Browsing by Subject "Intraocular Pressure / drug effects"
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Item Effect of ATP-sensitive Potassium Channel Openers on Intraocular Pressure in Ocular Hypertensive Animal Models(ARVO Journals, 2022-02-01) Roy Chowdhury, Uttio; Millar, J. Cameron; Holman, Bradley H.; Anderson, Kjerston J.; Dosa, Peter I.; Roddy, Gavin W.; Fautsch, Michael P.Purpose: To evaluate the effect of ATP-sensitive potassium channel openers cromakalim prodrug 1 (CKLP1) and diazoxide on IOP in three independent mouse models of ocular hypertension. Methods: Baseline IOP was measured in TGFbeta2 overexpression, steroid-induced, and iris dispersion (DBA/2J) ocular hypertension mouse models, followed by once daily eyedrop administration with CKLP1 (5 mM) or diazoxide (5 mM). The IOP was measured in conscious animals with a handheld rebound tonometer. Aqueous humor dynamics were assessed by a constant perfusion method. Effect of treatment on ocular tissues was evaluated by transmission electron microscopy. Results: CKLP1 decreased the IOP by 20% in TGFbeta2 overexpressing mice (n = 6; P < 0.0001), 24% in steroid-induced ocular hypertensive mice (n = 8; P < 0.0001), and 43% in DBA/2J mice (n = 15; P < 0.0001). Diazoxide decreased the IOP by 32% in mice with steroid-induced ocular hypertension (n = 13; P < 0.0001) and by 41% in DBA/2J mice (n = 4; P = 0.005). An analysis of the aqueous humor dynamics revealed that CKLP1 decreased the episcleral venous pressure by 29% in TGFbeta2 overexpressing mice (n = 13; P < 0.0001) and by 72% in DBA/2J mice (n = 4 control, 3 treated; P = 0.0002). Diazoxide lowered episcleral venous pressure by 35% in steroid-induced ocular hypertensive mice (n = 3; P = 0.03). Tissue histology and cell morphology appeared normal when compared with controls. Accumulation of extracellular matrix was reduced in CKLP1- and diazoxide-treated eyes in the steroid-induced ocular hypertension model. Conclusions: ATP-sensitive potassium channel openers CKLP1 and diazoxide effectively decreased the IOP in ocular hypertensive animal models by decreasing the episcleral venous pressure, supporting a potential therapeutic application of these agents in ocular hypertension and glaucoma.Item Effect of Cromakalim Prodrug 1 (CKLP1) on Aqueous Humor Dynamics and Feasibility of Combination Therapy With Existing Ocular Hypotensive Agents(ARVO Journals, 2017-11-01) Roy Chowdhury, Uttio; Rinkoski, Tommy A.; Bahler, Cindy K.; Millar, J. Cameron; Bertrand, Jacques A.; Holman, Bradley H.; Sherwood, Joseph M.; Overby, Darryl R.; Stoltz, Kristen L.; Dosa, Peter I.; Fautsch, Michael P.Purpose: Cromakalim prodrug 1 (CKLP1) is a water-soluble ATP-sensitive potassium channel opener that has shown ocular hypotensive properties in ex vivo and in vivo experimental models. To determine its mechanism of action, we assessed the effect of CKLP1 on aqueous humor dynamics and in combination therapy with existing ocular hypotensive agents. Methods: Outflow facility was assessed in C57BL/6 mice by ex vivo eye perfusions and by in vivo constant flow infusion following CKLP1 treatment. Human anterior segments with no trabecular meshwork were evaluated for effect on pressure following CKLP1 treatment. CKLP1 alone and in combination with latanoprost, timolol, and Rho kinase inhibitor Y27632 were evaluated for effect on intraocular pressure in C57BL/6 mice and Dutch-belted pigmented rabbits. Results: CKLP1 lowered episcleral venous pressure (control: 8.9 +/- 0.1 mm Hg versus treated: 6.2 +/- 0.1 mm Hg, P < 0.0001) but had no detectable effect on outflow facility, aqueous humor flow rate, or uveoscleral outflow. Treatment with CKLP1 in human anterior segments without the trabecular meshwork resulted in a 50% +/- 9% decrease in pressure, suggesting an effect on the distal portion of the conventional outflow pathway. CKLP1 worked additively with latanoprost, timolol, and Y27632 to lower IOP, presumably owing to combined effects on different aspects of aqueous humor dynamics. Conclusions: CKLP1 lowered intraocular pressure by reducing episcleral venous pressure and lowering distal outflow resistance in the conventional outflow pathway. Owing to this unique mechanism of action, CKLP1 works in an additive manner to lower intraocular pressure with latanoprost, timolol, and Rho kinase inhibitor Y27632.Item Glucocorticoid receptor GRbeta regulates glucocorticoid-induced ocular hypertension in mice(Springer Nature, 2018-01-16) Patel, Gaurang C.; Liu, Yang; Millar, J. Cameron; Clark, Abbot F.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 GRbeta acts as dominant negative regulator of GR activity, and it has been shown that overexpressing GRbeta 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 GRbeta isoform in the TM of mouse eyes treated with GCs, to precisely dissect the role of GRbeta in regulating steroid responsiveness. We show that overexpression of GRbeta inhibits GC effects on MTM cells in vitro and GC-induced OHT in mouse eyes in vivo. Ad5 mediated GRbeta overexpression reduced the GC induction of fibronectin, collagen 1, and myocilin in TM of mouse eyes both in vitro and in vivo. GRbeta also reversed DEX-Ac induced IOP elevation, which correlated with increased conventional aqueous humor outflow facility. Thus, GRbeta 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 GRbeta in regulating GC-OHT and GC-mediated gene expression in the TM.Item ID1 and ID3 are Negative Regulators of TGFbeta2-Induced Ocular Hypertension and Compromised Aqueous Humor Outflow Facility in Mice(ARVO Journals, 2021-05-03) Mody, Avani A.; Millar, J. Cameron; Clark, Abbot F.Purpose: In POAG, elevated IOP remains the major risk factor in irreversible vision loss. Increased TGFbeta2 expression in POAG aqueous humor and in the trabecular meshwork (TM) amplifies extracellular matrix (ECM) deposition and reduces ECM turnover in the TM, leading to a decreased aqueous humor (AH) outflow facility and increased IOP. Inhibitor of DNA binding proteins (ID1 and ID3) inhibit TGFbeta2-induced fibronectin and PAI-1 production in TM cells. We examined the effects of ID1 and ID3 gene expression on TGFbeta2-induced ocular hypertension and decreased AH outflow facility in living mouse eyes. Methods: IOP and AH outflow facility changes were determined using a mouse model of Ad5-hTGFbeta2C226S/C288S-induced ocular hypertension. The physiological function of ID1 and ID3 genes were evaluated using Ad5 viral vectors to enhance or knockdown ID1/ID3 gene expression in the TM of BALB/cJ mice. IOP was measured in conscious mice using a Tonolab impact tonometer. AH outflow facilities were determined by constant flow infusion in live mice. Results: Over-expressing ID1 and ID3 significantly blocked TGFbeta2-induced ocular hypertension (P < 0.0001). Although AH outflow facility was significantly decreased in TGFbeta2-transduced eyes (P < 0.04), normal outflow facility was preserved in eyes injected concurrently with ID1 or ID3 along with TGFbeta2. Knockdown of ID1 or ID3 expression exacerbated TGFbeta2-induced ocular hypertension. Conclusions: Increased expression of ID1 and ID3 suppressed both TGFbeta2-elevated IOP and decreased AH outflow facility. ID1 and/or ID3 proteins thus may show promise as future candidates as IOP-lowering targets in POAG.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.