Eye / Vision

Permanent URI for this collectionhttps://hdl.handle.net/20.500.12503/30436


Recent Submissions

Now showing 1 - 11 of 11
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    Evaluation of Neuroprotective Effect of hybrid compound SA-2 Nanosuspension in Optic Nerve Crush Mouse Model.
    (2021) Amankwa, Charles E.; Gondi, Sudershan; Funk, Arlene; Debnath, Biddut; Zhang, Wei; Li, Linya; Ferguson, Jonathan; Johnson, Gretchen; Ellis, Dorette; Stankowska, Dorota; Acharya, Suchismita
    This study examines the cytoprotective effects of hybrid antioxidant-nitric oxide (NO) donating compound SA-2 nanosuspension in normal trabecular meshwork-5 cells (NTM-5) and optic nerve crush (ONC) induced-mouse retinal ganglion cell (RGC) death model. SA-2 was encapsulated in poly-lactic-co-glycolic acid (PLGA) nanoparticles and reported earlier. Total nitrite concentrations of SA-2-NPs was determined using Griess assay in both buffer and NTM-5 cell supernatants. NTM-5 cells were treated with tert-butyl hydrogen peroxide (TBHP, EC50 = 5.5mM) at varying concentrations of SA-2 NPs (1, 0.5, 0.25, 0.125 % w/v) and cell viability was measured. 12-week-old C57BL/6J mice were subjected to ONC injury in the left eye and were administered with either vehicle or 1% SA-2-NPs via intravitreal injections (2uL) or eye drops (5uL). Pattern ERG (PERG) was used to assess retinal function and RGC survival was determined using RBPMS-positive RGCs. Data were presented as mean ± SEM, n=3-8. 1% SA-2-NPs dose dependently increased NTM-5 cell viability and total nitrite concentrations (~50%) lasting over 4 days. There was significant improvement in PERG amplitudes (1.7-1.9 times) following SA-2-NP administration as well as increased RGC numbers (by ~20%) relative to ONC-ed eyes. SA-2 nanosuspension shows strong positive trend in protecting RGC's from oxidative stress-induced apoptosis in ONC rodent model. Further investigation is being done for improved dosing and signaling changes in response to SA-2 NPs therapy.
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    Metabolic changes in optic nerve head astrocytes following glaucoma-associated deformation
    (2021) Pappenhagen, Nathaniel; Zaman, Khadiza; Prokai, Laszlo; Inman, Denise
    Purpose: The astrocytes of the optic nerve head (ONHAs) are the predominant glial cell in the unmyelinated portion of the optic nerve. We tested the metabolic changes these cells undergo in glaucoma by exposing them in vitro to degrees of deformation similarly experienced as a result of increased intraocular pressure in primary open angle glaucoma. Methods: Primary astrocytes were cultured from the cortices of P1 mouse pups or P7 optic nerve head explants then seeded on collagen-coated FlexCell plates. The astrocytes were then biaxially stretched by 12% for 24 hours using the FX-6000T FlexCell. ONHA extracellular acidification rate (ECAR) and oxygen consumption rate (OCR) were measured using the Seahorse XFe24 Analyzer, while changes in the proteome were measured using mass spectrometry. Results: Stretched cortical astrocytes showed at least twofold increases in metabolic proteins such as glutamate dehydrogenase 1, isocitrate dehydrogenase 1, and aldolase fructose-bisphosphate c. There was also at least a twofold decrease in citrate synthase, isocitrate dehydrogenase 2, glycogen phosphorylase B, and adenylate kinase 1. In the Seahorse Analyzer, stretched ONHAs showed increased glycolytic ECAR, maximal ECAR, and maximal OCR. Stretched astrocytes showed no difference in their dependence on pyruvate compared to controls, but a significant decrease in their capacity for mitochondrial respiration from pyruvate. Conclusions: Exposing astrocytes to glaucoma-associated deformation altered their metabolism in ways that indicated stronger commitment to glycolysis compared to control astrocytes, such as increases in glycolytic proteins, decreases in mitochondrial proteins, and increases in glycolytic ECAR.
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    Evaluating the Relationship between Race and Amblyogenic Risk Factors in Preschool Children in Fort Worth, Texas
    (2021) Karsaliya, Gopal; Omar, Salma; Manning, Sydney E.; Luna-Smith, Annabel; Aryal, Subhash; Mozdbar, Sima
    Background Amblyopia (lazy eye) is the most common cause of vision loss in children. The prevalence is between 2-5% in the United States. Amblyogenic risk factors include early visual deprivation, strabismus, anisometropia, and media opacities such as a cataract. If treatment for amblyopia is not initiated before the age of 7, the likelihood of successful correction begins to drastically decline with age. Previous studies have found correlations between race and various vision abnormalities. This study aims to assess the incidence of myopia, hyperopia, and astigmatism among pre-Kindergarten children of different racial groups in Fort Worth, TX, as well as evaluate any racial differences in the presence of amblyogenic risk factors. Methods Using the PlusoptiX refractometers at 37 local elementary schools, researchers collected refractive error data of N=2,258 children under the age of 6, allowing for the detection of hyperopia, myopia, astigmatism, and anisometropia. The children's age, race, and sex were also recorded. A chi-square test was done to compare proportions of male/female participants, and odds ratios were calculated for each amblyogenic risk factor between racial/ethnic groups. Results Those with at least one amblyogenic risk factor accounted for 27.82% of the sample. There was a significant difference in astigmatism as an amblyogenic risk factor between Black and Hispanic groups (OR=0.6039, 95%CI 0.46-0.79) and between White and Hispanic groups (OR=0.2387, 95%CI 0.17-0.34). Conclusion The results of this study suggest Hispanic children were at increased risk of developing amblyopia compared to Black and White children in Fort Worth.
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    TGFβ2 induces Chronic Endoplasmic Reticulum Stress in Trabecular Meshwork cells
    (2021) Patil, Shruti; Kasetti, Ramesh; Maddineni, Prabhavathi; Zode, Gulab
    Purpose:TGFβ2 induced extracellular matrix(ECM)accumulation is known to be associated with glaucomatous trabecular meshwork(TM)damage and IOP elevation. Previously, we have demonstrated that abnormal ECM accumulation leads to endoplasmic reticulum(ER)stress in TM. Here, we investigated whether TGFβ2 induces ER stress in TM cells and whether chronic ER stress plays a pathological role in dysfunction of TM cells. Methods:Primary human TM cells were treated with vehicle or recombinant TGFβ2(5ng/mL) to determine its effect on chronic ER stress markers(GRP78,ATF4 and CHOP)and ECM proteins(Fibronectin and Collagen-I/IV). Chronic ER stress-induced ATF4/CHOP were genetically knocked-down using targeted CRISPR/Cas9 expression plasmids, or by transducing with ad5-ATF4deltaRK that inhibits endogenous ATF4 activity. The effect on IOP of intravitreal gene delivery of active-TGFβ2 viral-vector in Chop-/- and C57BL/6J mice was evaluated. Pharmacological inhibition of ER stress using ATF4/CHOP inhibitor ISRIB and chemical chaperon sodium4phenylbutyrate(PBA) was also determined on TGFβ2-treated TM cells. Results:Westernblot and immunostaining demonstrated that TGFβ2 induced chronic ER stress markers along with increased levels of ECM proteins, suggesting TGFβ2-induced ECM deposition is associated with ER stress. Knockdown of key transcriptional factors, ATF4/CHOP, and ISRIB treatment prevented TGFβ2-induced ECM expression and reduced ER stress in TM cells. Moreover, activeTGFβ2 viral delivery caused no IOP elevation in Chop-/-mice compared to control C57BL/6J mice. Treatment of TM cells with PBA also inhibited TGFβ2-induced fibronectin deposition via induced expression and activation of MMP2/9. Conclusion:This study indicates that TGFβ2 induces chronic ER stress, which is associated with increased ECM accumulation.
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    A Systematic Pharmacology Analysis of the Age-Related Eye Disease Study 2 (AREDS2) formula and its role in preventing Age-Related Macular Degeneration (AMD)
    (2021) Au, My-Lien; Yu, Yu; Lou, Alexander; Garcia, Luis; Tran, Myhoa; Wu, Hongli
    Purpose: According to the major clinical trial sponsored by the National Eye Institute (NEI), oral supplementation with the Age-Related Eye Disease Study 2 (AREDS2) formulation (vitamins C and E, zinc, copper, lutein, and zeaxanthin) has been shown to delay the progression of advanced age- related macular degeneration (AMD). However, the detailed pharmacological mechanisms of AREDS2 are not fully understood at the molecular level. In this study, we intend to develop a systematic approach to predict AREDS2-associated targets and to map the drug-disease-target network. Methods: Genes of interest were identified via the NCBI database for compounds in the AREDS2 formula. Cytoscape software was used to visually create a network of source and target nodes to analyze the similarities between them. The formula's relation to AMD was analyzed via the Gene2Function and GeneCard databases. Results: A total of 158 genes were identified as the targets of the AREDS2 formula. 27 of these genes were a result of multiple components of the AREDS2 formula. The main pathways that these genes affect were identified and mapped out to include lipid metabolism, DNA damage responses, and oxidative stress. The top 5 genes regulated by the most components of the AREDS2 formula are GSTP1, Nrf2, VEGFA, HIF1A, and CXCL8. Conclusions: A systematic pharmacology-based approach provides beneficial information for elucidating the potential mechanisms of action of the AREDS2 formula in treating AMD. Furthermore, it provides future direction for AMD treatment which may focus on anti-adipogenic, anti-inflammatory, and anti-oxidant pathways.
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    Functions of glutaredoxin 2 (Grx2) in the retina: Mechanisms and Protection
    (2021) Tran, Myhoa; Yu, Yu; Wu, Hongli
    Purpose: Glutaredoxin 2 (Grx2) is a glutathione-dependent oxidoreductase that reduces S-glutathionylated proteins. Previously, we found that Grx2 could protect the retina from light-induced retinal degeneration (LIRD). However, the mechanisms that coordinate thiol-repair processes in the damaged retina remain unknown. To better understand the protective effects of Grx2, our study was extended to analyze the transcriptome changes of retinal tissue in light-exposed Grx2 knockout (KO) mice. Methods: Wild type (WT) and Grx2 KO mice were exposed to white light at 23,000 lux for 1 hour after dark adaptation for 10 hours. Retinal damage confirmed by electroretinogram (ERG) and spectral domain optical coherence tomography (SD-OCT). The transcriptome of the retinal tissue in WT and Grx2 KO mice were compared using transcriptome shotgun sequencing (RNA-seq). DESeq2 software utilized to analyze gene network. Real-time PCR and Western Blot further confirmed the genes of interest. Results: Light-exposed Grx2 KO mice showed compromised visual function indicated by loss of a- and b-wave amplitudes and thinning of the outer nuclear layer (ONL). Thousands of genes identified with statistically significant expression changes and were then classified into cellular processes and molecular pathways. Among these pathways, many genes that contribute to complement activation, inflammation, and cell survival system were significantly upregulated. Conclusions: Our results suggest that Grx2 could protect the retina from LIRD. Grx2 plays an important role in regulating light-induced retinal inflammation which may be associated with its ability to repair S-glutathionylated substrates.
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    Involvement of the c-Jun N-terminus kinase (JNK) pathway in Endothelin (ET-1) mediated neurodegeneration of retinal ganglion cells
    (2021) Kodati, Bindu; Stankowska, Dorota; Krishnamoorthy, Vignesh; Krishnamoorthy, Raghu
    PURPOSE: Endothelins contribute to neurodegeneration in glaucoma, however, the mechanisms are not completely understood. The goal of this study was to determine if JNK2 plays a causative role in endothelin-1 (ET-1)-mediated loss of RGCs in mice. METHODS: JNK2-/- and wild type (C57BL6) mice (n=4) were intravitreally injected in one eye with 2 nmoles of ET-1, while the contralateral eye was injected with 2 µl of vehicle. The mice were euthanized at 2 h and 24 h post-injection. Retinal sections from the JNK2-/- and wild type (C57BL6) mice were used for immunohistochemical analysis of the phosphorylation of JNK substrate, c-Jun. In a separate set of experiments, JNK2 -/- and wild type mice (n=6) were intravitreally injected with either 2 nmoles of ET-1 or vehicle, and euthanized 7 days post-injection. RGC counts and axonal degeneration were assessed. RESULTS: Intravitreal ET-1 administration produced a significant increase in immunostaining for phospho c-Jun in wild type mice which was appreciably lower in the JNK2 -/- mice. Following ET-1 administration, a significant (p< 0.05) 26% loss of RGCs was found in wild type mice, while JNK2-/- mice showed no significant (p=0.36) loss of RGCs. A significant decrease in the axonal counts and an increase in the collapsed axons was found in ET-1 injected eyes in wild type mice. CONCLUSION: JNK2 appears to play a major role in ET-1 mediated loss of RGCs in mice.
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    The Effects of TGFβ2 on Uveoscleral Outflow in the Mouse Eye
    (2021) Stevenson, Cooper; Clark, Abbot F.; Millar, J. Cameron
    Purpose: Elevated intraocular pressure (IOP) is a risk factor for primary open-angle glaucoma (POAG). Transforming growth factor beta-2 (TGFβ2) contributes to POAG pathogenesis via effects on the conventional aqueous outflow pathway. However, its effects on the uveoscleral (unconventional) aqueous outflow pathway, and effects of euthanasia and enucleation on TGFβ2 action, are unknown. Methods: We quantified the impact of overexpression of TGFβ2 on uveoscleral outflow (Fu) in mice. TGFβ2 was overexpressed in left (OS) eyes by intravitreal (IVT) injection with Ad5.CMV.hTGFβ2C226/228S (2×10-7pfu in 2?L). Right (OD) eyes were injected IVT with Ad5.CMV.null (same titer/volume). After 14 days IOP was elevated in OS (84.29%, p< 0.001). Fu was determined by perfusing the anterior aqueous chamber with FITC-dextran (2.5×10-3 M), followed by dissection of eyes into component tissues and measurement of each sample's fluorescence. Fu was inferred using a standard curve. Living animals, euthanatized animals, and enucleated eyes (exposed to air or submerged in PBS) from euthanatized animals were studied. Results: Fu was reduced in vivo in OS (0.0048±0.0017 µL/min) vs. OD (0.0987±0.0126 µL/min, p=0.025). In euthanatized mice, Fu was reduced in OS (0.0215±0.0101 µL/min) vs. OD (0.1543±0.0241 µL/min, p=0.010). In ex-vivo eyes exposed to air, Fu was reduced in OS (0.0702±0.0087 µL/min) vs. OD (0.1377±0.0106 µL/min, p=0.008). In ex-vivo eyes submerged in PBS, there was no difference between OS (0.0222±0.0065 µL/min) and OD (0.0137±0.0078 µL/min, p=0.175). Conclusions: This study quantifies TGFβ2 effects on uveoscleral outflow, providing insight on the effects TGFβ2 has on POAG pathophysiology.
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    miRNA Profiling of Human Optic Nerve Head Astrocytes Exposed to Cyclic Stretch
    (2021) Rangan, Rajiv; Tovar-Vidales, Tara
    Purpose: Elevated intraocular pressure (IOP) is the primary risk factor for glaucoma, a leading cause of irreversible blindness involving the progressive loss of retinal ganglion cells (RGCs) and their axons. Elevated IOP induces biomechanical aberrations within ocular tissues – including the transmission of biomechanical stretch through the lamina cribrosa (LC) region of the optic nerve head (ONH), the site where RGC axon damage first occurs. LC cells and ONH astrocytes (ONHA), the primary cells of the LC, respond to stretch in a manner that promotes pathological extracellular matrix (ECM) remodeling and mechanical damage of RGCs within the ONH. A complex set of molecular mechanisms regulate ECM remodeling. Part of this regulation may involve microRNAs (miRNAs), small RNA molecules that can indirectly inhibit gene expression by binding messenger RNA. miRNA dysregulation may contribute to ECM remodeling during glaucoma progression. In this study, we examined miRNA expression profiles of ONHA exposed to cyclic stretch. Methods: Primary human normal ONHA cell strains (n=3) were exposed to 0-12% cyclic stretch for 24 hours. miRNA PCR arrays were used to determine expression changes in profibrotic and anti-fibrotic miRNAs. Results: We found that specific miRNAs were consistently dysregulated across three independent strains of ONHA. Statistical significance could not be detected, but these patterns may represent biologically meaningful changes. Conclusion: Stretch modulates miRNA expression in cultured human ONHA and may be responsible for ECM alterations at the LC. Dysregulated miRNAs may serve as novel targets or models for future therapeutics.
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    Novel Compound SA-21 with Antioxidant Capability - The Prospect for Neuroprotection in Glaucoma.
    (2021) Ferguson, Jonathan; Johnson, Gretchen; Amankwa, Charles E.; Zhang, Wei; Li, Linya; Gondi, Sudershan; Funk, Arlene; Ellis, Dorette; Acharya, Suchismita; Stankowska, Dorota
    Purpose: Current treatments for glaucoma do not fully address neurodegeneration of retinal ganglion cells (RGCs). Our objective was to determine if compound SA-21, a hybrid superoxide dismutase and glutathione mimetic, could inhibit death of trabecular meshwork cells (NTM5), RGCs and rescue the functional decline of RGCs in an optic nerve crush (ONC) model. Methods: The structure of SA-21 was confirmed by magnetic resonance (NMR) spectroscopy and mass spectrometry. Reactive oxygen species (ROS) release was performed using pyrogallol assay. NTM5 cells were oxidatively stressed with TBHP (5.5mM) or vehicle in the presence of SA-21 (1mM, 100µM, 10µM, 1µM) for 24h. Cell survival was assessed by MTT assay. C57BL6 male mice (12-weeks old, n=4-5) were anesthetized, underwent ONC surgery, and at day 0 and 3 were intravitreally injected with 1% SA-21 (2µl) or vehicle. On day 7, pattern electroretinogram (PERG) was performed, animals were euthanized, and the number of surviving RBPMS-positive RGCs were counted. Results: SA-21 (1mM) treatment significantly decreased ROS production (~50%) measured by pyrogallol assay and increased NTM5 cell viability (~20%, p< 0.0094) following TBHP treatment compared to cells treated with the vehicle. ONC produced a 48% loss of RGCs, which was decreased in SA-21 treated mice (by ~10%) and demonstrated a trend in increase in PERG amplitude. Conclusions: SA-21 compound has antioxidant capability and protects NTM5 cells from oxidative stress. Intravitreally injected SA-21 at the selected dose in mice demonstrated trend in neuroprotection but further investigation is required.
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    Glucocorticoid-induced ocular hypertension alters synaptic plasticity and neurotransmission during the progression of glaucomatous neurodegeneration
    (2021) Maddineni, Prabhavathi; Kasetti, Ramesh; Zode, Gulab
    Purpose: Elevated intraocular pressure (IOP) is the major risk factor for Primary Open Angle Glaucoma (POAG). The neurodegeneration in POAG extends beyond the eye into the visual centers of the brain (VCB). Unfortunately, the underlying mechanisms responsible for IOP-induced neurodegeneration still remain unclear. We have developed a glucocorticoid (GC)-induced mouse model of glaucoma and determined the role of GC-induced ocular hypertension (OHT) on synaptic dysfunction, and how alterations in synaptic plasticity in VCB contribute to neurodegeneration. Methods: C57BL/6J mice were injected with either Dexamethasone (Dex) or Vehicle (Veh) via periocular-route, once a week for 10-weeks. IOP was measured every week and the electrical response of VCB was measured by VEP. Expression of synaptic markers in VCB were assessed by immunostaining. Results: Dex-induced OHT led to glaucomatous neurodegeneration in 10-weeks Dex treated mice compared to Veh mice. We observed RGC hyper excitability during the early stages of axonal damage with significantly increased VEP amplitudes with decreased latencies in 5-weeks Dex treated mice (32µV;73ms) compared to Veh mice (26µV;79ms). Interestingly, we observed complete collapse of neuronal excitability, with decreased VEP amplitudes and increased latencies in 10-weeks Dex treated mice (12µV;132ms) due to chronic OHT. Also, we observed an altered synaptic plasticity with decreased expression of both pre and post synaptic markers in the VCB of the 10-weeks Dex treated mouse. Conclusion: These data highlights that GC-induced OHT alters neurotransmission and axonal synaptic plasticity in VCB during the progression of glaucomatous neurodegeneration.