Browsing by Subject "eye"
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Item A Study of Some Aspects of the Role of Mast Cells in Experimental Autoimmune Uveitis(1994-06-01) Lee, Carol Hamberlin; Edward Orr; Robert Gracy; Laura S. LangLee, Carol Hamberlin, A Study of Some Aspects of the Role of Mast Cells in Experimental Autoimmune Uveitis. Doctor of Philosophy (Biomedical Sciences), June 1994, 141 pp., 6 tables, 29 illustrations, bibliography, 115 titles. Choroidal mast cells have been implicated in experimental autoimmune uveitis (EAU), an ocular inflammatory disease induced by S-antigen (Sag). Activation of ocular mast cells in Lewis rats was evaluated by determining changes in numbers of mast cells, levels of histamine, and wet weights of ocular tissues. A decrease in choroidal mast cells was confirmed statistically, and limbal mast cells were found to be activated earlier than choroidal mast cells. The ocular histamine distribution was altered during EAU, decreasing in the anterior eye, and increasing in the posterior eye. Retinal histamine levels increased when EAU symptoms occurred, but decreased while the disease was still intense. Levels of histamine methyltransferase, which degrades histamine, increased significiantly in retinal tissue when histamine levels fell. Signficant weight increases indicated edema, which can result from mast cell mediator action. Leflunomide, an immunomodulating drug that is known to affect mast cells in vitro, prevented induction of EAU. Leflunomide also suppressed changes in the mast cell-related parameters, histamine levels and wet weights. Mechanisms for activation of ocular mast cells in EAU were investigated. Results suggest that mast cell activation does not occur through mast cell surface IgE-antigen crosslinking. The adjuvant used, complete Freund’s adjuvant, is not conducive to IgE production. Histamine releasing factors, HRFs, are produced by various immune system cellular components. Preliminary efforts did not demonstrate HRF activity. Mast cell numbers, histamine levels, and wet weights were also evaluated in a milder form of EAU induced by M-peptide (Mpep), a peptide fragment of Sag. Mpep/EAU produces few disease symptoms in the anterior eye, but destroys the same retinal area as Sag/EAU—photoreceptor cells and their outer segments. Inflammation is less intense, restricted primarily to the target area. Mast cell numbers did not change, but histamine levels and wet weights changed significantly, suggesting that mast cells are also involved in Mpep/EAU. Overall, the results of this study add to evidence that mast cells are involved in pathogenesis of EAU. The results also point to topics of further investigation into the role of mast cells in EAU and in normal function in ocular tissues.Item Endothelin-1-Induced Proliferation of Human Optic Nerve Head Astrocytes Under Hypoxia(2003-11-01) Desai, Devashish; Thomas Yorio; Ganesh Prasanna; Clark, Abbot F.Desai, Devashish, Endothelin-1-Induced Proliferation of Cultured Human Optic Nerve Head Astrocytes under Hypoxia. Master of Science (Biomedical Sciences). Purpose: Optic nerve head astrocytes (ONAs) normally support and protect the axons of retinal ganglion cells exiting the eye. Along with effects related to elevated intraocular pressure (IOP), proliferation and activation of ONAs, known as ‘astrogliosis’, is also thought to contribute to the pathophysiology of glaucoma by distributing axonal transport and preventing axon regeneration. Concentrations of endothelin-1 (ET-1) are elevated in glaucomatous eyes and in animal models for glaucoma. ET-1 injection into the eye causes reduction of ocular blood flow. ET-1 causes a time-dependent proliferation of human ONAs. Tumor necrosis factor-α (TNF-α), a cytokine, which is also elevated in glaucomatous optic nerve head, promotes ET-1 release from ocular cells and could potentially stimulate ET-1 secretion from the ONAs. Hypoxia resulting from ischemia, which is produced by the elevation of IOP or vasospasm in the retinal vasculature, is considered a significant factor contributing to the stress as the glaucomatous optic nerve head. Methods: Concentrations of ET-1 secreted by hONAs into cell culture media after hypoxia and TNF-α treatment was measured using an enzyme-linked immunosorbent assay (ELISA). Proliferation of hONAs was measured using a proliferation assay (formazan assay), performed at the end of various time periods of incubation with TNPα and ET-1 under normoxia or hypoxia. The involvement of mitogen activated protein kinase (MAPK) in hONA proliferation was examined using MAPK inhibitors and Western blot analyses. Results: Cell culture media collected from hONAs after 24-hour hypoxia with concurrent TNF-α treatment showed a 500% increase in the irET-1. Under normoxia, both TNF-α and ET-1 caused moderate proliferation of hONAs. Under hypoxia, TNF-α-induced proliferation was greatly increased. Conclusion: Hypoxia augments TNF-a and ET-1 growth of optic nerve head astrocytes, by way of increasing ET-1 synthesis and release as well as mitogenesis. Therefore reactive ONAs could be the common denominator underlying optic nerve damage in glaucoma since their localization makes them susceptible to mechanistic and ischemic influences in addition to influences of ET-1 and TNF-α. Keywords: astrocyte; endothelin-1; tumor-necrosis factor-α; hypoxia; proliferation; astrogliosis; glaucoma; optic nerveItem Histamine Induced Changes in Phospholipase C Activity, Calcium Mobilization, and Contractility in Human Ciliary Muscle Cells(1996-06-01) Markwardt, Kerry L.; Michael W. Martin; Thomas Yorio; Eugene QuistMarkwardt, Kerry L., Histamine induced changes in phospholipase C activity, calcium mobilization, and contractility in human ciliary muscle cells. Doctor of Philosophy (Biomedical Sciences), June, 1996. Histamine has long been known to be an important mediator of inflammation and autocoid throughout the body. It has been shown to cause the contraction of many types of smooth muscle. Due to its known presence in many ocular structures and aqueous humor especially during inflammatory states, it was hypothesized that histamine could have an effect on intraocular pressure (IOP). This could occur if histamine triggered events which ultimately lead to contraction of the ciliary muscle, since it is established that contraction of the ciliary muscle affects aqueous humor outflow. Therefore, it was hypothesized in this study, the histamine causes increases in inositol phosphate production and intracellular calcium in human ciliary muscle cells which ultimately leads to contraction. To test this hypothesis, human ciliary muscle (CM) cells were cultured and used in various experiments to determine the effect of histamine on inositol phosphate production, intracellular calcium mobilization, and contractility. This study, for the first time in CM cells, showed that histamine, via an H1 receptor subtype, caused dose dependent increases in both inositol phosphates and intracellular calcium. Furthermore, it was shown that these histamine-induced events ultimately lead to contraction of the CM cells. Combining the results from all our studies, the data indicate that in human CM cells, histamine via an H1 receptor, activates phospholipase C which generates inositol phosphates such as inositol triphosphate (IP3). IP3 binds to an IP3 sensitive receptor on the endoplasmic reticulum causing the initial release of calcium which is sufficient to cause contraction of the CM cells. The intracellular release of calcium is also involved in activating a calcium channel which allows the influx of extracellular calcium into the cell. The results of these studies suggest that histamine could potentially have an IOP lowering effect in the eye due to contraction of the ciliary muscle. Overall, these studies contribute to a better understanding of the effect of histamine on a key IOP regulating tissue in the eye.Item Role of Glucocorticoids and Glucocorticoid Receptors in Glaucoma Pathogenesis(MDPI, 2023-10-27) Patel, Pinkal D.; Kodati, Bindu; Clark, Abbot F.The glucocorticoid receptor (GR), including both alternative spliced isoforms (GRalpha and GRbeta), has been implicated in the development of primary open-angle glaucoma (POAG) and iatrogenic glucocorticoid-induced glaucoma (GIG). POAG is the most common form of glaucoma, which is the leading cause of irreversible vision loss and blindness in the world. Glucocorticoids (GCs) are commonly used therapeutically for ocular and numerous other diseases/conditions. One serious side effect of prolonged GC therapy is the development of iatrogenic secondary ocular hypertension (OHT) and OAG (i.e., GC-induced glaucoma (GIG)) that clinically and pathologically mimics POAG. GC-induced OHT is caused by pathogenic damage to the trabecular meshwork (TM), a tissue involved in regulating aqueous humor outflow and intraocular pressure. TM cells derived from POAG eyes (GTM cells) have a lower expression of GRbeta, a dominant negative regulator of GC activity, compared to TM cells from age-matched control eyes. Therefore, GTM cells have a greater pathogenic response to GCs. Almost all POAG patients develop GC-OHT when treated with GCs, in contrast to a GC responder rate of 40% in the normal population. An increased expression of GRbeta can block GC-induced pathogenic changes in TM cells and reverse GC-OHT in mice. The endogenous expression of GRbeta in the TM may relate to differences in the development of GC-OHT in the normal population. A number of studies have suggested increased levels of endogenous cortisol in POAG patients as well as differences in cortisol metabolism, suggesting that GCs may be involved in the development of POAG. Additional studies are warranted to better understand the molecular mechanisms involved in POAG and GIG in order to develop new disease-modifying therapies to better treat these two sight threatening forms of glaucoma. The purpose of this timely review is to highlight the pathological and clinical features of GC-OHT and GIG, mechanisms responsible for GC responsiveness, potential therapeutic options, as well as to compare the similar features of GIG with POAG.Item Sigma-1 Receptor Signaling in the Eye(2008-03-12) Tchedre, Kissaou T.; Yorio, Thomas; Singh, Meharvan; Machu, TinaSIGMA-1 Receptor Signaling in the Eye Kissaou T. Tchedre, Department of Biomedical Sciences, University of North Texas Health Science Center Fort Worth, TX 76107. SUMMARY The sigma-1 receptor is a discovered transmembrane protein that mediates the regulation of ion channels. Sigma-1 receptor ligands have exhibited a wide variety of actions in the central nervous system including attenuation of the neuronal death associated with glutamate excitotoxicity both in vitro and in vivo. Although the sigma-1 receptor was cloned almost a decade ago, the molecular mechanism of the neuroprotective effect remains to be elucidated. In the current proposal it was hypothesized that activation of sigma-1 receptors promotes retinal ganglion cells survival by decreased calcium signaling pathways and factors linked to cell death. We showed by the ratiometric calcium imaging and patch clamp techniques that sigma-1 receptor activation could inhibit both calcium influx and intracellular calcium mobilization. The results showed that sigma-1 receptor overexpressing RGC-5 cells also had a lower glutamate-induced intracellular calcium mobilization compared to non-overexpressing RGC-5 cells. The survival assay data showed that the sigma-1 receptor agonist, (+)-SKF10047 protected RGC-5 cells showed a significant resistance to glutamate-induced apoptosis compared to the control RGC-5 cells. The sigma-1 receptor neuroprotective mechanism also included the down regulation of Bax, and caveolin-1 protein expression levels and inhibited caspase-3 activation. We also demonstrated for the first time using a co-immunoprecipitation technique, the association between L-type calcium channels and sigma-1 receptors. Thus sigma-1 receptor ligands may indirectly influence the voltage-gated calcium channels by interacting with the sigma-1 receptor associated voltage-gated calcium channel complex. In conclusion, activation of sigma-1 receptors can regulate calcium homeostasis and signaling in retinal ganglion cells. Activation of sigma-1 receptors regulate intracellular calcium levels and pro-apoptotic gene expression to promote retinal ganglion cell survival. Sigma-1 receptor ligands may be neuroprotective and targets for potential glaucoma therapeutics.Item The Effects of Ad5.CMV.hTGFβ2C226/228S on AHD in Mice(2021-05) Stevenson, Cooper H.; Millar, J. Cameron; Tovar-Vidales, Tara; Stankowska, Dorota L.Elevated intraocular pressure (IOP) is a key risk factor for the development of primary open-angle glaucoma (POAG), a leading cause of blindness in people over the age of 40 years. Transforming growth factor beta-2 is a cytokine known to contribute to the pathogenesis of POAG due to its deleterious effects on aqueous humor outflow via the conventional, or trabecular, outflow pathway in the eye. However, its effects on the rate of aqueous outflow (Fu) via the unconventional or uveoscleral outflow pathway, rate of aqueous humor production (Fin), and episcleral venous pressure (Pe) are unknown. Further, effects of euthanasia and enucleation in our hands on TGFβ2-mediated effects on Fu are also unknown. The goal of the present study was to quantify the impact of over-expression of TGFβ2 on aqueous humor dynamics (AHD) in the mouse eye, with special emphasis on Fu, Fin, and Pe in the mouse eye. To simulate TGFβ2 over-expression, left (OS) eyes were injected intravitreally (IVT) with a mutant form of TGFβ2 (Ad5.CMV.hTGFβ2C226/228S, 2×10⁷pfu in 2μL), while right (OD) eyes were injected IVT with a null virus (Ad5.CMV.null, same titer and volume). Following 14 days, after which time mean IOP (determined tonometrically in conscious mice) had become elevated in TGFβ2-injected eyes (84.29% increase in IOP, P < 0.001), Fu was determined directly by cannulating the anterior aqueous chamber (AC) and perfusing it with fluorescein isothiocyanate-dextran (1×10⁻⁹ M), followed by dissection of the retina/choroid/iris-ciliary body/scleral shell, homogenization, and measurement of each sample's fluorescence, and then inference of flow rate using a standard curve. Those perfusion were performed in living eyes, also in eyes in situ in the animal immediately following euthanasia, and enucleated eyes perfused in vivo either (i) exposed to air, or (ii) submerged in PBS. In a further group of experiments in living animals aqueous humor outflow conductance (C) (also known as aqueous humor outflow facility), and Pe were measured, and then Fin and Fu were calculated using a constant flow infusion method. Further, we sought to determine whether IOP elevation would lead to a reduction in RGC numbers in the retina, so retinal flat mounts from both treated and untreated eyes from 5 of our animals were prepared and RGC counts were made. For eyes perfused in-vivo, Fu was reduced in OS (0.0048 ± 0.0017 μL/min) compared to OD (0.0987 ± 0.0126 μL/min, P = 0.025). For eyes perfused in euthanatized mice, Fu was reduced in OS (0.0215 ± 0.0101 μL/min) compared to OD (0.1543 ± 0.0241 μL/min, P = 0.010). For eyes perfused ex-vivo while submerged in PBS, there was no difference in Fu between OS (0.0222 ± 0.0065 μL/min) and OD (0.0137 ± 0.0078 μL/min, P = 0.175). For eyes perfused ex-vivo while exposed to air, Fu was reduced in OS (0.0702 ± 0.0087 μL/min) compared to OD (0.1377 ± 0.0106 μL/min, P = 0.008). Fin showed a trend towards a reduction in the eyes in which TGFβ2 was over-expressed, but this effect did not reach statistical significance. There was a significant increase in Pe in eyes in which TGFβ2 was expressed (8.6 ± 0.7 mmHg in OS to 6.4 ± 0.2 mmHg in OD, P = 0.015). Given these results, the present study further quantifies the effect of TGFβ2 in POAG, providing more insight into its mechanism of action in this disease.