Qupath automated analysis of optic nerve degeneration in brown Norway rats

Barbara A. Mysona, Sharmila Segar, Cecilia Hernandez, Christian Kim, Jing Zhao, David Mysona, Kathryn E. Bollinger

Research output: Contribution to journalArticle

Abstract

Purpose: A novel application of QuPath open-source digital analysis software is used to provide in-depth morphological analysis of progressive optic nerve (ON) degeneration in rats. Methods: QuPath software was adapted to assess axon and gliotic morphology in toluidine blue-stained, Brown Norway rat ON light micrographs. QuPath axon numbers, density, size distributions, and gliotic areas were obtained from test images and ON cross-sections separated by damage grade. QuPath results were compared with manual counting, AxonJ, and electron microscopy axon estimates. Results: QuPath-derived axon number, density, and diameter decreased with increasing ON damage. Axon density negatively correlated with gliotic areas in test images (R2 = 0.759; P < 0.0001; N = 40) and in ON cross-sections (R2 = 0.803; P < 0.0004; N = 10). Although axon losses occurred across most axon diameters, large axons were more susceptible to degeneration. The exception was swollen axons > 2 μm, which increased in moderately but not severely damaged images. QuPath axon counts correlated strongly with manual counts of test images (R2 = 0.956; P < 0.0001). QuPath outperformed AxonJ on test images and total ON axon counts. Compared to electron microscopy analysis, QuPath undercounted ON axons; however, correlation between the methods was robust (R2 = 0.797; P < 0.001; N = 10). Conclusions: QuPath analysis reliably identified axon loss, axon morphology changes, and gliotic expansion that occurred in degenerating ONs.

Original languageEnglish (US)
Article number22
JournalTranslational Vision Science and Technology
Volume9
Issue number3
DOIs
StatePublished - Jan 1 2020

Keywords

  • Axons
  • Glaucoma
  • Gliosis
  • Image analysis
  • Optic nerve degeneration

ASJC Scopus subject areas

  • Biomedical Engineering
  • Ophthalmology

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