Evidence of Hypoxic Glial Cells in a Model of Ocular Hypertension

Abstract

PurposeReoxygenation after hypoxia can increase reactive oxygen species and upregulate autophagy. We determined, for the first time, the impact of elevated IOP on hypoxia induction, superoxide accumulation, and autophagy in a bead model of glaucoma.MethodOcular hypertension was achieved with magnetic bead injection into the anterior chamber. Before mice were killed, they were injected with pimonidazole for hypoxia detection and dihydroethidium (DHE) for superoxide detection. Total retinal ganglion cells (RGCs) and optic nerve (ON) axons were quantified, total glutathione (GSH) was measured, and retinal and ON protein and mRNA were analyzed for hypoxia (Hif-1α and Hif-2α), autophagy (LC3 and p62), and SOD2.ResultsWith IOP elevation (P < 0.0001), the retina showed significantly (P < 0.001) decreased GSH compared with control, and a significant decrease (P < 0.01) in RGC density compared with control. Pimonidazole-positive Müller glia, microglia, astrocytes, and RGCs were present in the retinas after 4 weeks of ocular hypertension but absent in both the control and after only 2 weeks of ocular hypertension. The ON showed significant axon degeneration (P < 0.0001). The mean intensity of DHE in the ganglion cell layer and ON significantly increased (P < 0.0001). The ratio of LC3-II to LC3-I revealed a significant increase (P < 0.05) in autophagic activity in hypertensive retinas compared with control.ConclusionsWe report a novel observation of hypoxia and a significant decrease in GSH, likely contributing to superoxide accumulation, in the retinas of ocular hypertensive mice. The significant increase in the ratio of LC3-II to LC3-I suggests autophagy induction.

Endothelin 1-induced retinal ganglion cell death is largely mediated by JUN activation

“…4b, c). Hypoxic glia and RGCs were also reported after induction of ocular hypertension, suggesting the potential importance of hypoxia in glaucoma pathology 45,46 . RGC death did not necessarily occur in close proximity to major vessels after EDN1 insult, but it is possible that loss of blood flow or damage to minor vessels and capillary beds contributes to EDN pathology.…”

“…EDN ligands cause vasoconstriction by binding to EDNRA expressed by vascular smooth muscle cells 4,7,8,43,44 . Thinner retinal vessels and evidence of hypoxia have been demonstrated in animal models of chronic glaucoma 5,6,45,46 . Importantly, JUN was found be an important regulator of ischemia-induced neuronal death [47][48][49][50][51] .…”

Autoantibodies Activating the β2-Adrenergic Receptor Characterize Patients With Primary and Secondary Glaucoma

“…Recent data showed that just an elevated IOP induced oxidative stress (via reduction in glutathione), axon degeneration of the optic nerve head and autophagy in the retinas. Thus, hypoxic glial cells can be detected in animals with OHT, even in the absence of a glaucomatous pathology (39). β2AR of astrocytes were also involved in the regulation of the glucose metabolism (40, 41) and are discussed to contribute to neuronal degeneration (42, 43).…”

Retinal Ganglion Cell Degeneration in a Rat Magnetic Bead Model of Ocular Hypertensive Glaucoma

“…Using a rat laser photocoagulation model of OHT, Chidlow et al 47 reported a highly variable degree of axon transport disruption and pimonidazole labeling that did not correspond statistically with the degree of IOP elevation. Furthermore, Jassim and Inman 23 observed no pimonidazole labeling in the retina 2 weeks after the induction of ocular hypertension in a mouse bead model of OHT; yet, cell loss was as great as 50% in some eyes. In this study, numerous pimonidazole-positive glia were detected at 4 weeks.…”

“… 16 – 18 To our knowledge, we were the first to propose the use of paramagnetic microparticles that could be directed with a magnet to the iridocorneal angle, allowing for control of distribution and avoidance of disruption to vision through occlusion of the pupil. 19 We have since used this model to investigate RGC dendritic atrophy, 20 , 21 and others have adapted its use for mice, 22 , 23 but a detailed characterization of the rat model has yet to be performed. We use the Brown Norway rat ( Rattus norvegicus ) because (1) it is the rat genetic standard (and as such will allow for detailed -omics analysis (e.g., KEGG pathways based on Brown Norway gene annotations); (2) their temperament is amenable to ophthalmic examination without the need for restraint or anesthesia, particularly with regard to recording reliable tonometry measurements; and (3) they are pigmented and as such avoid potential confounders present in albino rodents (e.g., Wistar and Sprague–Dawley rats), which have reduced ipsilateral projections.…”