods: Control TM and selected primary open angle glaucoma (POAG) TM samples were collected from cadaveric donors. Other TM samples, glaucomatous AQH and control AQH were procured during intraocular surgery. Lipid extraction was performed using modifications of the Bligh and Dyer method. Protein concentration was estimated using the Bradford colorimetric assay. Cholesterol and glycosphingolipids were identified and subjected to ratiometric quantification utilizing precursor ion scan and neutral ion loss scan in positive ion mode using appropriate class specific lipid standards (Cholesterol and Psychosine) on a TSQ Quantum Access Max mass spectrometer. Results: Control and glaucomatous AQH demonstrated 7 and 4 unique cholesterol species, whereas the TM demonstrated 7 and 12 unique species, respectively. The control and POAG AQH showed 6 and 0 whereas TM samples showed 5 and 1 unique glycosphingolipids, respectively. A total of 65 and 62 common cholesterol species and 59 and 58 common glycosphingolipids were found in AQH and TM, respectively. Increased zymosterol and glucopyranosyl cholesterol levels were found in glaucomatous AQH. Significantly decreased levels of galactosylceramide, glucosylceramide in glaucomatous TM were found compared to control TM. Conclusion: A high percentage of cholesterol and glycosphingolipid species was found to be common between control and POAG AQH and TM. Several cholesterol and glycosphingolipid species was found to be unique in a subset of POAG or controls. Glaucomatous aqueous humor and TM showed relatively higher levels of zymosterol (an intermediate precursor of cholesterol) and decreased glycoceramide levels, respectively.