Consensus Recommendations for Studies of Outflow Facility and Intraocular Pressure Regulation Using Ex Vivo Perfusion Approaches.

Intraocular pressure (IOP) elevation is the primary risk factor and currently the main treatable factor for progression of glaucomatous optic neuropathy. In addition to direct clinical and living animal in vivo studies, ex vivo perfusion of anterior segments and whole eyes is a key technique for studying conventional outflow function as it is responsible for IOP regulation. We present well-tested experimental details, protocols, considerations, advantages, and limitations of several ex vivo model systems for studying IOP regulation.

Ischemic Optic Neuropathy: A Review of Current and Potential Future Pharmacotherapies.

The treatment of arteritic anterior ischemic optic neuropathy (AAION), non-arteritic ischemic optic neuropathy (NAAION), and posterior ischemic optic neuropathy (PION) is a topic of ongoing research with mixed evidence on some pharmacotherapies and a need for more consensus. This manuscript provides an overview of these conditions' current, potential future, and attempted pharmacotherapies. AAION's current treatment regimen consists of high-dose steroids, with methotrexate, tocilizumab, and abatacept, being the most viable steroid-sparing therapy candidates.

Quantifying ABCA1/apoA-1 Signaling Pathways with AFM Imaging and Lipidomic Analysis.

The role of lipid metabolic pathways in the pathophysiology of primary open-angle glaucoma (POAG) has been thoroughly elucidated, with pathways involved in lipid-related disorders such as hypercholesterolemia and hyperlipoprotein accumulation being of particular interest. The ABCA1/apoA-1 transduction pathway moderates reverse cholesterol transport (RCT), facilitating the transport of free cholesterol (FC) and phospholipids (PL) and preventing intracellular lipid aggregates in retinal ganglion cells (RGCs) due to excess FCs and PLs. A deficiency of ABCA1 transporters, and thus, dysregulation of the ABCA1/apoA-1 transduction pathway, may potentiate cellular lipid accumulation, which affects the structural and mechanical features of the cholesterol-rich RGC membranes.

Diacylglycerol Signaling in Retinal Ganglion Cells.

With impaired retinal ganglion cell (RGC) function and eventual RGC death, there is a heightened risk of experiencing glaucoma-induced blindness or other optic neuropathies. Poor RGC efficiency leads to limited transmission of visual signals between the retina and the brain by RGC axons. Increased focus on studying lipid messengers found in neurons such as endocannabinoids (eCBs) has importance due to their potential axonal pathway regenerative properties.

Downregulation of ATP8B2 to Assess Plasmalogen Distribution and Far1 Expression in Primary Trabecular Meshwork Cells.

The trabecular meshwork (TM) from primary open-angle glaucoma (POAG) cases has been found to contain decreased levels of intracellular plasmalogens. Plasmalogens are a subset of lipids involved in diverse cellular processes such as intracellular signaling, membrane asymmetry, and protein regulation. Proper plasmalogen biosynthesis is regulated by rate-limiting enzyme fatty acyl-CoA reductase (Far1).

Analysis of Sphingosine and Sphinganine from the Aqueous Humor for Signaling Studies Using Ultrahigh-Performance Liquid Chromatography-Mass Spectrometry.

Sphingolipids, including sphingosine and sphinganine, are one of the major classes of lipids. They serve as constituents of cell membranes and lipid rafts and aid in the performance of cell-cell communication and adhesion. Abnormal levels of sphingolipids in the aqueous humor can indicate impaired sphingolipid metabolism and associated ocular pathologies.

Optimized Lipidomics Extraction of Sphingosine and Sphinganine from Optic Nerve for Signaling Studies.

Interconvertible sphingolipid metabolites represent germane constituents of eukaryotic membranes and are vital in the regulation of cellular homeostasis, proliferation, survival, and induction of autophagy. This protocol describes a step-by-step method for extractions of sphingosine and sphinganine from mammalian tissue samples, particularly from the murine optic nerve. These lipids are partitioned into a binary mixture of chloroform and methanol in a modified Bligh and Dyer method.

Association of Primary Open-Angle Glaucoma with Diabetic Retinopathy among Patients with Type 1 and Type 2 Diabetes: A Large Global Database Study.

Purpose: To assess the correlation between primary open-angle glaucoma (POAG) and the risk of developing diabetic retinopathy (DR) in patients with type 1 diabetes mellitus (T1DM) and type 2 diabetes mellitus (T2DM).

Design: A retrospective cohort study leveraging the global patient database of TriNetX Research Network.

Participants: The study included 44 359 patients with diabetes mellitus (DM) with POAG and 4 393 300 patients with DM without any glaucoma ≥ 18 years of age.

Currently available prostanoids for the treatment of glaucoma and ocular hypertension: A review.

Recent advancements in prostaglandin analogs (PGAs) have reinforced their role in managing intraocular pressure (IOP). Latanoprost excels in 24-h IOP control, while various PGAs offer similar effectiveness and side effects, generic PGAs perform as well as branded ones, and a notable IOP rise observed upon PGA discontinuation. Formulations with or without preservatives show comparable IOP reduction and adherence, often surpassing benzalkonium chloride (BAK)-preserved options.

Appropriate patient population for future visual system axon regeneration therapies.

A number of blinding diseases caused by damage to the optic nerve result in progressive vision loss or loss of visual acuity. Secondary glaucoma results from traumatic injuries, pseudoexfoliation or pigmentary dispersion syndrome. Progressive peripheral vision loss is common to all secondary glaucoma irrespective of the initial event.

Mass spectrometry dataset of LC-MS lipidomics analysis of optic nerve.

CNS injuries of the anuran amphibian, , are uniquely suited for studying the molecular compositions of neuronal regeneration of retinal ganglion cells (RGC) due to a functional recovery of optic axons disparate to adult mammalian analogues. RGCs and their optic nerve axons undergo irreversible neurodegeneration in glaucoma and associated optic neuropathies, resulting in blindness in mammals. Conversely, demonstrates RGC lifetime-spanning regenerative capabilities after optic nerve crush [1], inciting opportunities to compare de novo regeneration and develop efficient pharmaceutical approaches for vision restoration.

Glaucomatous aqueous humor vesicles are smaller and differ in composition compared to controls.

Cells communicate with each other using vesicles of varying sizes, including a specific repertoire known as exosomes. We isolated aqueous humor (AH)-derived vesicles using two different methods: ultracentrifugation and an exosome isolation kit. We confirmed a unique vesicle size distribution in the AH derived from control and primary open-angle glaucoma (POAG) patients using various techniques, including Nanotracker, dynamic light scattering, atomic force imaging, and electron microscopy.

Metabolomics dataset of zebrafish optic nerve regeneration after injury.

Zebrafish () have the capacity for successful adult optic nerve regeneration. In contrast, mammals lack this intrinsic ability and undergo irreversible neurodegeneration seen in glaucoma and other optic neuropathies. Optic nerve regeneration is often studied using optic nerve crush, a mechanical neurodegenerative model.

Ocular Treatments Targeting Separate Prostaglandin Receptors in Mice Exhibit Alterations in Intraocular Pressure and Optic Nerve Lipidome.

Prostaglandin (PG) receptor agonists are the first-line eyedrop medication treatment for glaucoma. The pathophysiology of this disease is not completely known, and elevated intraocular pressure (IOP) is the key risk factor. The membranes of the axons (of the retinal ganglion cells) passing through the optic nerve (ON) head experience significant damage.

Glycerophospholipid Analysis of Optic Nerve Regeneration Models Indicate Potential Membrane Order Changes Associated with the Lipidomic Shifts.

Optic nerve (ON) injury causes irreversible degeneration, leading to vision loss that cannot be restored with available therapeutics. Current therapies slow further degeneration but do not promote regeneration. New regenerative factors have been discovered that are successful .

C-Laurdan: Membrane Order Visualization of HEK293t Cells by Confocal Microscopy.

Membrane order is a biophysical characteristic dependent on cellular lipid makeup. Cells regulate the membrane structure as it affects membrane-bound protein activity levels and membrane stability. Spatial organization of membrane lipids, such as lipid rafts, is a proposed theory that has been indirectly measured through polarity-sensitive fluorescent dyes.

Characterization of Trabecular Meshwork Mechanical Property Modulation After Application of Lipids.

Treatment of lipids endogenous to the aqueous humor of the eye could serve as a potential therapy to slow the progression of glaucoma. Herein, we describe the method to treat trabecular meshwork samples in vitro with lipids and characterize changes in the samples' stiffness.

Capillary Electrophoresis Plasma Fractionation for Lipids Analysis.

Extraction of lipids has mainly used density liquid phase separation techniques; however, these methods are limited by their broad extraction and lack specificity. Complex mixtures like blood plasma contain multiple lipid classes, whose distribution in the body are mediated by protein-lipid interactions and integration of lipids in larger lipoprotein complexes. The capillary electrophoresis system separates complex mixtures by electrokinetic forces that preserve protein-lipid complexation and allow for the fractionation of samples.

Analyses and Localization of Phosphatidylcholine and Phosphatidylserine in Murine Ocular Tissue Using Imaging Mass Spectrometry.

Imaging mass spectrometry (IMS) allows for spatial visualization of proteins, lipids, and metabolite distributions in a tissue. Identifying these compounds through mass spectrometry, combined with mapping the compound distribution in the sample in a targeted or untargeted approach, renders IMS a powerful tool for lipidomics. IMS analysis for lipid species such as phosphatidylcholine and phosphatidylserine allows researchers to pinpoint areas of lipid deficiencies or accumulations associated with ocular disorders such as age-related macular degeneration and diabetic retinopathy.

Analysis of Cholesterol Lipids Using Gas Chromatography Mass Spectrometry.

An optimized Bligh and Dyer protocol and subsequent derivatization is described in this chapter for the extraction of free cholesterol and cholesterol esters from tissue samples. Quantification analysis of lipid species is then described utilizing gas chromatography-mass spectrometry, the ideal method for analysis of volatile organic compounds and extraction of sterols.

Lipidomics Analysis with Triple Quadrupole Mass Spectrometry.

Lipids serve an essential role in multiple cellular functions including signaling, metabolism, energy storage, and membrane constitution. Lipidomics, the study of lipids using analytical chemistry, allows for the study of disease states and cellular metabolism. Shotgun lipidomics is a technique that involves direct-infusion electrospray ionization (ESI) and analysis with a triple quadrupole mass spectrometer.

Characterization of Gangliosides from Mouse Optic Nerve Samples Using Mass Spectrometry.

This chapter focuses on identifying gangliosides in the optic nerve of the mouse using mass spectrometry techniques. The described protocol will also permit the characterization of the sample's lipidome. Two deuterium-labeled ganglioside standards and a general lipid class standard will be utilized for extraction efficiency and quantification.