Retinal Ischemia-reperfusion Injury Research Articles

Role of Curcumin in Retinal Diseases-A review.

PurposeTo review the role of curcumin in retinal diseases, COVID era, modification of the molecule to improve bioavailability and its future scope.MethodsPubMed and MEDLINE searches were pertaining to curcumin, properties of curcumin, curcumin in retinal diseases, curcumin in diabetic retinopathy, curcumin in age-related macular degeneration, curcumin in retinal and choroidal diseases, curcumin in retinitis pigmentosa, curcumin in retinal ischemia reperfusion injury, curcumin in proliferative vitreoretinopathy and curcumin in current COVID era.ResultsIn experimental models, curcumin showed its pleiotropic effects in retinal diseases like diabetic retinopathy by increasing anti-oxidant enzymes, upregulating HO-1, nrf2 and reducing or inhibiting inflammatory mediators, growth factors and by inhibiting proliferation and migration of retinal endothelial cells in a dose-dependent manner in HRPC, HREC and ARPE-19 cells. In age-related macular degeneration, curcumin acts by reducing ROS and inhibiting apoptosis inducing proteins and cellular inflammatory genes and upregulating HO-1, thioredoxin and NQO1. In retinitis pigmentosa, curcumin has been shown to delay structural defects of P23H gene in P23H-rhodopsin transgenic rats. In proliferative vitreoretinopathy, curcumin inhibited the action of EGF in a dose- and time-dependent manner. In retinal ischemia reperfusion injury, curcumin downregulates IL-17, IL-23, NFKB, STAT-3, MCP-1 and JNK. In retinoblastoma, curcumin inhibits proliferation, migration and apoptosis of RBY79 and SO-RB50. Curcumin has already proven its efficacy in inhibiting viral replication, coagulation and cytokine storm in COVID era.ConclusionCurcumin is an easily available spice used traditionally in Indian cooking. The benefits of curcumin are manifold, and large randomized controlled trials are required to study its effects not only in treating retinal diseases in humans but in their prevention too.

7,8-Dihydroxyflavone alleviates apoptosis and inflammation induced by retinal ischemia-reperfusion injury via activating TrkB/Akt/NF-kB signaling pathway.

Retinal ischemia-reperfusion injury (RIRI) is of common occurrence in retinal and optic nerve diseases. The BDNF/TrkB signaling pathway has been examined to be neuroprotective in RIRI. In this study, we investigated the role of a potent selective TrkB agonist 7,8-dihydroxyfavone (DHF) in rat retinas with RIRI. Our results showed that RIRI inhibited the conversion of BDNF precursor (proBDNF) to mature BDNF (mBDNF) and increased the level of neuronal cell apoptosis. Compared with RIRI, DHF+RIRI reduced proBDNF level and at the same time increased mBDNF level. Moreover, DHF administration effectively activated TrkB signaling and and downstream Akt and Erk signaling pathways which increased nerve cell survival. The combined effects of mBDNF/proBDNF increase and TrkB signaling activation lead to reduction of apoptosis level and protection of retinas with RIRI. Moreover, it was also found that astrocytes labeled by GFAP were activated in RIRI and NF-kB mediated the increased expressions of inflammatory factors and these effects were partially reversed by DHF administration. Besides, we also used RNA sequencing to analyze the differently expressed genes (DEGs) and their enriched (Kyoto Encyclopedia of Genes and Genomes) KEGG pathways between Sham, RIRI, and DHF+RIRI. It was found that 1543 DEGs were differently expressed in RIRI and 619 DEGs were reversed in DHF+RIRI. The reversed DEGs were typically enriched in PI3K-Akt signaling pathway, Jak-STAT signaling pathway, NF-kB signaling pathway, and Apoptosis. To sum up, the DHF administration alleviated apoptosis and inflammation induced by RIRI via activating TrkB signaling pathway and may serve as a promising drug candidate for RIRI related ophthalmopathy.

Preclinical investigation of Pegylated arginase 1 as a treatment for retina and brain injury

Arginase 1 (A1) is the enzyme that hydrolyzes the amino acid, L-arginine, to ornithine and urea. We have previously shown that A1 deletion worsens retinal ischemic injury, suggesting a protective role of A1. In this translational study, we aimed to study the utility of systemic pegylated A1 (PEG-A1, recombinant human arginase linked to polyethylene glycol) treatment in mouse models of acute retinal and brain injury. Cohorts of WT mice were subjected to retinal ischemia-reperfusion (IR) injury, traumatic optic neuropathy (TON) or brain cerebral ischemia via middle cerebral artery occlusion (MCAO) and treated with intraperitoneal injections of PEG-A1 or vehicle (PEG only). Drug penetration into retina and brain tissues was measured by western blotting and immunolabeling for PEG. Neuroprotection was measured in a blinded fashion by quantitation of NeuN (neuronal marker) immunolabeling of retina flat-mounts and brain infarct area using triphenyl tetrazolium chloride (TTC) staining. Furthermore, ex vivo retina explants and in vitro retina neuron cultures were subjected to oxygen-glucose deprivation (OGD) followed by reoxygenation (R) and treated with PEG-A1. PEG-A1 given systemically did not cross the intact blood-retina/brain barriers in sham controls but reached the retina and brain after injury. PEG-A1 provided neuroprotection after retinal IR injury, TON and cerebral ischemia. PEG-A1 treatment was also neuroprotective in retina explants subjected to OGD/R but did not improve survival in retinal neuronal cultures exposed to OGD/R. In summary, systemic PEG-A1 administration is neuroprotective and provides an excellent route to deliver the drug to the retina and the brain after acute injury.

Maintaining blood retinal barrier homeostasis to attenuate retinal ischemia-reperfusion injury by targeting the KEAP1/NRF2/ARE pathway with lycopene

Retinal ischemia-reperfusion (I/R) often results in intractable visual impairments, where blood retinal barrier (BRB) homeostasis mediated by retinal pigment epithelium (RPE) and retinal microvascular endothelium (RME) is crucial. However, strategies targeting the BRB are limited. Thus, we investigated the inconclusive effect of lycopene (LYC) in retinal protection under I/R. LYC elevated cellular viability and reversed oxidative stress in aRPE-19 cells/hRME cells under I/R conditions based on oxygen-glucose deprivation (OGD) in vitro. Molecular analysis showed that LYC promoted NRF2 expression and enhanced the downstream factors of the KEAP1/NRF2/ARE pathway: LYC increased the activities of antioxidants, including SOD and CAT, whereas it enhanced the mRNA expression of HO-1 (ho-1) and NQO-1 (nqo-1). The activation resulted in restrained ROS and MDA. On the other hand, LYC ameliorated the damage to retinal function and morphology in a mouse I/R model, which was established by unilateral ligation of the left pterygopalatine artery/external carotid artery and reperfusion. LYC promoted the expression of NRF2 in both the neural retina and the RPE choroid in vivo. This evidence revealed the potential of LYC in retinal protection under I/R, uncovering the pharmacological effect of the KEAP1/NRF2/ARE pathway in BRB targeting. The study generates new insights into scientific practices in retinal research.

Hydrogen Sulfide Reduces Ischemia and Reperfusion Injury in Neuronal Cells in a Dose- and Time-Dependent Manner.

Background: The ischemia-reperfusion injury (IRI) of neuronal tissue, such as the brain and retina, leads to possible cell death and loss of function. Current treatment options are limited, but preliminary observations suggest a protective effect of hydrogen sulfide (H2S). However, the dosage, timing, and mechanism of inhaled H2S treatment after IRI requires further exploration. Methods: We investigated possible neuroprotective effects of inhaled H2S by inducing retinal ischemia–reperfusion injury in rats for the duration of 1 h (120 mmHg), followed by the administration of hydrogen sulfide (H2S) for 1 h at different time points (0, 1.5, and 3 h after the initiation of reperfusion) and at different H2S concentrations (120, 80, and 40 ppm). We quantified the H2S effect by conducting retinal ganglion cell counts in fluorogold-labeled animals 7 days after IRI. The retinal tissue was harvested after 24 h for molecular analysis, including qPCR and Western blotting. Apoptotic and inflammatory mediators, transcription factors, and markers for oxidative stress were investigated. Histological analyses of the retina and the detection of inflammatory cytokines in serum assays were also performed. Results: The effects of inhaled H2S were most evident at a concentration of 80 ppm administered 1.5 h after IRI. H2S treatment increased the expression of anti-apoptotic Bcl-2, decreased pro-apoptotic Bax expression, reduced the release of the inflammatory cytokines IL-1β and TNF-α, attenuated NF-κB p65, and enhanced Akt phosphorylation. H2S also downregulated NOX4 and cystathionine β-synthase. Histological analyses illustrated a reduction in TNF-α in retinal ganglion cells and lower serum levels of TNF-α in H2S-treated animals after IRI. Conclusion: After neuronal IRI, H2S mediates neuroprotection in a time- and dose-dependent manner. The H2S treatment modulated transcription factor NF-κB activation and reduced retinal inflammation.

Inflammatory resolution and vascular barrier restoration after retinal ischemia reperfusion injury

BackgroundSeveral retinal pathologies exhibit both inflammation and breakdown of the inner blood-retinal barrier (iBRB) resulting in vascular permeability, suggesting that treatments that trigger resolution of inflammation may also promote iBRB restoration.MethodsUsing the mouse retinal ischemia-reperfusion (IR) injury model, we followed the time course of neurodegeneration, inflammation, and iBRB disruption and repair to examine the relationship between resolution of inflammation and iBRB restoration and to determine if minocycline, a tetracycline derivative shown to reverse microglial activation, can hasten these processes.ResultsA 90-min ischemic insult followed by reperfusion in the retina induced cell apoptosis and inner retina thinning that progressed for approximately 2 weeks. IR increased vascular permeability within hours, which resolved between 3 and 4 weeks after injury. Increased vascular permeability coincided with alteration and loss of endothelial cell tight junction (TJ) protein content and disorganization of TJ protein complexes. Shunting of blood flow away from leaky vessels and dropout of leaky capillaries were eliminated as possible mechanisms for restoring the iBRB. Repletion of TJ protein contents occurred within 2 days after injury, long before restoration of the iBRB. In contrast, the eventual re-organization of TJ complexes at the cell border coincided with restoration of the barrier. A robust inflammatory response was evident a 1 day after IR and progressed to resolution over the 4-week time course. The inflammatory response included a rapid and transient infiltration of granulocytes and Ly6C+ classical inflammatory monocytes, a slow accumulation of Ly6Cneg monocyte/macrophages, and activation, proliferation, and mobilization of resident microglia. Extravasation of the majority of CD45+ leukocytes occurred from the superficial plexus. The presence of monocyte/macrophages and increased numbers of microglia were sustained until the iBRB was eventually restored. Intervention with minocycline to reverse microglial activation at 1 week after injury promoted early restoration of the iBRB coinciding with decreased expression of mRNAs for the microglial M1 markers TNF-α, IL-1β, and Ptgs2 (Cox-2) and increased expression of secreted serine protease inhibitor Serpina3n mRNA.ConclusionsThese results suggest that iBRB restoration occurs as TJ complexes are reorganized and that resolution of inflammation and restoration of the iBRB following retinal IR injury are functionally linked.

Resveratrol attenuates retinal ganglion cell loss in a mouse model of retinal ischemia reperfusion injury via multiple pathways

BackgroundResveratrol (RES) is a natural polyphenol that has been shown to protect retinal ganglion cells (RGCs) following retinal ischemia reperfusion (I/R) injury. However, the molecular mechanisms of resveratrol function are yet to be fully elucidated. Thus, this study explored the potential mechanisms of resveratrol in vivo. MethodsA retinal ischemia reperfusion injury model was established in adult male C57BL/6 J mice. Intraperitoneal injection of resveratrol was administered continuously for 5 days. RGC survival was determined by immunofluorescence staining with Brn3a. Flash electroretinography (ERG) was conducted to assess visual function. Proteins of HIF-1a, VEGF, p38, p53, PI3K, Akt, Bax, Bcl2, and Cleaved Caspase3 were detected using Western blot. ResultsRES administration significantly ameliorated retinal thickness damage and increased Brn3a stained RGCs 7 days after I/R injury. We also found that administration of RES remarkably inhibited the upregulation of mitochondrial apoptosis-related protein Bax and Cleaved Caspase3, as well as increased the expression of Bcl2. Furthermore, RES administration significantly suppressed the I/R injury-induced upregulation of the HIF-1a/VEGF and p38/p53 pathways, while activating the I/R injury-induced downregulation of the PI3K/Akt pathway. Moreover, RES administration remarkably improved retinal function after I/R injury-induced functional impairment. ConclusionsOur data demonstrated that resveratrol can mitigate retinal ischemic injury induced RGC loss and retinal function impairment by inhibiting the HIF-1a/VEGF and p38/p53 pathways while activating the PI3K/Akt pathway. Therefore, our results further reinforce that resveratrol has potential for treating glaucoma.

Recent Advances and Disputes About Curcumin in Retinal Diseases.

Curcumin belongs to the group of so-called phytocompounds, biologically active molecules produced by plants exerting a beneficial effect on health. Curcumin shows a wide spectrum of different properties, being an anti-inflammatory, antioxidant, antimicrobial and antimutagenic molecule. The purpose of the review is to examine what literature reported on the characteristics of curcumin, particularly, on the beneficial and controversial aspects of this molecule, aiming for a better therapeutic management of retinal diseases. The retina is a constant target of oxidative stress, this tissue being characterized by cells rich in mitochondria and by vessels and being, obviously, continuously reached from photons affecting its layers. Particularly, the retinal ganglion cells and the photoreceptors are extremely sensitive to oxidative stress damage and it is well known that an imbalance in reactive oxygen species is often involved in several retinal diseases, such as uveitis, age-related macular degeneration, diabetic retinopathy, central serous chorioretinopathy, macular edema, retinal ischemia-reperfusion injury, proliferative vitreoretinopathy, hereditary tapeto-retinal degenerations, and retinal and choroidal tumors. To date, several studies suggest that oral treatment with curcumin is generally well tolerated in humans and, in addition, it seems to have no negative effects: therefore, curcumin is a promising candidate as a retinal disease therapy. Unfortunately, the primary limitation of curcumin is represented by its poor bioavailability, in fact only a minimal fraction of this substance can reach the blood stream in the form of a biologically active compound. However, many steps have been made in several fields. In the future, it is expected that the strategies developed until now to allow curcumin to reach the target tissues in adequate concentrations could be ameliorated and, above all, large in vivo studies on humans are needed to demonstrate the total safety of these compounds and their effectiveness in different eye diseases.

N-acetylserotonin alleviated the expression of interleukin-1β in retinal ischemia–reperfusion rats via the TLR4/NF-κB/NLRP3 pathway

This study aimed to explore the effects of N-acetylserotonin (NAS) on the expression of interleukin-1β (IL-1β) in the retina of retinal ischemia–reperfusion injury (RIRI) rats via the toll-like receptor 4 (TLR4)/nuclear factor-kappa B (NF-κB)/nod-like receptor pyrin domain containing 3 (NLRP3) signaling pathway. In this study, adult male Sprague Dawley rats were randomly divided into the sham, RIRI, RIRI + NAS and RIRI + TAK-242 + NAS groups. The rats in the RIRI + NAS and RIRI + TAK-242 + NAS groups were intraperitoneally injected with NAS 30 min before and after modeling. TAK-242, a selective TLR4 inhibitor, was administered by intraperitoneal injection in RIRI + TAK-242 + NAS group. The RIRI rat model was established by elevating the intraocular pressure to 110 mmHg for 60 min. The retinal structure and edema were assessed by H&E staining. The expression levels of TLR4, phosphorylated NF-κB (p–NF–κB), NLRP3, cleaved Caspase-1, and IL-1β in the retina of each group were detected using immunohistochemistry and Western blot. The correlations of the differences of TLR4+ and cleaved Caspase-1+ with IL-1β+ cells (between the NAS and the RIRI groups) were analyzed, using linear regression in the RIRI + NAS group. Results showed that thinner retina, more RGCs, and less TLR4+, p–NF–κB+, NLRP3+, cleaved Caspase-1+, and IL-1β+ cells in the retina were observed in the RIRI + NAS and RIRI + TAK-242 + NAS groups compared with the RIRI group 12 h after RIRI (all P < 0.01). Western blot analysis results showed that the expression of IL-1β in the RIRI + NAS group began to increase 6 h after RIRI, and it reached a high level 12 h after RIRI, and then decreased. And it was lower at each time point in the RIRI + NAS group than in the RIRI group, and there existed significant difference (all P < 0.01). Besides, the expression levels of TLR4, p–NF–κB, NLRP3, and cleaved Caspase-1 proteins in the RIRI + NAS and RIRI + TAK-242 + NAS groups decreased 12 h after RIRI compared with those in the RIRI group (all P < 0.01). The difference in IL-1β+ cells was significantly correlated with those of TLR4+ and cleaved Caspase-1+ cells in the RIRI + NAS group (r2 = 0.9054 or 0.7431, P < 0.01). In conclusion, NAS could attenuate the expression of IL-1β by inhibiting the TLR4/NF-κB/NLRP3 signaling pathway, reduce the retina edema, and promote the survival of RGCs, thereby alleviating the retinal injury and exert its neuroprotective effect.

Histological Assessment of Rat Retinas with Ischemia-Reperfusion Injury

Introduction: Retinal ischemia-reperfusion (IR) injury occurs in pathological situations that interrupt the blood flow to the retina, such as is the case during central retinal artery occlusion (CRAO). The animal models described in the literature are based on the pressure produced by the weight of a given quantity of saline elevated to a certain height; however, to establish these parameters it is necessary to perform mathematical calculations that cannot be easily redone in the case of punctual variations of intraocular pressure (IOP). The aim of this study was to present a new system that allows us to reproduce the conditions of retinal IR and thereby properly assess the level of injury in retinal histological samples. Methods: We developed a retinal IR model in WAG/RijHsd rats based on CRAO through increasing IOP. To develop this model, we produced ischemia for 1 h using a hydrostatic pressure system that maintained a constant high IOP and then allowed reperfusion for 1 h. The injury attributable to IR was assessed by histological examination of retinal samples, determining whether there was histological damage and/or dendritic swelling and counting the outer nuclear layer cells showing cytoplasmic swelling. Results: The increase in IOP to 150 mm Hg produced CRAO, in turn causing observable histological damage and dendritic swelling in all retinas subjected to IR. Counting the number of cells showing cytoplasmic swelling yielded a mean of 102.5 ± 35 cells/field. The contralateral retinas were healthy, showing no significant changes. Conclusion: The retinal IR model proposed is simple, reproducible, and allows variable durations of ischemia and reperfusion, and most importantly, it allows easy correction by adjusting the pressure of the sphygmomanometer, of any change in IOP to keep the ischemia stable, without having to recalculate the elevation height of the ischemia induction system. Moreover, the damage caused by IR can be effectively assessed by the type of histopathological assessment performed. For these reasons, it can be considered a reliable method for studying drugs that may prevent retinal IR injury.

Krüppel-like factor 7 protects retinal ganglion cells and promotes functional preservation via activating the Akt pathway after retinal ischemia-reperfusion injury

ObjectiveThe purpose of this study is to investigate the effects of Krüppel-like factor 7 (KLF7) on retinal ganglion cells (RGCs) and retinal function after retinal ischemia-reperfusion (RIR) injury in mice. MethodsMale C57BL/6J mice were intravitreally injected with recombinant adeno-associated vectors (rAAV-KLF7-EGFP or rAAV-EGFP), and subsequently used to induce RIR injury. Retinal cryosections were used to access the efficacy of virus transfection, 1, 2, 3, and 4 weeks after rAAV-KLF7-EGFP transfer. RGCs survival rate was observed and quantified by immunofluorescent staining, 7 days after RIR injury. Meanwhile, electroretinogram (ERG) and optomotor response were used to evaluate the electrophysiological functions and visual acuity. Apoptosis was evaluated by TUNEL staining 1 day after RIR injury. Expression of KLF7, Akt, phospho-Akt, Bcl-2, and Bax were further detected by western blot to excavate the underlying mechanism. ResultsThe transfection efficiency of rAAV-KLF7-EGFP was increased in a time-dependent manner, and the number of EGFP-positive cells was increased significantly 3 weeks after rAAV-KLF7-EGFP transfer. RGCs survival rates, amplitudes of ERG a-, b-wave, Ops, PhNR, and visual acuity of mice were decreased after RIR injury. With the increase of light intensity, the amplitudes of scotopic ERG a- and b-wave were gradually increased while the incubation period was gradually shortened. RGCs survival rates, amplitudes of ERG a-, b-wave, Ops, PhNR, and visual acuity of mice were increased after rAAV-KLF7-EGFP transfer. The protein level of KLF7 was up-regulated after rAAV-KLF7-EGFP transfer. Up-regulation of KLF7 significantly inhibited cells apoptosis, increased phospho-Akt and Bcl-2 expression, and decreased Bax expression. There were no significant changes in Akt expression. ConclusionOverexpression of KLF7 can not only prevent the loss of RGCs, but also preserve the electrophysiological function. In addition, overexpression of KLF7 can ameliorate the retinal dysfunction after RIR injury, and ultimately improve the visual acuity of mice. The activation of Akt pathway and the suppression of the mitochondrial apoptotic pathway contribute to the neuroprotection of KLF7.

CircRNA expression profile and functional analysis in retinal ischemia-reperfusion injury

Retinal ischemia-reperfusion (I/R) is involved in the pathogenesis of many vision-threatening diseases. circRNAs act as key players in gene regulation and human diseases. However, the global circRNA expression profile in retinal I/R injury has not been fully uncovered. Herein, we established a murine model of retinal I/R injury and performed circRNA microarrays to identify I/R-related circRNAs. 1265 differentially expressed circRNAs were identified between I/R retinas and normal retinas. Notably, the detection of cWDR37 level in aqueous humor could discriminate glaucoma patients from cataract patients (AUC = 0.9367). cWdr37 silencing protected against hypoxic stress- or oxidative stress-induced retinal ganglion cell (RGC) injury. cWdr37 silencing alleviated IR-induced retinal neurodegeneration as shown by increased NeuN staining, reduced retinal reactive gliosis, and decreased retinal apoptosis. Collectively, this study provides a novel insight into the pathogenesis of retinal I/R injury. cWdr37 is a promising target for the diagnosis or treatment of I/R-related ocular diseases.

LncRNA Ttc3-209 Promotes the Apoptosis of Retinal Ganglion Cells in Retinal Ischemia Reperfusion Injury by Targeting the miR-484/Wnt8a Axis.

PurposeApoptosis of the retinal ganglion cells (RGCs) can cause irreversible damage to visual function after retinal ischemia reperfusion injury (RIR). Using a lncRNA chip assay, we selected lncRNA Ttc-209 and characterized its role in RGCs during ischemia reperfusion (I/R)–induced apoptosis.MethodsWe created an ischemic model of RGCs by applying Hank's balanced salt solution containing 10 µM antimycin A and 2 µM calcium ionophore for 2 hours. RIR was induced in mice by elevating the intraocular pressure to 120 mm Hg for 1 hour by cannulation of the cornea; this was followed by reperfusion. Real-time quantitative PCR was used to detect the expression levels of long noncoding RNA (lncRNA), microRNA (miRNA), and target gene mRNA. Western blotting, flow cytometry, immunofluorescent staining, and TUNEL assays were performed to detect cell apoptosis. Dual-luciferase reporter assays and FISH were used to identify endogenous competitive RNA (ceRNA) mechanisms that link lncRNAs, miRNAs, and target genes. We also used scotopic electroretinography examinations to evaluate visual function in treated mice.ResultslncRNA Ttc3-209 was significantly upregulated after I/R injury and played a proapoptotic role in RGCs during I/R-induced apoptosis. Mechanistically, lncRNA Ttc3-209 is a ceRNA that competitively binds to miR-484 and upregulates the translation of its target (Wnt8a mRNA), thus promoting apoptosis in RGCs.ConclusionsReducing the expression of lncRNA Ttc3-209 had a protective effect against apoptosis in RGCs. This may provide a new therapeutic option for the prevention of RGC apoptosis in response to RIR injury.

Nutrigenetic reprogramming of oxidative stress.

Retinal disorders such as retinitis pigmentosa, age-related retinal degeneration, oxygen-induced retinopathy, and ischemia-reperfusion injury cause debilitating and irreversible vision loss. While the exact mechanisms underlying these conditions remain unclear, there has been a growing body of evidence demonstrating the pathological contributions of oxidative stress across different cell types within the eye. Nuclear factor erythroid-2-related factor (Nrf2), a transcriptional activator of antioxidative genes, and its regulator Kelch-like ECH-associated protein 1 (Keap1) have emerged as promising therapeutic targets. The purpose of this review is to understand the protective role of the Nrf2-Keap1 pathway in different retinal tissues and shed light on the complex mechanisms underlying these processes. In the photoreceptors, we highlight that Nrf2 preserves their survival and function by maintaining oxidation homeostasis. In the retinal pigment epithelium, Nrf2 similarly plays a critical role in oxidative stabilization but also maintains mitochondrial motility and autophagy-related lipid metabolic processes. In endothelial cells, Nrf2 seems to promote proper vascularization and revascularization through concurrent activation of antioxidative and angiogenic factors as well as inhibition of inflammatory cytokines. Finally, Nrf2 protects retinal ganglion cells against apoptotic cell death. Importantly, we show that Nrf2-mediated protection of the various retinal tissues corresponds to a preservation of functional vision. Altogether, this review underscores the potential of the Nrf2-Keap1 pathway as a powerful tool against retinal degeneration. Key insights into this elegant oxidative defense mechanism may ultimately pave the path toward a universal therapy for various inherited and environmental retinal disorders.

Houttuynia cordata Thunb rescues retinal ganglion cells through inhibiting microglia activation in a rat model of retinal ischemia-reperfusion.

To determine whether Houttuynia cordata Thunb (HCT) can increase the survival of the retinal ganglion cells (RGCs) and inhibit microglia activation following retinal ischemia-reperfusion (RIR) injury. Rat model of RIR was induced by transient elevation of the intraocular pressure (IOP). HCT was orally administered for 2d before the performance of retinal RIR model and once a day for the next 14d. After 14d of RIR injury, the rats were sacrificed for further analysis. Survival RGCs were stained with haematoxylin and eosin (H&E). Apoptosis of RGCs was detected by TUNEL staining. Retinal function was examined by flash-electroretinography (F-ERG). Retinal microglia were labeled using Iba-1, one specific marker for microglia. The mRNA expression levels of inducible nitric oxide synthase (iNOS), tumor necrosis factor alpha (TNF-α), and interleukin 1 beta (IL-1β) were assessed by quantitative real time reverse transcription polymerase chain reaction (qRT-PCR). Systemic HCT treatment significantly reduced RGCs death by H&E staining and exhibited an anti-apoptotic effect as assessed by TUNEL staining at day 14 after RIR injury. HCT greatly improved the retinal function as examined by F-ERG. The number of activated microglia significantly increased after RIR injury, which was significantly attenuated by HCT treatment. Besides, RIR injury induced a strong upregulation of pro-inflammatory genes TNF-α, iNOS and IL-1β mRNAs at day 14 post injury, which was suppressed by HCT. Neuroprotective effects of HCT encourage the survival of RGCs through inhibiting microglia activation due to RIR injury. Together these results support the use of HCT as promising therapy for the ischemic events of the retina diseases.

Involvement of the NLRC4 inflammasome in promoting retinal ganglion cell death in an acute glaucoma mouse model

PurposeTo explore the role of nucleotide-binding oligomerization domain-like receptors (NLRs) family caspase-activation and the recruitment domain containing 4 (NLRC4) inflammasome in retinal ganglion cell (RGC) injury induced by an acute glaucoma mouse model. MethodA mouse model of acute ocular hypertension, which can lead to retinal ischemia-reperfusion (I/R) injury, was established. The expression level of NLRC4 was detected by polymerase chain reaction and western blotting. Localized expression of NLRC4 was detected by examining immunofluorescence in eyeball sections. Intravitreal adeno-associated virus 2(AAV2) administration was used to knockdown retinal Nlrc4. Fluoro-Gold labeled RGCs and TdT-mediated dUTP nick end labeling were used to evaluate the survival and apoptosis of RGCs. Tlr4−/− mice were utilized to explore whether NLRC4 inflammasome is influenced by Toll-like receptor4 (TLR4). ResultsNLRC4, expressed in RGCs and microglial cells, was actively involved in mouse retinal I/R injury. Knockdown of Nlrc4 using an AAV2 vector caused an obvious reduction in the generation of IL-1β led by the rapidly elevated intraocular pressure, and thereby improved the RGC survival. In addition, activation of the NLRC4 inflammasome could influence the phosphorylation of p38 and Jun N-terminal kinase, which was largely dependent on TLR4 signaling. ConclusionOur study demonstrated the role of NLRC4 inflammasome in promoting RGC damage in mouse retinal I/R injury. Inhibition of NLRC4 might be leveraged as a potential therapeutic target in glaucomatous retinopathy.

Therapeutic Targeting of Retinal Immune Microenvironment With CSF-1 Receptor Antibody Promotes Visual Function Recovery After Ischemic Optic Neuropathy.

Retinal ischemia/reperfusion injury (RI) is a common cause of irreversible visual impairment and blindness in elderly and critical unmet medical need. While no effective treatment is available for RI, microglial activation and local immune responses in the retina are thought to play important roles in the pathophysiology of neurodegeneration. While survival and activation of microglia depend critically on colony-stimulating factor receptor (CSF-1R) signaling, it remains unclear if targeting the retinal immune microenvironments by CSF-1RAb after RI is sufficient to rescue vision and present a potentially effective therapy. Here we used rodent models of RI and showed that retinal ischemia induced by acute elevation of intraocular pressure triggered an early activation of microglia and macrophages in the retina within 12 h. This was followed by lymphocyte infiltration and increased production of pro-inflammatory cytokines. Intravitreal injection of CSF-1R neutralizing antibody (CSF-1RAb) after RI significantly blocked microglial activation and the subsequent T cell recruitment. This also led to improved retinal ganglion cell survival and function measured by cell quantification and electroretinogram positive scotopic threshold responses, as well as increased visual acuity and contrast sensitivity as assessed by optomotor reflex-based assays, when compared to the isotype-treated control group. Moreover, the administration of CSF-1RAb efficiently attenuated inflammatory responses and activation of human microglia in culture, suggesting a therapeutic target with human relevance. These results, together with the existing clinical safety profiles, support that CSF-1RAb may present a promising therapeutic avenue for RI, a currently untreatable condition, by targeting microglia and the immune microenvironment in the retina to facilitate neural survival and visual function recovery.

Application of Hydrogen in Ophthalmology.

This report reviews studies on the use of H2 in the ophthalmological field. In retinal diseases, particularly in a retinal ischemia-reperfusion injury, effects of H2 are remarkable in reducing retinal tissue damage. H2 treatment of corneal damage caused by alkali or UVB suppressed scar formation. The most unique application of H2 in the ophthalmological field appears to be its use in phacoemulsification cataract surgery. Ultrasound oscillation produces ·OH through the cavitation phenomenon in the anterior chamber of the eye, which induces oxidative insults in the corneal endothelium. Phacoemulsification using H2 dissolved in the irrigation solution significantly suppressed the corneal endothelial damage. The effect of H2 was direct and clear, as H2 instantly scavenges ·OH produced by ultrasound oscillation in the anterior chamber, thereby suppressing oxidative insults during the phacoemulsification procedure.

Overexpression of S100A4 protects retinal ganglion cells against retinal ischemia-reperfusion injury in mice

BackgroundGlaucoma is characterized by the neurodegeneration of retinal ganglion cells (RGCs) and the optic nerve. Numerous studies have reported that S100A4 participates in the metastasis of tumor cells and nerve protection. This study was intended to explore the role of S100A4 on RGCs under retinal ischemia-reperfusion (I/R) injury in mice. MethodsC57BL/6J mice were used to induce retinal I/R injury. The intravitreal administration of rAAV-EF1α-s100a4-EGFP-WPRE (rAAV-S100A4) or rAAV-EF1α-EGFP-WPRE-Pa was performed 4 weeks before I/R injury. Expression of S100A4 was detected by quantitative real-time PCR, immunofluorescence staining of retinal sections and western blot. Surviving RGCs were quantified using immunofluorescence staining. Staining of TUNEL was utilized to evaluate the apoptosis of retinal cells. Electroretinogram (ERG) was used to analyze retinal function. Expression of Akt, phospho-Akt, Bcl-2, and Bax were determined using western blotting to investigate the potential mechanisms of S100A4. ResultsRetinal S100A4 level had no statistical difference 7 days after I/R injury. The rAAV-S100A4 was clearly demonstrated by the green fluorescence protein in many layers of the retina after intravitreal injection and up-regulated the expression of S100A4. I/R injury resulted in an increase of the apoptosis of retinal cells and the reduction of surviving RGCs, however, overexpressed S100A4 inhibited the apoptosis of cells and a decrease of RGCs. ERG analysis showed a drop on amplitude of a-wave and b-wave was impeded to some extent by overexpressing of S100A4. Up-regulation of S100A4 raised the expression of phospho-Akt and reduced Bax expression. Nevertheless, there were no significant changes in the levels of Bcl-2 and total Akt. ConclusionOur results indicate the neuroprotective effects of overexpressed S100A4 on RGCs by activating the Akt pathway and then inhibiting the apoptosis of cells after I/R injury. The use of S100A4 protein may be a novel therapeutic strategy for glaucoma.

Inactivation of Endothelial ADAM17 Reduces Retinal Ischemia-Reperfusion Induced Neuronal and Vascular Damage

Retinal ischemia contributes to visual impairment in ischemic retinopathies. A disintegrin and metalloproteinase ADAM17 is implicated in multiple vascular pathologies through its ability to regulate inflammatory signaling via ectodomain shedding. We investigated the role of endothelial ADAM17 in neuronal and vascular degeneration associated with retinal ischemia reperfusion (IR) injury using mice with conditional inactivation of ADAM17 in vascular endothelium. ADAM17Cre-flox and control ADAM17flox mice were subjected to 40 min of pressure-induced retinal ischemia, with the contralateral eye serving as control. Albumin extravasation and retinal leukostasis were evaluated 48 h after reperfusion. Retinal morphometric analysis was conducted 7 days after reperfusion. Degenerate capillaries were assessed by elastase digest and visual function was evaluated by optokinetic test 14 and 7 days following ischemia, respectively. Lack of ADAM17 decreased vascular leakage and reduced retinal thinning and ganglion cell loss in ADAM17Cre-flox mice. Further, ADAM17Cre-flox mice exhibited a remarkable reduction in capillary degeneration following IR. Decrease in neurovascular degeneration in ADAM17Cre-flox mice correlated with decreased activation of caspase-3 and was associated with reduction in oxidative stress and retinal leukostasis. In addition, knockdown of ADAM17 resulted in decreased cleavage of p75NTR, the process known to be associated with retinal cell apoptosis. A decline in visual acuity evidenced by decrease in spatial frequency threshold observed in ADAM17flox mice was partially restored in ADAM17-endothelial deficient mice. The obtained results provide evidence that endothelial ADAM17 is an important contributor to IR-induced neurovascular damage in the retina and suggest that interventions directed at regulating ADAM17 activity can be beneficial for alleviating the consequences of retinal ischemia.