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Yalong Dang, Susannah Waxman, Ralitsa Loewen, Adrianna Jensen, CHAO WANG, Ming Sun, Nils Loewen; An ex vivo pigment dispersion glaucoma perfusion model. Invest. Ophthalmol. Vis. Sci. 2017;58(8):3147.
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© ARVO (1962-2015); The Authors (2016-present)
Pigment dispersion glaucoma (PDG) is a common form of secondary open angle glaucoma with early age of onset, high intraocular pressure (IOP) and higher resistance to treatment than primary open angle glaucoma. The aim of this study was to develop and characterize a realistic ex vivo PDG model to develop novel, specific therapies.
Anterior segments of twenty-four porcine eyes were dissected, mounted, and cultured in constant-rate perfusion units. We produced size (1.03 +/-0.11) micron pigment granules by freeze-thaw, fragmentation, filtration and washing iris fragments. After IOP stabilization, eyes in the treatment group were perfused with pigment granules (1.67*10^7 particles/ml) for 196 hours. Control samples continued perfusion with pigment-free media. IOP was recorded continually. The TM was examined using hematoxylin and eosin stained sections and electron microscopy. We quantified how phagocytosis, adhesion, migration, and cytoskeleton changes in vitro when challenged with pigment. To further investigate pathways and mechanisms, a gene expression (GeneChip® Pico Kit, Affymetrix) and pathway analysis (Ingenuity Pathway Analysis) was performed.
Pigment granules generated by free-thaw method had no cytotoxic effects to primary TM cells at a concentration of 1.67*10^7 particles per ml. Baseline IOPs were (16.8 +/- 7.3) mmHg and began to elevate after 12 hours of pigment treatment. IOP peaked at 72 hours (29.4 +/- 8.1 mmHg) and remained steady to the end of this study (Figure 1). Histologically, pigment granules did not cause a physical outflow obstructions in between TM cells but were instead phagocytosed and aggregated around cell nuclei. Electron microscopy indicated TM cell apoptosis and lysosome activation. Significant decrease of phagocytosis, disruption of actin cytoskeleton, decrease of cell-cell and cell-matrix adhesion, and increase of TM migration in vitro and ex vivo were observed. The gene microarray and pathway analysis revealed 1025 genes to be upregulated or downregulated more than 1.5 fold compared to the control that were associated with cell phagocytosis, migration, cytoskeleton, and extracellular matrix formation (Figure 2).
We developed an affordable PDG ex vivo model that resembles human PDG. The discovery of decreased phagocytosis and increased migration suggest that manipulation of these pathways can be examined in this model to evolve disease-specific therapies.
This is an abstract that was submitted for the 2017 ARVO Annual Meeting, held in Baltimore, MD, May 7-11, 2017.
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