Acute Ocular Hypertension Research Articles

A Minimally Invasive Experimental Model of Acute Ocular Hypertension with Acute Angle Closure Characteristics.

PurposeTo describe a minimally invasive experimental model of acute ocular hypertension (OHT) with characteristics of acute angle closure (AAC).MethodsAdult C57/Bl6 mice (n = 31) were subjected to OHT in one eye using a modified circumlimbal suture technique that elevated intraocular pressure (IOP) for 30 minutes. Contralateral un-operated eyes served as controls. IOP, anterior segment optical coherence tomography, and fundus fluorescein angiography (FFA) were performed. The positive scotopic threshold response (pSTR) and a-wave and b-wave amplitudes were also evaluated. Retinal tissues were immunostained for the retinal ganglion cell (RGC) marker RBPMS and the glial marker GFAP.ResultsOHT eyes developed shallower anterior chambers and dilated pupils. FFA showed focal leakage in 32.2% of OHT eyes, but in none of the control eyes. pSTR was significantly reduced at week 1 in OHT eyes compared to control eyes (57.3 ± 7.2 µV vs. 106.9 ± 24.8 µV; P < 0.05), but a- and b-waves were unaffected. GFAP was upregulated in OHT eyes but not in control eyes or eyes that had been sutured without OHT. RGC density was reduced in OHT eyes after 4 weeks (3857 ± 143.8) vs. control eyes (4469 ± 176.0) (P < 0.05).ConclusionsOur minimally invasive model resulted in acute OHT with characteristics of AAC in the absence of non-OHT-related neuroinflammatory changes arising from ocular injury alone.Translational RelevanceThis model provides a valuable approach to studying specific characteristics of a severe blinding disease in an experimental setting. Focal areas of ischemia were demonstrated, consistent with clinical studies of acute angle closure patients elsewhere, which may indicate the need for further research into how this could affect visual outcome in these patients.

ScAAV2-Mediated C3 Transferase Gene Therapy in a Rat Model with Retinal Ischemia/Reperfusion Injuries

Glaucoma is characterized by retinal ganglion cell (RGC) death and axonal loss. Therefore, neuroprotection is important in treating glaucoma. In this study, we explored whether exoenzyme C3 transferase (C3)-based gene therapy could protect retinas in an ischemia/reperfusion (I/R) injury rat model. Self-complementary adeno-associated virus 2 (scAAV2) vectors encoding either C3 protein (scAAV2-C3) or enhanced green fluorescence protein (scAAV2-EGFP) were intravitreally delivered into both eyes of rats, and I/R models (acute ocular hypertension) were made in one eye of each rat at day 7 after the injection. The rats were divided into six groups: scAAV2-C3, scAAV2-C3 with I/R, scAAV2-EGFP, scAAV2-EGFP with I/R, blank control, and blank control with I/R. TUNEL (terminal deoxynucleotidyltransferase-mediated deoxyuridine triphosphate nick end labeling), immunohistochemistry of cleaved caspase-3, NeuN and Brn-3a, and H&E staining were used to detect apoptotic cells and other changes in the retina. The results showed that scAAV2-C3 significantly reduced the number of apoptotic RGCs and decreased cell loss in the ganglion cell layer after I/R injury, and the I/R-injured retinas treated with scAAV2-C3 were the thickest in all I/R groups. These results suggest that scAAV2-mediated C3 gene therapy is able to protect the rat retina from I/R injury and has potential in the treatment of glaucoma in the future.

Protective effects of carnosic acid on retinal ganglion cells in acute ocular hypertension rats.

To observe the protective effects of carnosic acid on rat retinal ganglion cells (RGCs) among acute ocular hypertension rats. Sixty male SPF (specific-pathogen-free) SD rats (10weeks) were randomly assigned to untreated group, carnosic-acid-treated group and hypertensive group with 20 rats for each. The acute ocular hypertension animal model was induced by the perfusion of normal saline solution into anterior chamber of eyes to elevate the intraocular pressure (IOP) to 110mmHg for 60min in the rats of the carnosic-acid-treated group and hypertensive group. Then, the carnosic acid dissolving in dimethyl sulfoxide (DMSO) was intraperitoneally injected for consecutive 7days in the carnosic-acid-treated group, and only DMSO was used in the same way in the hypertensive group. The rats were killed 2weeks after experiment, and retinal sections were prepared for histopathological and apoptotic retinal ganglion cells (RGCs) examination by hemotoxylin and eosin staining and TUNEL staining. Use immunofluorescence employed to examine the survival of RGCs. This study protocol was approved by the Ethic Committee for Experimental Animal of Three Gorges University. The retinal morphology and structure were clear in the untreated group. The edema of retinal tissue, loosely arranged RGCs and swollen nucleus were seen in the hypertensive group. In the carnosic-acid-treated group, the retinal morphology and structure were regular. The retinal nerve fiber layer (RNFL) thickness was (32.96 ± 1.63), (58.96 ± 1.57) and (50.11 ± 2.37) μm, and the apoptotic cell number was (6.92 ± 2.96), (29.85 ± 6.40) and (14.69 ± 2.98)/field, and the survived cell number was (2363.17 ± 148.45), (1308.67 ± 106.02) and (1614.17 ± 96.39)/0.235mm2 in the untreated group, hypertensive group and carnosic-acid-treated group, respectively, showing significant differences among groups (F = 339.284, 81.583, 122.68, all at P < 0.01). Compared with the untreated group, the RNFL thickness was thickened, the number of apoptotic RGCs was much more, and the number of survived RGCs was decreased in the hypertensive group. In the carnosic-acid-treated group, the RNFL thickness was thinner, the number of apoptotic RGCs was reduced, and the number of survived RGCs was increased in comparison with the untreated group (all at P < 0.01). Carnosic acid plays a protective effect on RGCs by inhibiting the cell apoptosis in acute ocular hypertension rats.

Protective Effects Oncorneal Endothelium During Intracameral Irrigation Using N-(2)-l-alanyl-l-Glutamine.

Corneal endothelial disease is a global sight-threatening disease, and corneal transplantation using donor corneas remains the sole therapeutic option. A previous work demonstrated that N (2)-alanyl-glutamine (Ala-Gln) protected against apoptosis and cellular stress, and maintained intestinal tissue integrity. In this pursuit, the present study aimed to examine the effect of Ala-Gln in the protection of the corneal endothelium and expand its range of potential clinical applications. Mice in the control group were intracamerally irrigated with Ringers lactate injection, whereas those in the experimental group were irrigated with Ringers lactate injection containing Ala-Gln. The mean intraocular pressure increased to 44 ± 3.5 mm Hg during intracameral irrigation (normal range 10.2 ± 0.4 mmHg). In vivo confocal microscopy results showed that the addition of Ala-Gln protected the morphology, structure, and density of the corneal endothelial cells. Optical Coherence Tomography (OCT) measurements showed that corneal thickness was not significantly different between the two groups, because of the immediate corneal edema after irrigation, but the addition of Ala-Gln obviously promoted the recovery of the corneal edema. Scanning electron microscopy indicated that the corneal endothelial cells were severely ruptured and exfoliated in the Ringer’s group accompanied with cellular edema, when compared with the Ala-Gln group. The intracameral irrigation using Ala-Gln protected the structure and expression of cytoskeleton and Na-K-ATPase, which exhibited a regular distribution and significantly increased expression in comparison to Ringer’s group. Furthermore, Ala-Gln maintained the mitochondrial morphology and increased the activity of mitochondria. Moreover, transmission electron microscopy showed that intracameral irrigation of Ala-Gln reversed the ultrastructural changes induced by the acute ocular hypertension in mice. Our study demonstrates that the intracameral irrigation of Ala-Gln effectively maintained the corneal endothelial pump function and barrier function by protecting the mitochondrial function and preventing the rearrangement of cytoskeleton in acute ocular hypertension in mice.

Expression and role of autophagy related protein p62 and LC3 in the retina in a rat model of acute ocular hypertension.

To investigate the expression and possible role of the autophagy related protein p62 and LC3 in the retina based on a rat model of acute ocular hypertension. Fifty rats were randomized into five groups: control group A, B, C, and D. Groups A to D all received normal saline perfusion into the anterior chamber with pressure of 80 mm Hg for one hour, and retina tissue was obtained at 6, 12, 24 and 48h after perfusion respectively, to investigate the activation of autophagy following ischemia-reperfusion. The distribution and semi-quantification of autophagy related protein p62 and LC3 in the retina were detected using immunohistochemistry technique. The expression level of these two proteins was evaluated using Western blot. The number of retinal ganglion cells (RGCs) decreased with increasing reperfusion time, and significant reduction in the retinal thickness was observed 48h after perfusion. In normal adult rats, LC3 protein was mainly expressed in the ganglion cell layer (GCL), and p62 protein was expressed in the nerve fiber layer (NFL), GCL, inner plexiform layer (IPL), inner nuclear layer (INL) and outer plexiform layer (OPL). In comparison to the control group, the expression level of LC3- II was higher in all the experimental groups (P<0.05), with the peak expression at 12h after reperfusion. Additionally, the expression level of p62 was higher in all the experimental groups than the control (P<0.05, except for group A), with the peak level occurred 24h after reperfusion. Both p62 and LC3 show low level and uneven expression in the retina of normal adult rats. Acute ocular hypertension can lead to upregulation of LC3- II and p62 expression in the retina. Autophagy flux is damaged 12h after reperfusion, potentially resulting in further loss of RGCs.

Lycium barbarum polysaccharides related RAGE and Aβ levels in the retina of mice with acute ocular hypertension and promote maintenance of blood retinal barrier.

Our previous study verified the protective effects of Lycium barbarum polysaccharides (LBP) on retinal neurons and blood vessels in acute ocular hypertension (AOH) mice. To investigate the effect of LBP on the reactivity of retinal glial cells, an AOH mouse model was established in one eye by maintaining ocular hypertension of 90 mmHg for 60 minutes. Either LBP solution (1 mg/kg) or phosphate-buffered saline was administrated to the mice by gavage daily, starting 7 days before the AOH insult and continuing until the mice were sacrificed for specimen collection on day 4 post-insult. After AOH insult, increased numbers of astrocytes and microglia were observed, together with decreased expression of the following glial cell biomarkers in the retinal ganglion cells of AOH mice: glial fibrillary acidic protein, glutamine synthetase, aquaporin-4, S-100 proteins, ionized calcium-binding adaptor molecule 1, amyloid precursor protein and receptor of advanced glycosylation end-products. After intervention with LBP, the above changes were significantly reduced. Remarkably, morphological remodeling of blood vessel-associated retinal astrocytes, marked by glial fibrillary acidic protein, was also observed. These results, taken together, suggest that LBP regulated the production of amyloid-β and expression of receptor of advanced glycosylation end-products, as well as mediating the activity of retinal glial cells, which may lead to the promotion of better maintenance of the blood-retinal barrier and improved neuronal survival in AOH insult. This study was approved by the Committee for the Use of Live Animals in Teaching and Research (approval No. CULTRA-#1664-08).

Functional and morphological alterations in a glaucoma model of acute ocular hypertension.

To study short and long-term effects of acute ocular hypertension (AOHT) on inner and outer retinal layers, in adult Sprague-Dawley rats AOHT (87mmHg) was induced for 90min and the retinas were examined longitudinally in vivo with electroretinogram (ERG) recordings and optical coherent tomography (OCT) from 1 to 90 days (d). Ex vivo, the retinas were analyzed for rod (RBC) and cone (CBC) bipolar cells, with antibodies against protein kinase Cα and recoverin, respectively in cross sections, and for cones, horizontal (HZ) and ganglion (RGC) cells with antibodies against arrestin, calbindin and Brn3a, respectively in wholemounts. The inner retina thinned progressively up to 7d with no further changes, while the external retina had a normal thickness until 30d, with a 20% thinning between 30 and 90d. Functionally, the a-wave showed an initial reduction by 24h and a further reduction from 30 to 90d. All other main ERG waves were significantly reduced by 1d without significant recovery by 90d. Radial sections showed a normal population of RBCs but their terminals were reduced. The CBCs showed a progressive decrease with a loss of 56% by 30d. In wholemount retinas, RGCs diminished to 40% by 3d and to 16% by 30d without further loss. Cones diminished to 58% and 35% by 3 and 7d, respectively and further decreased between 30 and 90d. HZs showed normal values throughout the study. In conclusion, AOHT affects both the inner and outer retina, with a more pronounced degeneration of the cone than the rod pathway.

Rod bipolar cells dysfunction occurs before ganglion cells loss in excitotoxin-damaged mouse retina

Progressive degeneration of retinal ganglion cells (RGCs) will cause a blinding disease. Most of the study is focusing on the RGCs itself. In this study, we demonstrate a decline of the presynaptic rod bipolar cells (RBCs) response precedes RGCs loss and a decrease of protein kinase Cα (PKCα) protein expression in RBCs dendrites, using whole-cell voltage-clamp, electroretinography (ERG) measurements, immunostaining and co-immunoprecipitation. We present evidence showing that N-methyl D-aspartate receptor subtype 2B (NR2B)/protein interacting with C kinase 1 (PICK1)-dependent degradation of PKCα protein in RBCs contributes to RBCs functional loss. Mechanistically, NR2B forms a complex with PKCα and PICK1 to promote the degradation of PKCα in a phosphorylation- and proteasome-dependent manner. Similar deficits in PKCα expression and response sensitivity were observed in acute ocular hypertension and optic never crush models. In conclusion, we find that three separate experimental models of neurodegeneration, often used to specifically target RGCs, disrupt RBCs function prior to the loss of RGCs. Our findings provide useful information for developing new diagnostic tools and treatments for retinal ganglion cells degeneration disease.

XEN in extreme cases

AbstractPurposeThe Xen implant allows to perform a filtering procedure using a collagen tube of 6 mm in length and 45 microns of lumen usually placed in the upper nasal quadrant. Its main indication is primary open angle glaucoma, however, its versatility has allowed it to be used in other types of glaucoma cases. The aim of this communication is to present some cases in which the Xen implant has been used to treat complex situations.Methods &amp; ResultsCase 1 ‐ A 31‐year‐old woman with congenital glaucoma, a history of goniotomy and a drainage device carrier of double‐plate glaucoma who presented intraocular pressure (IOP) of 17 mmHg with maximum topical and oral treatment and severe toxic ocular disease. As the upper nasal quadrant was occupied, a XEN device was implanted in the inferior nasal quadrant. During the postoperative period, displacement of the implant towards the anterior chamber was evident, but no endothelial cell loss was observed and control of the IOP was maintained at 1 year of follow‐up with a significant reduction in medication and improvement of the ocular surface. Case 2 ‐ A 34‐year‐old man presented with contusive trauma in his left eye complicated by 2‐mm hyphema and (IOP &gt; 40 mmHg ) that could not be controlled with maximum medication. After performing a drainage of the hyphema a XEN implant was placed in the upper nasal quadrant. Since then, IOP was controlled and the visual acuity completely reestablished. Case 3 ‐ A 58‐year‐old woman with severe scleromalacia affecting all four quadrants. She had a history of trabeculectomy in the upper quadrant and a superior nasal sector of healthy conjunctiva that was normocolored and mobilizable was observed. After placing the Xen in the upper nasal sector, the patient has remained with an IOP of 16 mmHg with a beta‐blocker medication.Conclusions1. The Xen implant can be useful in the management of glaucoma in some challenging situations. 2. The inferior nasal location of the Xen is raised exceptionally. It can make it more susceptible to displacement, although this does not necessarily affect its functioning. 3. The Xen device can be useful in acute uncontrolable ocular hypertension (ex trauma) in the context of an open angle.

The neuroprotective effects of novel estrogen receptor GPER1 in mouse retinal ganglion cell degeneration

PurposeTo investigate the potential protective effect of novel G protein coupled estrogen receptor (GPER1) against the neurotoxicity induced by NMDA in the mouse retina. MethodsWe induce retinal ganglion cells (RGCs) toxic injury through intravitreal injection of NMDA or acute ocular hypertension (AOH) induced by anterior chamber infusion with saline. Endogenous ligand 17-β-estradiol (E2), GPER1 agonist (G-1), and E2 with GPER1 antagonist (G-15) or classic estrogen receptor α and β (ERα and ERβ) antagonist tamoxifen (TAM) were subcutaneous administered before NMDA to identify the possible involved receptors. Immunofluorescence staining was performed to explore the survival of RGCs and Müller cell gliosis. TUNEL staining was used to evaluate the RGC apoptosis. The involved molecular pathway was detected via antibody array expression profiling. ResultsActivation of estrogen receptor by E2 or G-1 could significantly rescue the RGCs injury in NMDA administration. The protective effect was carried exclusively by GPER1 activation. E2 application can still mimicked the protective function when estrogen receptor α and β (ERα and ERβ) blocked by tamoxifen (TAM), while the effects was blocked by GPER1 antagonist G-15. Moreover, the TUNEL positive RGCs and GFAP expression level were both attenuated in G-1 application and the effects could be reversed by G-15. In addition, application of the PI3K/Akt antagonist LY294002 counteracted the effect of G-1. And a number of apoptosis regulatory factors decreased dramatically in the G-1 group, including Bad, Caspase 3, Caspase 7, Smad2, P-53 and TAK1. Also, similar protective effect of G-1 was spotted in acute ocular hypertension (AOH) model. ConclusionEstrogen played a protective role via a novel estrogen receptor, GPER1, instead of classical receptors ERα or ERβ. Activation of GPER1 attenuated RGCs apoptosis and Müller cells gliosis, indicating GPER1 as a potential treatment target in RGCs degeneration diseases.

Lycium Barbarum Polysaccharides Rescue Neurodegeneration in an Acute Ocular Hypertension Rat Model Under Pre- and Posttreatment Conditions.

To investigate the posttreatment neuronal rescue effects of Lycium barbarum polysaccharides (LBP) in an acute ocular hypertensive (AOH) model. Intraocular pressure (IOP) was elevated manometrically to 80 mm Hg (AOH) or 15 mm Hg (sham) for 120 minutes in adult Sprague-Dawley rats. Five experimental groups were considered: Three AOH groups were pretreated with PBS (vehicle) (n = 9), LBP 1 mg/kg (n = 8), or 10 mg/kg (n = 13), and one AOH group was posttreated with LBP 10 mg/kg (n = 8), once daily. The sham cannulation group (n = 5) received no treatment. Pretreatments commenced 7 days before and posttreatment 6 hours after AOH, and continued up through postcannulation day 28. All the animals underwent optical coherence tomography and electroretinogram measurements at baseline and postcannulation days 10 and 28. The ganglion cell layer (GCL) densities were quantified at day 28. Both inner retinal layer thickness (IRLT) and positive scotopic threshold response (pSTR) underwent significant reduction (≥50% of thickness and amplitude) in the vehicle group (P < 0.05). Pretreatment with LBP 1 and 10 mg/kg retained 77 ± 11% and 89 ± 8% of baseline IRLT, respectively, and preserved pSTR functions. The posttreatment group showed a significant reduction in IRLT (-35 ± 8%, P < 0.001) and pSTR (∼48% of baseline, P < 0.001) on day 10. By day 28, there was an improvement in functional pSTR (∼72% of baseline, P > 0.05) with no significant further thinning (-40 ± 8%, P = 0.15) relative to day 10. GCL density was reduced in vehicle control (P = 0.0001), but did not differ between sham and pre- and posttreated AOH groups. The rescue effect of LBP posttreatment was observed later, which arrested the secondary degeneration and improved the retinal function.

Effect of BDNF-TrKB pathway on apoptosis of retinal ganglion cells in glaucomatous animal model.

The aim of this paper is to investigate the effect of BDNF-TrKB pathway on AOH by accessing its regulatory role in retinal ganglion cell apoptosis. Acute ocular hypertension (AOH) model in rats was established by anterior chamber perfusion to increase intraocular tension. Rats were randomly divided into AOH group, control group and k252a group, with ten rats in each group. Rats were sacrificed 72 h after animal procedures and eyeballs were harvested. HE staining was used to observe retinal structural changes at different time points. Immunohistochemical staining was used to observe the BDNF-positive cells in retinal tissues. TUNEL staining was conducted to measure apoptosis of retinal ganglion cells. Reverse Transcriptase-Polymerase Chain Reaction (RT-PCR) and Western blot were performed to detect mRNA and protein levels of BDNF, TrKB, PI3K and ERK1 in retinal tissues, respectively. HE and TUNEL staining showed significant pathological changes and abundant apoptotic cells in rat retina of AOH group and k252a group compared with those of the control group (p<0.05). The number of survived retinal ganglion cells in the AOH group was lower than that of the control group (p<0.05). K252a group had the lowest number of survived retinal ganglion cells. Immunohistochemical results showed that BDNF was rarely expressed in rat retinal tissues of the control group, which was remarkably pronounced in the AOH group and k252a group. The number of BDNF-positive cells in the k252a group was higher than that in the AOH group (p<0.05). RT-PCR and Western blot indicated that mRNA and protein levels of relative genes in BDNF-TrKB and PI3K/ERK1 pathways were upregulated in AOH group (p<0.05), but were significantly downregulated in k252a group (p<0.05). BNDF-TrKB pathway exerts a protective effect on retina against acute ocular hypertension by reducing retinal cell apoptosis.

Vagal Nerve Stimulation Attenuates Ischemia-Reperfusion Induced Retina Dysfunction in Acute Ocular Hypertension

Purpose: The present study aimed to investigate whether cervical vagal nerve stimulation (VNS) could prevent retinal ganglion cell (RGC) loss and retinal dysfunction after ischemia/reperfusion (I/R) injury.Methods: First, rats were randomly divided into sham group (n = 4) and VNS group (n = 12). Activation of the nodose ganglia (NOG), nucleus of the solitary tract (NTS), superior salivatory nucleus (SSN), and pterygopalatine ganglion (PPG) neural circuit were evaluated by c-fos expression at 0 h after sham VNS and at 0 h (n = 4), 6 h (n = 4), 72 h (n = 4) after VNS. Secondly, rats were randomly assigned to I/R group (pressure-induced retinal ischemia for 1 h and reperfusion for 1 h in the right eye, n = 16) and I/R+VNS group (right cervical VNS for 2 h during the I/R period, n = 16). The left eye of each rat served as a control. Electroretinogram (ERG), RGC numbers, tumor necrosis factor-α (TNF-α) and vasoactive intestinal polypeptide (VIP) levels in retina were determined. Additionally, the level of VIP in PPG was evaluated.Results: In the first part of the study, compared with the sham group, the VNS group exhibited significantly increased expression of c-fos in NOG, NTS, SSN, and PPG tissues at 0, 6, and 72 h. In the second part of the study, compared with left eyes, retinal function in right eyes (as assessed by the a-wave, b-wave and the oscillatory potential amplitudes of ERG and RGC data) was significantly decreased by I/R. The decreased retinal function was attenuated by VNS. In addition, I/R induced an increase in inflammation, which was reflected by elevated TNF-α expression in the retina. VNS significantly attenuated the increase in I/R-induced inflammation. Moreover, VIP expression in the retina and PPG, which may contribute to the inhibition of the inflammatory response, was significantly increased after VNS.Conclusion: VNS could protect against retinal I/R injury by downregulating TNF-α. Upregulation of VIP expression due to activation of the NOG-NTS-SSN-PPG neural circuit may underlie to the protective effects of VNS.

Methyl 3,4-dihydroxybenzoate protects retina in a mouse model of acute ocular hypertension through multiple pathways

Methyl 3,4 dihydroxybenzoate (MDHB) is a small molecule that shows neuroprotective effects in vitro and in a photoreceptor-degenerative mouse model. Here we investigated whether MDHB protects retina in a mouse model of acute ocular hypertension (AOH) and explores the underlying mechanisms. AOH was induced in mice by increasing intraocular pressure to approximately 90 mmHg for 60 min, then MDHB or vehicle was intraperitoneally injected daily up to 7 days. Immunostaining and multi-electrode array recordings were performed to examine the structure and function of retinas receiving the treatments. Western-blotting was applied to test the expression of several proteins related to oxidative stress and brain-derived neurotrophic factor (BDNF)-initiated signaling. Results showed that AOH injury reduced the number of Brn3a-stained retinal ganglion cells (RGCs) and ChAT-amacrine cells; thinned the inner retinal layers and induced apoptosis. Physiologically, AOH decreased the response of OFF and ON-OFF RGCs. All of these changes were reversed by MDHB-treatment. Mechanistically, MDHB appeared to work on three parallel pathways: (1) MDHB decreased the production of reactive oxygen species, the expression of nuclear factor erythroid 2-related factor 2 (Nrf2) and cytosol heme oxygenase 1 (HO-1); (2) It upregulated the expression of BDNF and its receptor tropomyosin-related kinase B (TrkB), and activated the downstream AKT pathways; (3) It inhibited reactive gliosis by reducing GFAP and Iba-1 expression. Thus our results suggest that MDHB protects retina against AOH injury by inhibiting oxidative stress, activating the BDNF/AKT signaling and inhibiting inflammatory pathways. Therefore, MDHB may serve as a promising candidate to treat retinal ischemia.

Neuronal apoptosis, axon damage and synapse loss occur synchronously in acute ocular hypertension

Retinal ganglion cells (RGCs) apoptosis and their axon degeneration are pivotal features in glaucoma. Previous studies suggest that the process of RGCs soma degeneration is distinct from axon degeneration and that both of them lead to vision loss but separately. However, since a normal visual function relies on the integrity of axon, synapse and soma in the retina, a comprehensive understanding of the changes of these neuron components in glaucoma is desired. Therefore, in an acute ocular hypertension (AOH) model in mice, we systematically evaluated retinal neuron soma, axon and synapse alteration at certain time points. We found that ocular hypertension led to a progressive apoptosis of retinal neural cells which proceeded from peripheral to central retina in the wholemount, meanwhile, started in the ganglion cell layer (GCL) and spread to the inner nuclear layer (INL) and then the outer nuclear layer (ONL) as time went on. The type of apoptotic cells was identified as RGCs in GCL, amacrine cells in INL and cone photoreceptor cells in ONL. Axon degeneration was observed at the same time as soma degenerated and also progressed from peripheral to central retina. More interestingly, accumulation of neurofilament in the soma caused by axon transport failure was detected synchronously. We also found that presynaptic and postsynaptic vesicle proteins were downregulated. Taken together, these data support a view that retinal neuronal apoptosis happens not only in RGCs, but also other neurons in laminar layers. Axon damage and synapse loss occur synchronously with soma loss in AOH. The combination of these three parameters might facilitate a systematic evaluation of the disease progression and treatment strategies in glaucoma.

Effect of the Notch signaling pathway on retinal ganglion cells and its neuroprotection in rats with acute ocular hypertension.

To explore the effect of the Notch signaling pathway on retinal ganglion cells (RGCs) and optic nerve in rats with acute ocular hypertension (OH). Totally 48 Sprague-Dawley (SD) rats were included, among which 36 rats were selected to establish acute OH models. OH rats received a single intravitreal injection of 2 µL phosphate buffered solution (PBS) and another group of OH rats received a single intravitreal injection of 10 µmol/L γ-secretase inhibitor (DAPT). Quantitative real-time polymerase chain reaction (qPCR) and Western blot assay were adopted to determine the mRNA level of Notch and the protein levels of Notch, Bcl-2, Bax, caspase-3, and growth-associated protein 43 (GAP-43). The RGC apoptosis conditions were assessed by TUNEL staining. The OH rats and PBS-injected rats had increased expression levels of Notch1, Bax, caspase-3, and GAP-43, decreased expression levels of Bcl-2, and increased RGC apoptosis, with severer macular edema and RGCs more loosely aligned, when compared with the normal rats. The DAPT-treated rats displayed increased expression levels of Notch1, Bax, caspase-3, and GAP-43, decreased expression levels of Bcl-2, and increased RGC apoptosis, in comparison with the OH rats and PBS-injected rats. RGCs were hardly observed and macular edema became severe in the DAPT-treated rat. The Notch signaling pathway may suppress the apoptosis of retinal ganglion cells and enhances the regeneration of the damaged optic nerves in rats with acute OH.

Gastrodin protects retinal ganglion cells through inhibiting microglial-mediated neuroinflammation in an acute ocular hypertension model.

To investigate the neuroprotective effect of gastrodin on retinal ganglion cells (RGCs) in an acute ocular hypertension (AOH) rat model and to identify its possible mechanism. AOH rat model was performed in a randomly selected eye by anterior chamber perfusion and either received an intraperitoneal injection with various concentrations of gastrodin or normal saline. After 2wk, the rats were sacrificed. FluoroGold was used to label survival RGCs. Immunostaining with anti-Iba1 in the retinal flat mounts to calculate the microglia density in the ganglion cell layer (GCL). Changes in microglial cytokines, tumour necrosis factor-alpha (TNF-α) and inducible NO synthase (iNOS) were examined with Western blot and reverse transcription-quantitative polymerase chain reaction. Expression levels of total and phosphorylated p38 mitogen activated protein kinase (MAPK) were determined by Western blot. Results showed that AOH induced significant loss of RGCs and severe microglia activation in the GCL. Besides, AOH increased the phosphorylation of p38 MAPK and promoted the release of microglial cytokines in the retinas. Intraperitoneal injection with dose-dependent gastrodin significantly reduced the loss of RGCs and inhibited retinal microglia activation, accompanied with the decreased expression levels of microglial cytokines and p38 MAPK phosphorylation. Gastrodin exerts a neuroprotective effect on RGCs in an acute glaucoma animal model via inhibiting microglia activation and microglial-mediated neuroinflammation. The finding demonstrates the potential application of gastrodin in the neuroprotective therapy of acute glaucoma and other retinal neurodegenerative diseases characterized by microglia activation and RGCs death.

Caveolin-1 Protects Retinal Ganglion Cells against Acute Ocular Hypertension Injury via Modulating Microglial Phenotypes and Distribution and Activating AKT pathway

Glaucoma, a group of eye diseases, causes gradual loss of retinal ganglion cells (RGCs) and ultimately results in irreversible blindness. Studies of the underlying mechanisms of glaucoma and clinical trial are far from satisfactory. Results from a genome-wide association study have suggested that the CAV1/CAV2 locus is associated with glaucoma, but this association and its potential underlying mechanisms need to be confirmed and further explored. Here, we studied the function of caveolin-1 (Cav1) in an acute ocular hypertension glaucoma model. Cav1 deficiency caused an aggregated lesion in the retina. In addition, treatment with cavtratin, a membrane permeable Cav1 scaffolding domain peptide, enhanced RGC survival. After cavtratin treatment, microglial numbers decreased significantly, and the majority of them migrated from the inner retinal layer to the outer retinal layers. Furthermore, cavtratin promoted a change in the microglia phenotype from the neurotoxic pro-inflammatory M1 to the neuroprotective anti-inflammatory M2. In a molecular mechanism experiment, we found that cavtratin activated the phosphorylation of both AKT and PTEN in cultured N9 cells. Our data highlights the neuroprotective effect of Cav1 on acute ocular hypertension and suggests that Cav1 may serve as a novel therapeutic target for the treatment of glaucoma. We further propose that cavtratin is a therapeutic candidate for glaucoma clinical trials.

Elevation of intraocular pressure in relation to retinal diseases

SummaryIntraocular pressure remains the only modifiable risk factor in glaucoma, a disease characterized by progressive loss or retinal ganglion cells (RGCs) and visual field deficits leading to blindness. Using two rat models of chronic (COH) or acute (AOH) ocular hypertension we study the response of the general population of RGCs responsible for image‐forming visual information and the intrinsically photosensitive population of RGCs responsible for nonimage‐forming visual information, identified with Brn3a and melanopsin antibodies, respectively. For AOH the anterior chamber of the left eye was cannulated and connected to saline elevated 1½ meters for 75 minutes. COH was induced by laser‐photocoagulation of limbar tissues. In both models prior to OH a single intravitreal injection of brain derived neurotrophic factor (5 μg BDNF in 1% albumin PBS) or vehicle was administered. Re retinas were examined at 12 or 15 days after COH or 3, 7, 14, or 45 days after AOH to identify surviving Brn3a+RGCs and m+RGCs. COH induced comparable loss of RGCs but only Brn3a+RGCs responded to BDNF. After AOH, Brn3a+RGCs died progressively while m+RGCs did not, and BDNF induced a permanent protection up to 45 days after AOH in both types of RGCs.

Acute ocular hypertension disrupts barrier integrity and pump function in rat corneal endothelial cells

Acute ocular hypertension (AOH) frequently compromises corneal endothelial cell (CEC) function in clinical practice. This type of stress induces corneal oedema and a decrease in the corneal endothelial cell density (ECD). The anterior chamber of the right eye of Sprague-Dawley rats was irrigated with Balanced Salt Solution (BSS) for two hours, and the left eye served as a control to determine the time-dependent effects of AOH on endothelial cell morphology and function. The average intraocular pressure (IOP) increased to 82.6 ± 2.3 mmHg (normal range: 10.2 ± 0.4 mmHg) during anterior irrigation. Very soon after initiating irrigation, corneal oedema became evident and the cornea exhibited a significant increase in permeability to FITC-dextran. The peripheral ECD was significantly reduced, and the morphology of CECs became irregular and multiform. The structures of the zonula occludens-1 (ZO-1) and F-actin were severely disrupted. In addtion, Na,K-ATPase exhibited a dispersed expression pattern. Two days after irrigation, obvious CEC proliferation was observed, the ECD recovered to a normal level, and F-actin was dispersed throughout the cytoplasm. Seven days later, the CEC structure and function were nearly normalized. Based on the results obtained using this model, an acute IOP crisis exerts transient deleterious effects on CEC structure and function in rats.