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Updated: Mar 13, 2026
A Mouse Model of Retinal Ischemia-Reperfusion Injury Through Elevation of Intraocular Pressure
Published on: July 14, 2016
Ischemia Reperfusion Injury Triggers TNFα Induced-Necroptosis in Rat Retina
Cho Rong Kim1, Jie Hyun Kim1, Hae-Young Lopilly Park1
1a Department of Ophthalmology and Visual Science , Seoul St. Mary's Hospital, College of Medicine, The Catholic University of Korea , Seoul , Korea.
Purpose:
A recent study revealed a novel form of cell death, termed necroptosis, or programmed necrosis. Previous research indicated that after ischemia-reperfusion (IR) injury to the retina, Tumor Necrosis Factor α (TNFα) is increased, which may activate necroptosis. This study observed macroglial cell activation, and in particular, astrocyte activation, after the release of TNFα and other necroptosis factors in the rat retina due to IR.
Materials And Methods:
IR was induced in the retinas of adult male Sprague-Dawley rats by increasing the intraocular pressure to 160 mmHg and then allowing reperfusion. In addition, to inhibit necroptosis, Nec-1 (necrostatin-1) was injected intravitreally after IR. Rats were sacrificed after reperfusion at 12 hours, 1, 3, and 5 days, and 1 and 2 weeks. Retinas from each time point were analyzed by immunohistochemistry (IHC) and Western blotting (WB) to identify the initiator of necroptosis, TNFα, the expression of necroptosis factors, such as receptor interacting protein (RIP) 1, 3, and inactive caspase 8, and Brn3a.
Results:
Cell death in the IR-injured retinas was identified by cell counting. We found decreased retinal cell numbers in the inner and outer nuclear layers (INL and ONL), as well as in the ganglion cell layer (GCL). Increased glial cell activation was detected by using glial fibrillary acidic protein (GFAP) IHC. TNFα, RIP1, RIP3, and inactive caspase 8 were mainly expressed in the GCL after IR, as determined by IHC and WB. Nec-1 inhibited RIP1, a necroptosis factor, indicating protection against retinal cell loss after IR injury.
Conclusions:
We showed that IR injury triggered increases in both activation of astrocytes and the expression of TNFα. In addition, TNFα, which was activated by IR, triggered the release of necroptosis factors, particularly, in GCL. Inhibition of necroptosis using Nec-1 decreased the level of RIP1 and retinal cell loss in IR-injured retinas.
Insights
Ischemia-reperfusion injury activates necroptosis in the retina, leading to cell death. Inhibiting necroptosis with Nec-1 protects retinal cells from this damage, offering a potential therapeutic strategy.
Area of Science:
- Ophthalmology
- Neuroscience
- Cell Biology
Background:
- Ischemia-reperfusion (IR) injury is a significant cause of retinal damage.
- Tumor Necrosis Factor alpha (TNFα) is implicated in IR-induced retinal injury.
- Necroptosis, a form of programmed necrosis, is a potential mechanism of cell death in the retina.
Purpose of the Study:
- To investigate the role of necroptosis in IR-induced retinal injury.
- To examine the activation of necroptosis factors, including TNFα, RIP1, and RIP3, in the rat retina after IR.
- To evaluate the potential of necrostatin-1 (Nec-1) in inhibiting necroptosis and protecting retinal cells.
Main Methods:
- Induction of IR injury in rat retinas by increasing intraocular pressure.
- Intravitreal injection of Nec-1 to inhibit necroptosis.
- Analysis of retinal tissues using immunohistochemistry (IHC) and Western blotting (WB) at various time points post-IR.
- Assessment of cell death, glial cell activation (GFAP), and expression of necroptosis markers (TNFα, RIP1, RIP3, caspase 8).
Main Results:
- IR injury led to significant retinal cell loss in the inner and outer nuclear layers and the ganglion cell layer.
- Increased activation of astrocytes and expression of TNFα were observed after IR.
- TNFα, RIP1, RIP3, and inactive caspase 8 were primarily expressed in the ganglion cell layer post-IR.
- Nec-1 treatment inhibited RIP1 expression and reduced retinal cell loss.
Conclusions:
- IR injury activates astrocytes and upregulates TNFα in the retina.
- Activated TNFα triggers the release of necroptosis factors, particularly in the ganglion cell layer.
- Inhibition of necroptosis with Nec-1 mitigates RIP1 levels and protects against retinal cell loss in IR-injured retinas.
