7, 8-Dihydroxyflavone, a TrkB receptor agonist, provides minimal protection against retinal vascular damage during oxygen-induced ischemic retinopathy

Retinopathy of prematurity (ROP) is one of the main causes of blindness in children worldwide. Brain-derived neurotrophic factor (BDNF) and its receptor, tropomyosin-related kinase B (TrkB), play critical protective roles in the development and function of neurons and vasculature. Lack of BDNF expression results in increased endothelial cell apoptosis and reduced endothelial cell-cell contact. Premature babies who develop ROP tend to have lower serum BDNF levels. BDNF expression is also significantly lower in mouse retinas following exposure to hyperoxia compared to those reared in room air. Specifically, BDNF promotes angiogenic tube formation of endothelial cells (EC), and it is considered an EC survival factor required for stabilization of intramyocardial vessels. We hypothesized that the activation of TrkB receptor protects retinal vasculature in the mice during oxygen-induced ischemic retinopathy (OIR), a model of ROP. To test this hypothesis, we treated neonatal mice with 7,8-dihydroxyflavone (DHF) (5 mg/kg body weight), a TrkB receptor agonist. We examined its potential protective effects on retinal vessel obliteration and neovascularization, two hallmarks of ROP and OIR. We found that retinas from DHF treated postnatal day 8 (P8) and P12 mice have similar levels of vessel obliteration as retinas from age-matched control mice subjected to OIR. Similarly, DHF showed no significant effect on mitigation of retinal neovascularization during OIR in P17 mice. Collectively, our studies demonstrate that the TrkB receptor agonist DHF provides no significant protective effects during OIR.

Mural cell SRF controls pericyte migration, vessel patterning and blood flow

Rationale: Pericytes (PCs) and vascular smooth muscle cells (vSMCs), collectively known as mural cells(MCs), are recruited through PDGFB-PDGFRB signaling. MCs are essential for vascular integrity, and their loss has been associated with numerous diseases. Most of this knowledge is based on studies in which MCs are insufficiently recruited or fully absent upon inducible ablation. In contrast, little is known about the physiological consequences that result from impairment of specific MC functions. Objective: Here, we characterize the role of the transcription factor serum response factor (SRF) in MCs and study its function in developmental and pathological contexts. Methods and Results: We generated a mouse model of MC-specific inducible Srf gene deletion and studied its consequences during retinal angiogenesis. By postnatal day (P)6, PCs lacking SRF were morphologically abnormal and failed to properly co-migrate with angiogenic sprouts. As a consequence, PC-deficient vessels at the retinal sprouting front became dilated and leaky. By P12, also the vSMCs had lost SRF, which coincided with the formation of pathological arteriovenous (AV) shunts. Mechanistically, we show that PDGFB-dependent SRF activation is mediated via MRTF co-factors. We further show that MRTF-SRF signaling promotes pathological PC activation during ischemic retinopathy. RNA-sequencing, immunohistology, in vivo live imaging and in vitro experiments demonstrated that SRF regulates expression of contractile SMC proteins essential to maintain the vascular tone. Conclusions: SRF is crucial for distinct functions in PCs and vSMCs. SRF directs PC migration downstream of PDGFRB signaling and mediates pathological PC activation during ischemic retinopathy. In vSMCs, SRF is essential for expression of the contractile machinery, and its deletion triggers formation of AV shunts. These essential roles in physiological and pathological contexts provide a rational for novel therapeutic approaches through targeting SRF activity in MCs.

Effect of anti-VEGF treatment on nonperfusion areas in ischemic retinopathy

In recent years, retinal ischemia such as that which occurs in diabetic retinopathy (DR) and retinal vein occlusion (RVO) has become a hotspot of ischemic retinopathy research. High levels of vascular endothelial growth factor (VEGF) are recognized as a major cause of macular edema (ME) in DR and RVO. High concentrations of VEGF in the vitreous can lead to serious retinal ischemia and hypoxia and form retinal nonperfusion areas (NPAs). Different levels of retinal ischemia can represent disease severity and progression. Anti-VEGF therapy as the first-line treatment for ME has been found to be effective in improving vision, but there are still disputes about whether anti-VEGF therapy could improve retinal ischemia and achieve reperfusion of previously developed retinal NPAs. Here, we review and summarize studies of the effects of anti-VEGF drugs on retinal ischemia, especially NPAs.

Retinopathy of prematurity shows alterations in Vegfa164 isoform expression

Background Pathologic ocular neovascularization in retinopathy of prematurity (ROP) and other proliferative retinopathies are characterized by dysregulation of vascular endothelial growth factor-A (VEGF-A). A study of Vegfa isoform expression during oxygen-induced ischemic retinopathy (OIR) may enhance our understanding of Vegf dysregulation. Methods Following induction of OIR, immunohistochemistry and polymerase chain reaction (PCR) was performed on room air (RA) and OIR mice. Results Total Vegfa messenger RNA (mRNA) expression was stable in RA mice, but increased in OIR mice with a peak at postnatal day 17 (P17), before returning to RA levels. Vegfa164a expression was similar in both OIR and RA mice at P10 (Phase 1 OIR), but 2.4-fold higher in OIR mice compared to RA mice at P16 (Phase 2 OIR). At P10, Vegfa164b mRNA was similar in OIR vs RA mice, but was expressed 2.5-fold higher in OIR mice compared to RA mice at P16. At P10 and P16, Vegfr2/Vegfr1 expression was increased in OIR mice compared to RA mice. Increased activation of microglia was seen in OIR mice. Conclusions Vegfa164a, Vegfa164b, and Vegfr1 were overexpressed in OIR mice, leading to abnormal signaling and angiogenesis. Furthers studies of mechanisms of Vegf dysregulation may lead to novel therapies for ROP and other proliferative retinopathies. Impact Vegfa164 has two major isoforms, a proangiogenic, Vegfa164a, and an antiangiogenic, Vegfa164b, with opposing receptors, inhibitory Vegfr1, and stimulatory Vegfr2, but their role in OIR is unclear. In Phase 1 OIR, both isoforms and receptors are expressed similarly. In Phase 2 OIR, both isoforms are overexpressed, with an increased ratio of inhibitory Vegfr1. Modulation of angiogenesis by Vegf regulation enables pruning of excess angiogenesis during physiology, but results in ineffective angiogenesis during OIR. Knowledge of VEGF dysregulation may have novel therapeutic implications in the management of ROP and retinal proliferative diseases.

Retinal macroaneurysm after carotid endarterectomy in patient with chronic ischemic retinopathy associated with hemodynamically significant stenosis of the internal carotid artery (clinical case)

Purpose. To present a clinical case of development of retinal macroaneurysm after carotid endarterectomy (CE) in patient with chronic ischemic retinopathy (CIR) associated with hemodynamically significant stenosis of the internal carotid artery (ICA). Material and methods. Patient R., 74 years old, diagnosed with CIR of the 1st degree of severity of the right eye. In addition to standard ophthalmic examination methods, Doppler ultrasound with registration of blood flow in the orbital artery and spiral computed tomography of the ICA were performed. Results. Panretinal photocoagulation (PRP) of the retina led to obliteration of the retinal macroaneurysm, resorption of intraretinal hemorrhages, and stabilization of the course of CIR after CE of the ICA. Conclusion. The appearance of retinal macroaneurysm and intraretinal hemorrhages after CE in patient with CIR associated with hemodynamically significant ICA stenosis can be regarded as reperfusion complications after restoration of blood flow in the orbital artery, which requires timely retinal PRP of the retina. Key words: chronic ischemic retinopathy, retinal macroaneurysm, internal carotid artery, carotid endarterectomy.

The Regulatory NOD-Like Receptor NLRC5 Promotes Ganglion Cell Death in Ischemic Retinopathy by Inducing Microglial Pyroptosis

Retinal ischemia is a common pathological event that can result in retinal ganglion cell (RGC) death and irreversible vision loss. The pathogenic mechanisms linking retinal ischemia to RGC loss and visual deficits are uncertain, which has greatly hampered the development of effective treatments. It is increasingly recognized that pyroptosis of microglia contributes to the indirect inflammatory death of RGCs. In this study, we report a regulatory NOD-like receptor, NOD-, LRR- and CARD-containing 5 (NLRC5), as a key regulator on microglial pyroptosis and the retinal ischemia process. Through an in-depth analysis of our recently published transcriptome data, we found that NLRC5 was significantly up-regulated in retina during ischemia–reperfusion injury, which were further confirmed by subsequent detection of mRNA and protein level. We further found that NLRC5 was upregulated in retinal microglia during ischemia, while NLRC5 knockdown significantly ameliorated retinal ischemic damage and RGC death. Mechanistically, we revealed that knockdown of NLRC5 markedly suppressed gasdermin D (GSDMD) cleavage and activation of interleukin-1β (IL-1β) and caspase-3, indicating that NLRC5 promotes both microglial pyroptosis and apoptosis. Notably, we found that NLRC5 directly bound to NLRP3 and NLRC4 in inflammasomes to cooperatively drive microglial pyroptosis and apoptosis mediating retinal ischemic damage. Overall, these findings reveal a previously unidentified key contribution of NLRC5 signaling to microglial pyroptosis under ischemia or hypoxia conditions. This NLRC5-dependent pathway may be a novel therapeutic target for treatment of ischemic retinopathy.

Blocking the interaction between interleukin-17A and endoplasmic reticulum stress in macrophage attenuates retinal neovascularization in oxygen-induced retinopathy

Background Neovascularization is a leading cause of visual loss typically associated with diabetic retinopathy (DR) and retinopathy of prematurity (ROP). Interleukin-17A (IL-17A) and endoplasmic reticulum (ER) stress both have been demonstrated to play a proangiogenic role in ischemic retinopathies. However, the relationship between IL-17A and ER stress in retinal neovascularization (RNV) under hypoxic conditions and its underlying mechanisms remain unclear. Methods In this study, oxygen-induced retinopathy (OIR) mice model was established and intravitreal injections were conducted. Changes of IL-17A and ER stress markers in retinas and cultured primary bone marrow derived macrophage (BMDM) under normoxic or hypoxic conditions were detected. Western blotting, Real-Time RT-PCR, Immunofluorescence assays were conducted to explore the roles and relationship of IL-17A and ER stress in RNV, as well as its underlying mechanisms. Results Compared to that in normal controls, IL-17A and ER stress markers were all remarkably increased under hypoxic conditions both in vivo and in vitro. Neutralization or knock out of IL-17A decreased ER stress. ER stress inhibitor 4-phenylbutyrate (4-PBA), attenuated the production of IL-17A, suggesting a positive feedback loop between IL-17A and ER stress. Inhibition of IL-17A or ER stress decreased areas of nonperfusion and neovascularization in OIR retinas. As TXNIP/NLRP3 pathway activation has been demonstrated to be involved in increased retinal vascular permeability of ischemic retinopathy, we observed that TXNIP/NLRP3 pathway mediated in the interaction between IL-17A and ER stress under hypoxic conditions. Conclusion The interplay between IL-17A and ER stress contributes to RNV in macrophages via modulation of TXNIP/NLRP3 signaling pathway under hypoxic conditions. The feedback loops may become an innovative and multiple pharmacological therapeutic target for ischemic retinopathy.