Dataset Information

Morphometric Analysis of Retinal Ganglion Cell Axons in Normal and Glaucomatous Brown Norway Rats Optic Nerves.

ABSTRACT:

Purpose

A reference atlas of optic nerve (ON) retinal ganglion cell (RGC) axons could facilitate studies of neuro-ophthalmic diseases by detecting subtle RGC axonal changes. Here we construct an RGC axonal atlas for normotensive eyes in Brown Norway rats, widely used in glaucoma research, and also develop/evaluate several novel metrics of axonal damage in hypertensive eyes.

Methods

Light micrographs of entire ON cross-sections from hypertensive and normotensive eyes were processed through a deep learning-based algorithm, AxoNet2.0, to determine axonal morphological properties and were semiquantitatively scored using the Morrison grading scale (MGS) to provide a damage score independent of AxoNet2.0 outcomes. To construct atlases, ONs were conformally mapped onto an ON "template," and axonal morphometric data was computed for each region. We also developed damage metrics based on myelin morphometry.

Results

In normotensive eyes, average axon density was ∼0.3 axons/µm2 (i.e., ∼80,000 axons in an ON). We measured axoplasm diameter, eccentricity, cross-sectional area, and myelin g-ratio and thickness. Most morphological parameters exhibited a wide range of coefficients of variation (CoV); however, myelin thickness CoV was only ∼2% in normotensive eyes. In hypertensive eyes, increased myelin thickness correlated strongly with MGS (P < 0.0001).

Conclusions

We present the first comprehensive normative RGC axon morphometric atlas for Brown Norway rat eyes. We suggest objective, repeatable damage metrics based on RGC axon myelin thickness for hypertensive eyes.

Translational relevance

These tools can evaluate regional changes in RGCs and overall levels of damage in glaucoma studies using Brown Norway rats.

SUBMITTER: Goyal V

PROVIDER: S-EPMC10020949 | biostudies-literature | 2023 Mar

REPOSITORIES: biostudies-literature

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Publications

Morphometric Analysis of Retinal Ganglion Cell Axons in Normal and Glaucomatous Brown Norway Rats Optic Nerves.

Goyal Vidisha V Read A Thomas AT Brown Dillon M DM Brawer Luke L Bateh Kaitlyn K Hannon Bailey G BG Feola Andrew J AJ Ethier C Ross CR

Translational vision science & technology 20230301 3

<h4>Purpose</h4>A reference atlas of optic nerve (ON) retinal ganglion cell (RGC) axons could facilitate studies of neuro-ophthalmic diseases by detecting subtle RGC axonal changes. Here we construct an RGC axonal atlas for normotensive eyes in Brown Norway rats, widely used in glaucoma research, and also develop/evaluate several novel metrics of axonal damage in hypertensive eyes.<h4>Methods</h4>Light micrographs of entire ON cross-sections from hypertensive and normotensive eyes were processed ...[more]

PMID: 36917118

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Project description:PurposeTo determine whether lysophospholipid (LPL) profiles and corresponding conversion enzymes in the LPL pathways are altered in the optic nerve (ON) between human control and glaucoma samples.MethodsLipids extracted from control (n = 11) and glaucomatous (n = 12) ON samples using the Bligh and Dyer method were subjected to high-resolution mass spectrometry on a Q-exactive mass spectrometer coupled with a high-performance liquid chromatography (Accela 600) system. Analysis was performed for LPLs (lysophosphatidylcholines, lysophosphatidylserines, lysophosphatidylethanolamines, lysophosphatidylinositols, and lysosphingomyelines) using LipidSearch v.4.1, MZmine v.2.0, and MetaboAnalyst v.4.0. LPL synthesis and degradation pathway maps, utilizing UniProt and BRENDA database entries as needed, were created using Kyoto Encyclopedia of Genes and Genomes (KEGG)-based tools. The mRNA expression level in normal and glaucomatous human ON were analyzed using Gene Expression Omnibus (GEO) entry GSE45570. Protein amounts were determined using PHAST gel and dot blot and were used for normalization of protein amounts across samples. Western blot, ELISA, and protein quantification were performed using established protocols.ResultsPrincipal component analysis of ON LPL profile placed control and glaucomatous ONs in two distinct separate groups. Mass spectrometric analysis of ON revealed decrease in lysophosphatidic acid, lysophosphatidylethanolamine, lysophosphatidylcholine, and significant increase in diacylglycerol in glaucomatous ON. Statistical analysis of LPL conversion enzymes revealed significant overexpression of phosphatidate phosphatase LPIN2, phospholipid phosphatase 3, phosphatidylcholine-sterol acyltransferase, and calcium-dependent phospholipase 2, and significant downregulation of glycerol-3-phosphate acyltransferase 4 at mRNA level in glaucomatous ON. Western blot and ELISA confirmed proteomic differences between normal and diseased ON.ConclusionsOur analysis revealed alterations in specific LPL levels and corresponding select enzyme-level changes in glaucomatous ON.

Project description:PurposeTo quantify the morphological features of the optic nerve head using radial polynomials, to use these morphometric models as the basis for classification of glaucomatous optic neuropathy via an automated decision tree induction algorithm, and to compare these classification results with established procedures.MethodsA cohort of patients with high-risk ocular hypertension or early glaucoma (n=179) and a second cohort of normal subjects (n=96) were evaluated for glaucomatous optic neuropathy using stereographic disc photography and confocal scanning laser tomography. Morphological features of the optic nerve head region were modeled from the tomography data using pseudo-Zernike radial polynomials and features derived from these models were used as the basis for classification by a decision tree induction algorithm. Decision tree classification performance was compared with expert classification of stereographic disc photographs and analysis of neural retinal rim thickness by Moorfields Regression Analysis (MRA).ResultsRoot mean squared error of the morphometric models decreased asymptotically with additional polynomial coefficients, from 62±0.5 (32 coefficients) to 32±5.7 μm (256 coefficients). Optimal morphometric classification was derived from a subset of 64 total features and had low sensitivity (69%), high specificity (88%), very good accuracy (80%), and area under the receiver operating characteristic curve (AUROC) was 88% (95% confidence interval, 78%-98%). In comparison, MRA classification of the same records had a comparatively poorer sensitivity (55%), but had higher specificity (95%), with similar overall accuracy (78%) and AUROC curve, 83% (95% CI, 70%-96%).ConclusionsPseudo-Zernike radial polynomials provide a mathematically compact and faithful morphological representation of the structural features of the optic nerve head. This morphometric method of glaucomatous optic neuropathy classification has greater sensitivity, and similar overall classification performance (AUROC) when compared with classification by neural retinal rim thickness by MRA in patients with high-risk ocular hypertension and early glaucoma.

Dataset Information

Morphometric Analysis of Retinal Ganglion Cell Axons in Normal and Glaucomatous Brown Norway Rats Optic Nerves.

Purpose

Methods

Results

Conclusions

Translational relevance

Publications

Morphometric Analysis of Retinal Ganglion Cell Axons in Normal and Glaucomatous Brown Norway Rats Optic Nerves.

Similar Datasets

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