Ultrastructural and Biochemical Evaluation of the Porcine Anterior Chamber Perfusion Model

Bachmann, Björn; Birke, Marco T.; Kook, Daniel; Eichhorn, Michael; Lütjen–Drecoll, Elke

doi:10.1167/iovs.05-1393

Cited by 69 publications

(61 citation statements)

References 20 publications

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“…Porcine eyes are a good model for human eyes 28,29 as a model for studying glaucoma, 15,28,30,31 aqueous humor outflow physiology, [31][32][33] and chronic IOP elevation, 28,29 and similar to human, outflow facility in both is reduced in response to TGFb 2 . 32 Porcine TM cells are already widely used by glaucoma researchers as a surrogate for human TM cells.…”

Section: Discussionmentioning

confidence: 99%

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Oxidative Stress Impact on Barrier Function of Porcine Angular Aqueous Plexus Cell Monolayers

Lei

Stamer

et al. 2013

Invest. Ophthalmol. Vis. Sci.

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Citation: Lei Y, Stamer WD, Wu J, Sun X. Oxidative stress impact on barrier function of porcine angular aqueous plexus cell monolayers. Invest Ophthalmol Vis Sci. 2013;54:4827-4835. DOI:10.1167/iovs.12-11435 PURPOSE. Our goal was to investigate the effect of chronic oxidative stress on angular aqueous plexus (AAP, functional equivalent to human Schlemm's canal) endothelial cells from porcine eyes.METHODS. AAP cells were differentially isolated from porcine outflow tissues using puromycin selection. Confluent cultures of porcine AAP cells were grown for 2 weeks in physiological (5% O 2 ) or hyperoxic conditions (40% O 2 ) to model elevated oxidative stress associated with ageing. Cell growth rate, size, transendothelial electrical resistance (TEER), and hydraulic conductivity (HC) were measured. The expression of senescence-associated b-galactosidase and DNA damage marker 8-hydroxy-2 0 -deoxyguanosine (8-OHdG) was monitored, and the levels of cytoskeletal and cell-cell adhesion proteins such as F-actin, phospho-myosin light chain (phosphor-MLC), occludin, claudin-5, ZO-1, b-catenin, and VE-cadherin were measured by immunofluorescence staining and Western blot analysis.RESULTS. Data showed that chronic hyperoxia inhibited cell growth rate from day 3 onward, the cell size increased by 18.2% 6 5.1%, and cells stained positive for bgalactosidase and 8-OHdG. Hyperoxia resulted in a significant 30% increase in TEER compared with the control group (P < 0.05, n ¼ 6). When perfused in the basal-to-apical direction at 4 mm Hg, HC of AAP cells was 1.97 6 0.12 and 1.54 6 0.13 lL/mm Hg/min/ cm 2 in control and hyperoxia groups, respectively (P < 0.05, n ¼ 6). Stressed cells expressed a significantly greater abundance of F-actin, phospho-MLC, occludin, claudin-5, b-catenin, and VE-cadherin compared to the control group by both immunofluorescence and Western blot analyses.CONCLUSIONS. Chronic exposure of AAP cells to oxidative stress decreased cell monolayer permeability and up-regulated cytoskeletal and cell-cell adhesion protein expression; suggesting that, with age and increased oxidative stress, resistance at the level of Schlemm's canal increases.

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Section: Discussionmentioning

confidence: 99%

“…32 Porcine TM cells are already widely used by glaucoma researchers as a surrogate for human TM cells. In this study we employed an established method of isolating AAP cells published by our group previously.…”

Section: Discussionmentioning

confidence: 99%

Oxidative Stress Impact on Barrier Function of Porcine Angular Aqueous Plexus Cell Monolayers

Lei

Stamer

et al. 2013

Invest. Ophthalmol. Vis. Sci.

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“…It is found abundantly in TM beams and the JCT region, with slightly higher levels in the basement membranes of TM beam cells, JCT cells and SC inner wall cells (Keller et al, manuscript in review). (Floyd et al 1985;Murphy et al 1987;Zhou et al 1998a;Bachmann et al 2006;Keller et al 2007a) In the JCT at the ultrastructural level, fibronectin is found in the amorphous material associated with the basement membranes and with the peripheral SD material of the elastic microfibrils. (Yue 1996;Brilakis et al 2001;Ueda et al 2002;Ueda and Yue 2003) In addition to forming a thin fibrous array that appears to encircle TM cells in culture, (Zhou et al 1998b) fibronectin also forms thick fibrous strands.…”

Section: Fibronectin and Vitronectinmentioning

confidence: 99%

Extracellular matrix in the trabecular meshwork

Acott

Kelley

2008

Experimental Eye Research

417

443

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The extracellular matrix (ECM) of the trabecular meshwork (TM) is thought to be important in regulating intraocular pressure (IOP) in both normal and glaucomatous eyes. IOP is regulated primarily by a fluid resistance to aqueous humor outflow. However, neither the exact site nor the identity of the normal resistance to aqueous humor outflow has been established. Whether the site and nature of the increased outflow resistance, which is associated with open-angle glaucoma, is the same or different from the normal resistance is also unclear.The ECMs of the TM beams, juxtacanalicular region (JCT) and Schlemm's canal (SC) inner wall are comprised of fibrillar and non-fibrillar collagens, elastin-containing microfibrils, matricellular and structural organizing proteins, glycosaminoglycans (GAGs) and proteoglycans. Both basement membranes and stromal ECM are present in the TM beams and JCT region. Cell adhesion proteins, cell surface ECM receptors and associated binding proteins are also present in the beams, JCT and SC inner wall region.The outflow pathway ECM is relatively dynamic, undergoing constant turnover and remodeling. Regulated changes in enzymes responsible for ECM degradation and biosynthetic replacement are observed. IOP homeostasis, triggered by pressure changes or mechanical stretching of the TM, appears to involve ECM turnover. Several cytokines, growth factors and drugs, which affect the outflow resistance, change ECM component expression, mRNA alternative splicing, cellular cytoskeletal organization or all of these. Changes in ECM associated with open-angle glaucoma have been identified.

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“…The anterior segment of the porcine eye has an acceptable degree of anatomical and ultrastructural resemblance with the human eye (Bachmann et al 2006), and isolated porcine eyes (WikbergMatsson et al 2000;Wikberg-Matsson & Simonsen 2001) or isolated porcine iris muscles (Toda et al 1999) are established models for investigating ocular adrenoceptors (Toda et al 1999;Wikberg-Matsson et al 2000;Wikberg-Matsson & Simonsen 2001). In addition, for an intracameral mydriatic to be usable in humans safety issues, possible adverse reactions et cetera also have to be addressed.…”

Section: Discussionmentioning

confidence: 99%

Stimulation of adrenergic β‐receptors enhances mydriasis in a porcine eye model

Janbaz

Lundberg

Behndig

2011

Acta Ophthalmologica

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Abstract.Purpose: To compare the mydriatic effect of intracamerally injected isoprenaline plus phenylephrine to phenylephrine alone and to epinephrine in a porcine eye model, aiming to eventually find the best combination of adrenergic substances for surgical mydriasis in humans.Methods: In this study, we used 89 intact eyes from newly slaughtered pigs, pretreated with 2.0 mg of intracameral acetylcholine. After waiting 60 seconds for miosis to develop, 0.15 ml 0.3% isoprenaline and 0.15 ml 3.0% phenylephrine were injected sequentially with a 90‐second interval in 21 eyes. In another 22 eyes, the same substances were given in the reverse order. In 20 eyes, 0.15 ml of 0.025% epinephrine was injected, and as a negative control 0.15 ml of balanced salt solution was injected in 26 eyes. The pupils were filmed during the treatments, and the mean pupil diameters were measured every 15 seconds from the video recordings.Results: Phenylephrine injected after isoprenaline had a larger mydriatic effect than epinephrine (p < 0.01). Without isoprenaline pretreatment, the mydriatic effect of phenylephrine was significantly smaller than that of epinephrine (p < 0.05). Isoprenaline also exhibited a small mydriatic effect of its own.Conclusions: The β‐receptor stimulator isoprenaline enhances the mydriatic effect of intracameral phenylephrine, indicating a role for the β‐receptor in the mydriatic response. Mydriasis mediated by β‐receptors may explain why nonspecific adrenergic stimulators such as epinine and epinephrine can have larger mydriatic effects than the specific α1‐receptor stimulator phenylephrine.

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Ultrastructural and Biochemical Evaluation of the Porcine Anterior Chamber Perfusion Model

Cited by 69 publications

References 20 publications

Oxidative Stress Impact on Barrier Function of Porcine Angular Aqueous Plexus Cell Monolayers

Oxidative Stress Impact on Barrier Function of Porcine Angular Aqueous Plexus Cell Monolayers

Extracellular matrix in the trabecular meshwork

Stimulation of adrenergic β‐receptors enhances mydriasis in a porcine eye model

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