Abstract
Purpose :
To determine major differences in lipid profile between human control and glaucomatous optic nerve. To assess major enzymes in lipid pathway if aberration is revealed for a lipid class by profiling.
Methods :
Optic nerve (ON) samples were obtained from human cadaveric donors [control (n=11) and primary open angle glaucoma (POAG; n=12)], the lipids were extracted using Bligh and Dyer methods. The cadaveric tissue was collected and research was coducted following the tenets of the Declaration of Helsinki. Control and glaucoma donors were all Caucasian age 72.3±5.9 and 70.3± 10.5 (inclusive of both genders) respectively. High resolution mass spectrometry was performed using a Q-exactive mass spectrometer coupled with an EASY-nLC 1000 liquid chromatograph instrument. Bioinformatics and statistical analysis were performed using LipidSearch v.4.1 and MetaboAnalyst 4.0/STATA 14.2. Protein amounts were determined using the Bradford’s method. Western blot, ELISA and immunohistochemistry utilized established protocols for protein quantification and localization, respectively.
Results :
Principal component analysis (PCA) placed control and glaucomatous optic nerves in two distinct groups based on analysis of lipid profiles. Total lipid, total phospholipids, total ceramide and total sphingolipids were similar (without significant difference) between control and glaucoma. However, we found a significant increase in glucosylsphingosine in glaucoma compared to control samples. We found similar levels of glucocerebrosidase (GBA), ceramide glucosyltransferase (UGCG), decreased non-lysosomal glucocerebrosidase (GBA2) and increased lysosomal and non-lysosomal acylsphingosine amidohydrolase (ASAH1& ASAH2) levels in glaucomatous ON compared to control.
Conclusions :
We found significant differences in glucosylsphingosine lipids, consistent with decreased GBA and GBA2 and increased ASAH1 and ASAH2 immunoreactivity in glaucoma, suggesting the potential impairment of sphingolipid enzymatic pathways in lysosomal and non-lysosomal cellular compartments.
This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.