Glucocorticoid-induced Ocular Hypertension Research Articles

Understanding the Ocular Hypertension Model in Mice Induced by Dexamethasone-21-Acetate – Implications for Glaucoma Research

Purpose This study aimed to assess the effectiveness of monocular and bilateral injections of Dexamethasone-21-acetate (Dex-21-Ac) into the murine fornix twice a week as a glucocorticoid-induced ocular hypertension model and investigated potential systemic side effects. Methods Dex-21-Ac was administered twice weekly in three groups: bilateral injections, monocular injections, and a control group receiving the vehicle solution bilateral. After 21 days, enucleated eyes were examined using immunocytochemistry (ICC), and organ histology was performed. Results All groups receiving Dex-21-Ac injections had a significant increase in intraocular pressure (IOP). Monocular injections also resulted in a significant increase in IOP in the fellow eye. The Dex-21-Ac-treated groups showed a bilateral increase in IOP of approximately 8 mmHg, accompanied by elevated expression of alpha smooth muscle actin and fibronectin in the anterior chamber angle. There were no significant changes in weight progression. Hepatic steatosis was observed in all Dex-21-Ac-treated animals, and some suffered from residual neuromuscular blockade under fentanyl anesthesia. Conclusion Bilateral injections of Dex-21-Ac twice a week lead to a significant increase in daytime IOP and fibrotic changes in the trabecular meshwork. Unilateral application has a significant impact on the fellow eye. Local dexamethasone leads to notable systemic effects independent of changes in animal weight. Considering liver damage and associated influence on metabolization, hepatically eliminated injection anesthetics may lead to overdosing and are not recommended. They should be replaced by inhalation anesthesia.

Intracellular Zn2+ promotes extracellular matrix remodeling in dexamethasone-treated trabecular meshwork.

Given the widespread application of glucocorticoids in ophthalmology, the associated elevation of intraocular pressure (IOP) has long been a vexing concern for clinicians, yet the underlying mechanisms remain inconclusive. Much of the discussion focuses on the extracellular matrix (ECM) of trabecular meshwork (TM). It is widely agreed that glucocorticoids impact the expression of matrix metalloproteinases (MMPs), leading to ECM deposition. Since Zn2+ is vital for MMPs, we explored its role in ECM alterations induced by dexamethasone (DEX). Our study revealed that in human TM cells treated with DEX, the level of intracellular Zn2+ significantly decreased, accompanied by impaired extracellular Zn2+ uptake. This correlated with changes in several Zrt-, Irt-related proteins (ZIPs) and metallothionein. ZIP8 knockdown impaired extracellular Zn2+ uptake, but Zn2+ chelation did not affect ZIP8 expression. Resembling DEX's effects, chelation of Zn2+ decreased MMP2 expression, increased the deposition of ECM proteins, and induced structural disarray of ECM. Conversely, supplementation of exogenous Zn2+ in DEX-treated cells ameliorated these outcomes. Notably, dietary zinc supplementation in mice significantly reduced DEX-induced IOP elevation and collagen content in TM, thereby rescuing the visual function of the mice. These findings underscore zinc's pivotal role in ECM regulation, providing a novel perspective on the pathogenesis of glaucoma.NEW & NOTEWORTHY Our study explores zinc's pivotal role in mitigating extracellular matrix dysregulation in the trabecular meshwork and glucocorticoid-induced ocular hypertension. We found that in human trabecular meshwork cells treated with dexamethasone, intracellular Zn2+ significantly decreased, accompanied by impaired extracellular Zn2+ uptake. Zinc supplementation rescues visual function by modulating extracellular matrix proteins and lowering intraocular pressure, offering a direction for further exploration in glaucoma management.

Glucocorticoid-Induced Ocular Hypertension and Glaucoma.

Glucocorticoid (GC) therapy is indicated in many diseases, including ocular diseases. An important side-effect of GC therapy is GC-induced ocular hypertension (GIOHT), which may cause irreversible blindness known as GC-induced glaucoma (GIG). Here, we reviewed the pathological changes that contribute to GIOHT including in the trabecular meshwork and Schlemm's canal at cellular and molecular levels. We also discussed the clinical aspects of GIOHT/GIG including disease prevalence, risk factors, the type of GCs, the route of GC administration, and management strategies.

The Effects of ROCK Inhibitor on Prevention of Dexamethasone-Induced Glaucoma Phenotype in Human Trabecular Meshwork Cells.

This study investigated the effects of dexamethasone (Dex) on human trabecular meshwork (TM) cells, a model of glucocorticoid-induced glaucoma, and evaluated the impact of ripasudil (Rip) as a co-delivery or sequential dosing strategy. In vitro experiments were conducted to assess the effects of Dex and Rip on TM cells. Confocal microscopy was used to evaluate the impact of Dex and Rip on F-actin staining signals. Contractility of the TM cells upon Dex and Rip treatment mimicking co-delivery and sequential delivery was quantified using collagen gel contraction assay. Transepithelial electrical resistance (TEER) values and fluorescein isothiocyanate (FITC)-dextran permeability were also measured to assess the impact of Dex and Rip on TM cells. Dex and Rip did not exhibit cytotoxicity at the maximum tested concentration (20 µM). Dex-treated TM cells exhibited higher F-actin staining signals compared to controls, which were reduced when co-treated with Rip. Rip inhibited Dex-induced collagen gel contraction activity in both co-delivery and sequential treatments. Dex resulted in increased TEER values as the dose increased, whereas TEER values were maintained when co-treated with Rip. Co-delivery of Rip has the potential to prevent glaucoma symptoms when patients are treated with Dex. This study highlights the importance of identifying strategies to reduce the side effects of prolonged use of glucocorticoids, such as Dex, in the treatment of various diseases. This study demonstrates the potential of co-delivering ripasudil with dexamethasone to mitigate glucocorticoid-induced ocular hypertension and a secondary glaucoma that resembles primary open-angle glaucoma, providing insights for the development of novel preventive strategies in clinical care.

Short and long-term effect of dexamethasone on the transcriptome profile of primary human trabecular meshwork cells in vitro

In the quest of identifying newer molecular targets for the management of glucocorticoid-induced ocular hypertension (GC-OHT) and glaucoma (GCG), several microarray studies have attempted to investigate the genome-wide transcriptome profiling of primary human trabecular meshwork (TM) cells in response to dexamethasone (DEX). However, no studies are reported so far to demonstrate the temporal changes in the expression of genes in the cultured human TM cells in response to DEX treatment. Therefore, in the present study, the time-dependent changes in the genome-wide expression of genes in primary human TM cells after short (16 hours: 16 h) and long exposure (7 days: 7 d) of DEX was investigated using RNA sequencing. There were 199 (118 up-regulated; 81 down-regulated) and 525 (119 up-regulated; 406 down-regulated) DEGs in 16 h and 7 d treatment groups respectively. The unique genes identified in 16 h and 7 d treatment groups were 152 and 478 respectively. This study found a distinct gene signature and pathways between two treatment regimes. Longer exposure of DEX treatment showed a dys-regulation of Wnt and Rap1 signaling and so highlighted potential therapeutic targets for pharmacological management of GC-OHT/glaucoma.

Determination of the Role of the Distal Outflow Pathway Tissue in Glucocorticoid-induced Ocular Hypertension

Introduction
 Prolonged application of glucocorticoids (GCs) induces ocular hypertension (OHT) and glaucoma. This increased intraocular pressure (IOP) is due to pathological changes in the trabecular meshwork (TM) outflow pathway tissues including impaired cell functions and extracellular matrix deposition. The changes and role of the TM in GC-induced OHT have been well studied. However, the role of the tissues distal to the TM (distal outflow tissues) is unclear. This study aims to further uncover the role of distal outflow tissue in GC-induced OHT using a novel perfusion organ culture (POC) model.
 Methods
 Huma corneal rims tissues were attached to 3D printed transparent perfusion plates using a combination of thin and thick glues. The artificial anterior chamber was perfused with DMEM-low glucose medium at 2ul/min to mimic aqueous humor production, and IOP was recorded using pressure transducers and a computerized system. To determine the role of distal tissue in GC-induced IOP changes, the TM tissue was carefully removed from both eyes, and one eye was treated with ethanol (EtOH) and the fellow eye with dexamethasone (DEX).
 Results
 The model was validated through a comparison of the IOP and TM stiffness of glue contaminated to non-contaminated corneal rims. The glue contaminated rim showed highly increased IOP and TM stiffness while the non-contaminated rim showed normal values. After validation, the TM was removed from paired corneal rims. One rim was treated with 100nM DEX and the fellow rim with 0.1% EtOH. The DEX treated rim showed increase in IOP while the EtOH control showed little change.
 Conclusion
 We created a novel corneal rim perfusion culture model for the study of GC-induced OHT. This model showed promising results of distal outflow involvement in glucocorticoid induced ocular hypertension. Further studies are needed to elucidate the role of distal outflow tissues in GC responsiveness in the eye.

Optogenetic Modulation of Intraocular Pressure in a Glucocorticoid-Induced Ocular Hypertension Mouse Model.

PurposeSteroid-induced glaucoma is a common form of secondary open angle glaucoma characterized by ocular hypertension (elevated intraocular pressure [IOP]) in response to prolonged glucocorticoid exposure. Elevated IOP occurs with increased outflow resistance and altered trabecular meshwork (TM) function. Recently, we used an optogenetic approach in TM to regulate the 5-phosphatase, OCRL, which contributes to regulating PI(4,5)P2 levels. Here, we applied this system with the aim of reversing compromised outflow function in a steroid-induced ocular hypertension mouse model.MethodsElevated IOP was induced by chronic subconjunctival dexamethasone injections in wild-type C57Bl/6j mice. AAV2 viruses containing optogenetic modules of cryptochrome 2 (Cry2)-OCRL-5ptase and CIBN-GFP were injected into the anterior chamber. Four weeks after viral expression and dexamethasone exposure, IOP was measured by tonometer and outflow facility was measured by perfusion apparatus. Human TM cells were treated with dexamethasone, stimulated by light and treated with rhodamine-phalloidin to analyze actin structure.ResultsDexamethasone treatment elevated IOP and decreased outflow facility in wild-type mice. Optogenetic constructs were expressed in the TM of mouse eyes. Light stimulation caused CRY2-OCRL-5ptase to translocate to plasma membrane (CIBN-CAAX-GFP) and cilia (CIBN-SSTR3-GFP) in TM cells, which rescued the IOP and outflow facility. In addition, aberrant actin structures formed by dexamethasone treatment were reduced by optogenetic stimulation in human TM cells in culture.ConclusionsSubcellular targeting of inositol phosphatases to remove PIP2 represents a promising strategy to reverse defective TM function in steroid-induced ocular hypertension.Translational RelevanceTargeted modulation of OCRL may be used to decrease steroid-induced elevated IOP.

Anti-fibrotic activity of a rho-kinase inhibitor restores outflow function and intraocular pressure homeostasis.

Glucocorticoids are widely used as an ophthalmic medication. A common, sight-threatening adverse event of glucocorticoid usage is ocular hypertension, caused by dysfunction of the conventional outflow pathway. We report that netarsudil, a rho-kinase inhibitor, decreased glucocorticoid-induced ocular hypertension in patients whose intraocular pressures were poorly controlled by standard medications. Mechanistic studies in our established mouse model of glucocorticoid-induced ocular hypertension show that netarsudil both prevented and reduced intraocular pressure elevation. Further, netarsudil attenuated characteristic steroid-induced pathologies as assessed by quantification of outflow function and tissue stiffness, and morphological and immunohistochemical indicators of tissue fibrosis. Thus, rho-kinase inhibitors act directly on conventional outflow cells to prevent or attenuate fibrotic disease processes in glucocorticoid-induced ocular hypertension in an immune-privileged environment. Moreover, these data motivate the need for a randomized prospective clinical study to determine whether netarsudil is indeed superior to first-line anti-glaucoma drugs in lowering steroid-induced ocular hypertension.

The Canonical Wnt Signaling Pathway Inhibits the Glucocorticoid Receptor Signaling Pathway in the Trabecular Meshwork

Glucocorticoid-induced glaucoma is a secondary open-angle glaucoma. About 40% of the general population may develop elevated intraocular pressure on prolonged glucocorticoid treatment secondary to damages in the trabecular meshwork (TM), a tissue that regulates intraocular pressure. Therefore, identifying the key molecules responsible for glucocorticoid-induced ocular hypertension is crucial. In this study, Dickkopf-related protein 1 (Dkk1), a canonical Wnt signaling inhibitor, was found to be elevated in the aqueous humor and TM of glaucoma patients. At the signaling level, Dkk1 enhanced glucocorticoid receptor (GR) signaling, whereas Dkk1 knockdown or Wnt signaling activators decreased GR signaling in human TM cells as indicated by luciferase assays. Similarly, activation of the GR signaling inhibited Wnt signaling. At the protein level, glucocorticoid-induced extracellular matrix was inhibited by Wnt activation using Wnt activators or Dkk1 knockdown in primary human TM cells. In contrast, inhibition of canonical Wnt signaling by β-catenin knockdown increased glucocorticoid-induced extracellular matrix proteins. At the physiological level, adenovirus-mediated Wnt3a expression decreased glucocorticoid-induced ocular hypertension in mouse eyes. In summary, Wnt and GR signaling inhibit each other in the TM, and canonical Wnt signaling activators may prevent the adverse effect of glucocorticoids in the eye.

Mitochondria and Autophagy Dysfunction in Glucocorticoid-Induced Ocular Hypertension/Glaucoma Mice Model

ABSTRACTPurpose: This study is aimed to investigate the effects of periocular steroids induction on intraocular pressure (IOP), retinal ganglion cells (RGCs) and trabecular meshwork (TM) ultrastructure in glucocorticoid-induced ocular hypertension mice model.Materials and Methods: Dexamethasone-21-acetate (Dex-Ace) was administered through periocular conjunctival fornix injection every 3 days in C57BL/6J mice. Intraocular pressure was measured weekly by rebound tonometry. RGCs were examined with immunofluorescent staining of BRN3a at week 1, 4, and 8. TM morphology was visualized with electron microscopy. Autophagy was evaluated with immunoblotting in TM tissues.Results: Dex-Ace rapidly and significantly induced IOP, which peaked at week 4. The absolute increase in IOP in the Dex-Ace-treated mice was 8.1 ± 1.4 mmHg, a 60% induction (p < .0001) compared with that in the vehicle-treated mice. The IOP sustained a higher level in the Dex-Ace group from week 4 to week 8. Dex-Ace treatment decreased the number of RGCs in a time-dependent manner, suggesting that high IOP resulted in optic neuropathy. In addition, Dex-Ace thickened trabecular beams and decreased intertrabecular spaces, with marked accumulation of fibrillar and amorphous granular extracellular material. Moreover, Dex-Ace induced swollen and elongated mitochondria in TM cells. The average mitochondria area was 0.090 ± 0.044 µm2 in the vehicle-treated mice, and increased to 0.161 ± 0.094 µm2 (p < .0001), 0.121 ± 0.029 µm2 (p = .0223) and 0.171 ± 0.076 µm2 (p < .0001) in the Dex-Ace-treated mice at weeks 1, 4 and 8, respectively. Autophagy was also increased by Dex-Ace treatment, indicating by the upregulation of LC3-I, LC3-II and beclin-1, and downregulation of p62.Conclusion: Dex-Ace administration decreased RGCs and changed TM ultrastructure, mimicking hallmarks of human glucocorticoid-induced glaucoma (GIG). In addition, mitochondria and autophagy dysfunction suggested abnormal energy metabolism in TM cells, which warranted further study to fully elucidate the pathogenesis of GIG.

Glucocorticoid Receptor Transactivation Is Required for Glucocorticoid-Induced Ocular Hypertension and Glaucoma.

PurposeGlucocorticoid (GC)–induced ocular hypertension (GC-OHT) is a serious side effect of prolonged GC therapy that can lead to glaucoma and permanent vision loss. GCs cause a plethora of changes in the trabecular meshwork (TM), an ocular tissue that regulates intraocular pressure (IOP). GCs act through the glucocorticoid receptor (GR), and the GR regulates transcription both through transactivation and transrepression. Many of the anti-inflammatory properties of GCs are mediated by GR transrepression, while GR transactivation largely accounts for GC metabolic effects and side effects of GC therapy. There is no evidence showing which of the two mechanisms plays a role in GC-OHT.MethodsGRdim transgenic mice (which have active transrepression and impaired transactivation) and wild-type (WT) C57BL/6J mice received weekly periocular dexamethasone acetate (DEX-Ac) injections. IOP, outflow facilities, and biochemical changes to the TM were determined.ResultsGRdim mice did not develop GC-OHT after continued DEX treatment, while WT mice had significantly increased IOP and decreased outflow facilities. Both TM tissue in eyes of DEX-treated GRdim mice and cultured TM cells isolated from GRdim mice had reduced or no change in the expression of fibronectin, myocilin, collagen type I, and α-smooth muscle actin (α-SMA). GRdim mouse TM (MTM) cells also had a significant reduction in DEX-induced cytoskeletal changes, which was clearly seen in WT MTM cells.ConclusionsWe provide the first evidence for the role of GR transactivation in regulating GC-mediated gene expression in the TM and in the development of GC-OHT. This discovery suggests a novel therapeutic approach for treating ocular inflammation without causing GC-OHT and glaucoma.

Glucocorticoid Receptor Transactivation is Required for Glucocorticoid-Induced Ocular Hypertension and Glaucoma: A Deleterious Effect of Chronic Glucocorticoid Therapy

Glucocorticoid (GC)-induced ocular hypertension (OHT) is a serious side-effect of prolonged GC therapy that can lead to glaucoma and permanent vision loss. GCs cause a plethora of changes in the trabecular meshwork (TM), an ocular tissue that regulates intraocular pressure (IOP). GCs act through the glucocorticoid receptor (GR), and the GR regulates transcription both through transactivation and transrepression. Many of the anti-inflammatory properties of GCs are mediated by GR transrepression, while GR transactivation largely accounts for GC metabolic effects and side effects of GC therapy. There is no evidence showing which of the two mechanisms play role in GC- OHT. Methods: GRdim transgenic mice (which have active transrepression and impaired transactivation) and wild type (WT) C57BL/6J mice received weekly periocular DEX-Ac injections. IOP, outflow facilities, and biochemical changes to the TM were determined. Results: GRdim mice did not develop GC-OHT DEX treatment, while WT mice had significantly increased IOP and decreased outflow facilities. Both TM tissue in eyes of DEX-treated GRdim mice and cultured TM cells isolated from GRdim mice had reduced or no change in the expression of fibronectin, myocilin, collagen type I, and α-SMA. GRdim MTM cells also had a significant reduction in DEX-induced cytoskeletal changes, which was clearly seen in WT MTM cells. Conclusion: We provide the first evidence for the role of GR transactivation in regulating GC-mediated gene expression in the TM and in the development of GC- OHT. This discovery suggests a novel therapeutic approach for treating ocular inflammation that will not cause the serious side effect of GC-OHT and glaucoma. Funding: NIH/NEI EY016242. Declaration of Interest: AFC is a consultant for Goodmans LLP, LayerBio, Kala Pharmacuticals and has received research support from Unity Biotechnology and Lung Therapeutics. However, none of these are related to the study presented. The other authors declare that they have no competing interests. Ethical Approval: All experimental protocols were approved by University of North Texas Health Science Center Institutional Animal Care and Use Committee regulations.

Transforming growth factor β2 (TGFβ2) signaling plays a key role in glucocorticoid-induced ocular hypertension

Elevation of intraocular pressure (IOP) is a serious adverse effect of glucocorticoid (GC) therapy. Increased extracellular matrix (ECM) accumulation and endoplasmic reticulum (ER) stress in the trabecular meshwork (TM) is associated with GC-induced IOP elevation. However, the molecular mechanisms by which GCs induce ECM accumulation and ER stress in the TM have not been determined. Here, we show that a potent GC, dexamethasone (Dex), activates transforming growth factor β (TGFβ) signaling, leading to GC-induced ECM accumulation, ER stress, and IOP elevation. Dex increased both the precursor and bioactive forms of TGFβ2 in conditioned medium and activated TGFβ-induced SMAD signaling in primary human TM cells. Dex also activated TGFβ2 in the aqueous humor and TM of a mouse model of Dex-induced ocular hypertension. We further show that Smad3-/- mice are protected from Dex-induced ocular hypertension, ER stress, and ECM accumulation. Moreover, treating WT mice with a selective TGFβ receptor kinase I inhibitor, LY364947, significantly decreased Dex-induced ocular hypertension. Of note, knockdown of the ER stress-induced activating transcription factor 4 (ATF4), or C/EBP homologous protein (CHOP), completely prevented Dex-induced TGFβ2 activation and ECM accumulation in TM cells. These observations suggested that chronic ER stress promotes Dex-induced ocular hypertension via TGFβ signaling. Our results indicate that TGFβ2 signaling plays a central role in GC-induced ocular hypertension and provides therapeutic targets for GC-induced ocular hypertension.

Glucocorticoid receptor GR\u03b2 regulates glucocorticoid-induced ocular hypertension in mice

Prolonged glucocorticoid (GC) therapy can cause GC-induced ocular hypertension (OHT), which if left untreated progresses to iatrogenic glaucoma and permanent vision loss. The alternatively spliced isoform of glucocorticoid receptor GRβ acts as dominant negative regulator of GR activity, and it has been shown that overexpressing GRβ in trabecular meshwork (TM) cells inhibits GC-induced glaucomatous damage in TM cells. The purpose of this study was to use viral vectors to selectively overexpress the GRβ isoform in the TM of mouse eyes treated with GCs, to precisely dissect the role of GRβ in regulating steroid responsiveness. We show that overexpression of GRβ inhibits GC effects on MTM cells in vitro and GC-induced OHT in mouse eyes in vivo. Ad5 mediated GRβ overexpression reduced the GC induction of fibronectin, collagen 1, and myocilin in TM of mouse eyes both in vitro and in vivo. GRβ also reversed DEX-Ac induced IOP elevation, which correlated with increased conventional aqueous humor outflow facility. Thus, GRβ overexpression reduces effects caused by GCs and makes cells more resistant to GC treatment. In conclusion, our current work provides the first evidence of the in vivo physiological role of GRβ in regulating GC-OHT and GC-mediated gene expression in the TM.

Differential response and withdrawal profile of glucocorticoid-treated human trabecular meshwork cells

The goal of the study was to examine secreted protein response and withdrawal profiles from cultured human trabecular meshwork (HTM) cells following short- and long-term glucocorticoid treatment. Primary cultures of five human HTM cell strains isolated from 5 different individual donor eyes were tested. Confluent HTM cells were differentiated in culture media containing 1% FBS for at least one week, and then treated with Dexamethasone (Dex, 100 nM) 3 times/week for 1 or 4 weeks. Cell culture supernatants were collected 3 times per week for 8 weeks. Secretion profiles of myocilin (MYOC), matrix metalloproteinase-2 (MMP2) and fibronectin (FN) were determined by Western blot analysis and MMP2 activity by zymography. Dex treatment reduced MMP2 expression and activity, returning to normal levels shortly after Dex withdrawal in 5 HTM cell strains. All five cell strains significantly upregulated MYOC in response to Dex treatment by an average of 17-fold, but recovery to basal levels after Dex withdrawal took vastly different periods of time depending on cell strain and treatment duration. Dex treatment significantly increased FN secretion in all strains but one, which decreased FN secretion in the presence of Dex. Interestingly, secretion of FN and MYOC negatively correlated during a 4 week recovery period following 4 weeks of Dex treatment. Taken together, the time course and magnitude of response and recovery for three different secreted, extracellular matrix-associated proteins varied greatly between HTM cell strains, which may underlie susceptibility to glucocorticoid-induced ocular hypertension.

Clinical relevance of the glucocorticoid receptor gene polymorphisms in glucocorticoid-induced ocular hypertension and primary open angle glaucoma.

To avoid the side effects of ocular hypertension of glucocorticoid (GC) usage in eye, we must identify susceptible individuals, which exists in about one-third of all population. Further, the majority of all primary open angle glaucoma (POAG) patients show this phenotype. Glucocorticoid receptor (GR) regulates C responsiveness in trabecular meshwork (TM) cells. In this study, single nucleotide polymorphism (SNP) genotyping was used to determine whether there are differences in the BclI (rs41423247) and N363S (rs6195) polymorphisms of the GR gene in healthy and POAG patients, and glucocorticoid-induced ocular hypertension (GIOH) populations. Three hundred and twenty-seven unrelated Chinese adults, including 111 normal controls, 117 GIOH subjects and 99 POAG patients, were recruited. DNA samples were prepared and the BclI and N363S polymorphisms were screened using real-time polymerase chain reaction (RT-PCR)-restriction fragment length polymorphism (RFLP) analysis. Frequencies of the BclI and N363S polymorphisms were determined and compared using Fisher's exact test and the Chi-squared test. Only the BclI polymorphism was identified in the Chinese Han population. The frequency of the G allele was 21.6 % in normal controls, 18.3% in GIOH patients, and 13.64% in the POAG patients. There was no significant difference in polymorphism or allele frequency in the 3 groups. Furthermore, no N363S polymorphism was found in the study subjects. The BclI polymorphisms in GR gene had no association with GIOH and POAG patients, and N363S polymorphism might not exist in the Chinese Han population. Therefore, the BclI polymorphism might not be responsible for the development of GC-induced ocular hypertension or POAG.

A useful mouse model of glucocorticoid-induced ocular hypertension.

Glucocorticoid (GC)-induced ocular hypertension and iatrogenic glaucoma still are a major side effect associated with GC therapy, especially with the increased use of intravitreally administered GCs for a variety of retinal conditions. In this issue, Overby and colleagues1 investigated GC-induced ocular hypertension in mice. They built on the success of Whitlock and coworkers,2 who previously reported GC-induced ocular hypertension in mice by systemic delivery of the potent GC dexamethasone (DEX) using osmotic minipumps. Overby et al.1 showed that systemic DEX delivery significantly elevated IOP (by approximately 3 mm Hg), which correlated with a 52% reduction in the aqueous outflow facility. They also showed DEX-induced changes in the extracellular matrix of the trabecular meshwork (TM) that were very similar to the histologic changes observed in humans with steroid-induced ocular hypertension and POAG, including increased fibrillar material in juxtacanalicular tissue, plaque-like sheath material surrounding elastic fibers in the TM, and myofibroblasts on the outer wall of Schlemm's canal. It should be noted that an important side effect of this route of DEX administration was considerable weight loss in a significant percentage of animals during the 4 weeks of systemic exposure. This will be an extremely useful animal model to study the molecular mechanisms responsible for GC-induced ocular hypertension as well as POAG. The power of mouse genetics will allow determination of the involvement of specific genes and signaling pathways in this model. In addition, there are significant differences in human responsiveness to GC-induced ocular hypertension, so a mouse strain survey may help uncover the mechanisms responsible for these differences in GC sensitivity.

Ocular-specific ER stress reduction rescues glaucoma in murine glucocorticoid-induced glaucoma

Administration of glucocorticoids induces ocular hypertension in some patients. If untreated, these patients can develop a secondary glaucoma that resembles primary open-angle glaucoma (POAG). The underlying pathology of glucocorticoid-induced glaucoma is not fully understood, due in part to lack of an appropriate animal model. Here, we developed a murine model of glucocorticoid-induced glaucoma that exhibits glaucoma features that are observed in patients. Treatment of WT mice with topical ocular 0.1% dexamethasone led to elevation of intraocular pressure (IOP), functional and structural loss of retinal ganglion cells, and axonal degeneration, resembling glucocorticoid-induced glaucoma in human patients. Furthermore, dexamethasone-induced ocular hypertension was associated with chronic ER stress of the trabecular meshwork (TM). Similar to patients, withdrawal of dexamethasone treatment reduced elevated IOP and ER stress in this animal model. Dexamethasone induced the transcriptional factor CHOP, a marker for chronic ER stress, in the anterior segment tissues, and Chop deletion reduced ER stress in these tissues and prevented dexamethasone-induced ocular hypertension. Furthermore, reduction of ER stress in the TM with sodium 4-phenylbutyrate prevented dexamethasone-induced ocular hypertension in WT mice. Our data indicate that ER stress contributes to glucocorticoid-induced ocular hypertension and suggest that reducing ER stress has potential as a therapeutic strategy for treating glucocorticoid-induced glaucoma.

Perfusion-Cultured Bovine Anterior Segments as an Ex Vivo Model for Studying Glucocorticoid-Induced Ocular Hypertension and Glaucoma

To determine whether perfusion-cultured bovine anterior segments would be a suitable model for glaucoma research. Fresh bovine eyes were dissected and sealed on a custom-made acrylic dish with an O-ring. Perfusion medium was infused by a syringe pump at a constant infusion rate of 5 μL/min. After intraocular pressure (IOP) was stable, bovine eyes were perfused with medium containing either a vehicle control (0.1% ethanol [ETH]) or dexamethasone (DEX) for up to 7 days. IOP was recorded by a pressure transducer and a computerized system. Perfusion medium was collected for Western immunoblot analysis of myocilin (MYOC). The morphology of the bovine trabecular meshwork after perfusion culture was similar to that of freshly dissected, nonperfused bovine eyes. Treatment with DEX elevated IOP in some bovine eyes, whereas others showed little change. The authors analyzed the data from 18 ETH-treated control eyes and defined 2.82 mm Hg as the threshold of ocular hypertension (OHT), which equals mean pressure change + 2× SD. Approximately 40% (12/29) of the bovine eyes were DEX responders, which is very close to the DEX-responsive rates observed in human and monkey eyes. Western blot data showed that DEX treatment induced the expression of the DEX-inducible gene MYOC only in the perfusion-cultured anterior segments with DEX-induced OHT. OHT can be induced by DEX in perfusion-cultured bovine anterior segments. This is a fast, convenient, affordable, and reliable model for studying DEX-induced OHT and the mechanisms of trabecular outflow.

Expression of glucocorticoid receptors alpha and beta in mononuclear cells of peripheral blood in patients with glucocorticoid-induced ocular hypertension

To study the expression of glucocorticoid receptors (alpha and beta) in mononuclear cells of peripheral blood (PBMCs) in patients with glucocorticoid-induced ocular hypertension (GIOH) and the relationship between of the expression of GRalpha and GRbeta in PBMCs and its susceptibility to GIOH. Case control study. Thirty-two patients with anaphylactic conjunctivitis were enrolled in this study. All of whom were received topical application of 0.1% dexamethasone. The patients with the elevation of intraocular pressure (IOP) over 8 mm Hg(1 mm Hg = 0.133 kPa) or higher within two weeks were defined as GIOH and the others were considered as control group. There were no statistical difference in sex, average age, and baseline IOP between two groups. Each group consisted of sixteen patients. GRalpha and GRbeta mRNA (relative value) in PBMCs were detected with semi-quantitative reverse transcriptional polymerase chain reaction (RT-PCR), GRalpha and GRbeta protein (positive ratio) in PBMCs examined with fluorescent labelling flow cytometry. The GRalpha mRNA/GAPDH ratio of GIOH group was significantly (t = 3.872, P < 0.05) higher than that of control group (1.152 +/- 0.057 vs 1.048 +/- 0.031). The GRbeta mRNA/GAPDH of GIOH group was lower than that of control group's (1.055 +/- 0.034 vs 1.063 +/- 0.035), but was not statistically different (t = 0.419, P > 0.05). GRalpha positive percentage was significantly (t = 8.513, P < 0.01) higher in PBMCs of GIOH group than that of the control group (93.10 +/- 7.35)% vs (46.00 +/- 13.11)%, whereas GRbeta positive percentage was significantly (t = 4.842 P < 0.01) lower in PBMCs of GIOH group than that in control group (12.50 +/- 4.18)% vs (23.83 +/- 3.92)%. The higher expression of GRalpha in PBMCs in patients may have an implication of susceptibility to GIOH and warrant further investigation.