QuPath Automated Analysis of Optic Nerve Degeneration in Brown Norway Rats

Area of Science:

• Neuroscience
• Ophthalmology
• Digital Pathology

Background:

• Optic nerve (ON) degeneration is a hallmark of various neurological disorders, including glaucoma.
• Accurate morphological analysis of ON pathology is crucial for understanding disease progression and evaluating therapeutic interventions.
• Current methods for ON analysis can be time-consuming and subjective.

Purpose of the Study:

• To introduce and validate a novel application of QuPath open-source software for in-depth morphological analysis of progressive optic nerve degeneration in rats.
• To assess the utility of QuPath in quantifying axon and glial morphology in relation to ON damage severity.

Main Methods:

• QuPath software was adapted to analyze toluidine blue-stained rat ON light micrographs.
• QuPath was used to measure axon numbers, density, size distributions, and gliotic areas.
• QuPath results were compared against manual counting, AxonJ software, and electron microscopy estimates.

Main Results:

• QuPath accurately quantified decreased axon number and density, and increased gliotic areas with progressive ON damage.
• Axon density showed a strong negative correlation with gliotic areas.
• Large axons were more susceptible to degeneration, with an increase in swollen axons observed in moderate damage.
• QuPath counts demonstrated high correlation with manual counts and robust correlation with electron microscopy.

Conclusions:

• QuPath provides a reliable and automated method for analyzing axon loss, morphological changes, and gliotic expansion in degenerating optic nerves.
• The QuPath approach offers unbiased, in-depth analysis essential for reproducible preclinical studies of new glaucoma treatments.