A Comparison of Trabecular Meshwork Sphingolipids and Ceramides of Ocular Normotensive and Hypertensive States of DBA/2J Mice

Abstract

Purpose: To determine the differential profiles of sphingomyelin, sphingoid base, sphingoid base-1-phosphate, and ceramide and their quantitative differences between trabecular meshwork (TM) derived from normotensive and hypertensive intraocular pressure states of DBA/2J mice. Methods: Normotensive and hypertensive state TM were collected from mice and analyzed. Lipid extraction was performed using the Bligh and Dyer method, and the protein concentrations were determined using the Bradford method. The lipids were identified and quantified using appropriate standards with a TSQ Quantum Access Max triple quadrupole mass spectrometer applying class-specific lipid identification settings. Results: The comparative profiles of sphingomyelin, sphingoid base, sphingoid base-1-phosphate, and ceramide between normotensive and hypertensive TM showed several species unique to a phase and as well common between states. Conclusion: The presence or absence of several sphingolipids and ceramides in the normotensive or hypertensive states may contribute to better understanding of the glaucomas.

Endogenous ocular lipids as potential modulators of intraocular pressure

“…Samples were injected as direct infusion using a Triversa Nanomate driven by Chipsoft 8.3 (Advion Inc). The samples were run with and without addition of lipid standards for each class of lipids analysed as done in our previous studies . In addition, an unrelated lipid standard was used during extraction.…”

“…Normal AH has been shown to contain diverse phospholipids (PLs) . The TM and AH phospholipids and sphingolipids of human control and POAG as well as normotensive and ocular hypertensive DBA/2J mouse have been previously analysed using moderate‐resolution mass spectrometers …”

Sphingolipids and ceramides of mouse aqueous humor: Comparative profiles from normotensive and hypertensive DBA/2J mice

“…DBA/2J enables studying AH composition in normotensive and hypertensive states and thus helps capture lipid/metabolites associated with each of these states. In DBA/2J mice, the IOP elevation and development of glaucoma is natural, age-related, variable, progressive and asynchronous [24, 25]. The ocular hypertension in DBA/2J is generally thought to be due to pigmentary dispersion [53].…”

“…Further investigations will show the function of Cer(d18:0/18:1(9Z)) and its role in IOP elevation. Cer(d18:0/24:1(15Z)) was found to be unique in both female and male DBA/2J mice in the hypertensive phase when compared to the normotensive state (Tables 2-3) and was found to be unique in the TM of normotensive DBA/2J male mice [25]. The appearance of Cer(d18:0/24:1(15Z)) in normotensive TM and in hypertensive AH suggests that IOP elevation leads to the ceramide species being released from the TM to the AH or that it was sequestered in the AH.…”

“…For the normotensive group no single IOP measurement had been >17 mm of Hg and average IOP over two weeks of measurement had been ≤15 mm of Hg. At 8-9 months of age the IOP elevation in DBA/2J mice is asynchronous [24, 25] and thus any mice that showed an IOP <18 mm of Hg at any single point measurement was excluded from the study.…”