Publications
75 results found
Bertrand JA, Sherwood JM, Schicht M, et al., 2019, Blockade of the BK-<i>a</i>/β4 potassium ion channel reduces outflow facility in mice, Annual Meeting of the Association-for-Research-in-Vision-and-Ophthalmology (ARVO), Publisher: ASSOC RESEARCH VISION OPHTHALMOLOGY INC, ISSN: 0146-0404
Torres ER, Sherwood JM, Overby DR, 2019, Aqueous humour outflow requires active cellular metabolism, Annual Meeting of the Association-for-Research-in-Vision-and-Ophthalmology (ARVO), Publisher: ASSOC RESEARCH VISION OPHTHALMOLOGY INC, ISSN: 0146-0404
McDonnell F, Perkumas KM, Ashpole NE, et al., 2019, Elevated IOP increases Shear Stress in Schlemm's Canal, Annual Meeting of the Association-for-Research-in-Vision-and-Ophthalmology (ARVO), Publisher: ASSOC RESEARCH VISION OPHTHALMOLOGY INC, ISSN: 0146-0404
Overby DR, Spenlehauer A, Cairoli A, et al., 2019, Actomyosin contractility and the vimentin cytoskeleton influence giant vacuole life-cycle in Schlemm's canal endothelial cells, Annual Meeting of the Association-for-Research-in-Vision-and-Ophthalmology (ARVO), Publisher: ASSOC RESEARCH VISION OPHTHALMOLOGY INC, ISSN: 0146-0404
Madekurozwa M, 2019, Aqueous Humour Outflow Dynamics in Mice
Glaucoma is the leading cause of irreversible blindness worldwide. The major risk factor of glaucoma is sustained elevation of intraocular pressure (IOP), and lowering IOP is the only proven method for halting the progression of glaucomatous blindness. IOP is determined by the balance between aqueous humour (AH) production and drainage through pressure-dependent and pressure-independent outflow pathways. Elevated IOP is caused by increased hydraulic resistance through the pressure-dependent outflow pathway. Most glaucoma therapies aimed at lowering IOP do not effectively target pressure-dependent outflow due to an incomplete understanding of its regulation. We aim to use mice to study outflow regulation in the context of glaucoma.Mice are commonly used to study IOP regulation due to their resemblance to human ocular anatomy, genetics and pharmacology. However, while the bulk of AH drainage passes through the pressure-dependent pathway in humans, it has been reported to predominantly flow through the pressure-independent pathway in mice, which if true would invalidate the mouse as a model for studying outflow as occurs in humans. Here we present the first direct measurement of pressure-independent outflow in mice, showing it to be indistinguishable from zero which supports the mouse being a good model for pressure-dependent outflow as occurs in humans.We also investigated the role of the ocular pulse in outflow facility regulation, which arises due to cardiac pulsations in ocular blood volume. To do this we designed an apparatus to apply a sinusoidal pressure waveform superimposed onto a steady pressure whilst simultaneously measuring outflow resistance. We show that the ocular pulse leads to immediate decrease in outflow resistance in mice, and the effect was partly mediated through nitric oxide synthase.Finally, we developed a new apparatus and method to measure outflow resistance in living mice accounting for the influence of anaesthesia that introduces time-depen
Sherwood JM, Stamer WD, Overby DR, 2019, A model of the oscillatory mechanical forces in the conventional outflow pathway, Journal of the Royal Society Interface, Vol: 16, ISSN: 1742-5662
Intraocular pressure is regulated by mechanosensitive cells within the conventional outflow pathway, the primary route of aqueous humour drainage from the eye. However, the characteristics of the forces acting on those cells are poorly understood. We develop a model that describes flow through the conventional outflow pathway, including the trabecular meshwork (TM) and Schlemm’s canal (SC). Accounting for the ocular pulse, we estimate the time-varying shear stress on SC endothelium and strain on the TM. We consider a range of outflow resistances spanning normotensive to hypertensive conditions. Over this range, the SC shear stress increases significantly and becomes highly oscillatory. TM strain also increases, but with negligible oscillations. Interestingly, TM strain responds more to changes in outflow resistance around physiological values, while SC shear stress responds more to elevated levels of resistance. A modest increase in TM stiffness, as observed in glaucoma, suppresses TM strain and practically eliminates the influence of outflow resistance on SC shear stress. As SC and TM cells respond to mechanical stimulation by secreting factors that modulate outflow resistance, our model provides insight regarding the potential role of SC shear and TM strain as mechanosensory cues for homeostatic regulation of outflow resistance and hence intraocular pressure.
Li G, Lee C, Agrahari V, et al., 2019, In vivo measurement of trabecular meshwork stiffness in a corticosteroid-induced ocular hypertensive mouse model, Proceedings of the National Academy of Sciences of the United States of America, Vol: 116, Pages: 1714-1722, ISSN: 0027-8424
Ocular corticosteroids are commonly used clinically. Unfortunately, their administration frequently leads to ocular hypertension, i.e., elevated intraocular pressure (IOP), which, in turn, can progress to a form of glaucoma known as steroid-induced glaucoma. The pathophysiology of this condition is poorly understood yet shares similarities with the most common form of glaucoma. Using nanotechnology, we created a mouse model of corticosteroid-induced ocular hypertension. This model functionally and morphologically resembles human ocular hypertension, having titratable, robust, and sustained IOPs caused by increased resistance to aqueous humor outflow. Using this model, we then interrogated the biomechanical properties of the trabecular meshwork (TM), including the inner wall of Schlemm’s canal (SC), tissues known to strongly influence IOP and to be altered in other forms of glaucoma. Specifically, using spectral domain optical coherence tomography, we observed that SC in corticosteroid-treated mice was more resistant to collapse at elevated IOPs, reflecting increased TM stiffness determined by inverse finite element modeling. Our noninvasive approach to monitoring TM stiffness in vivo is applicable to other forms of glaucoma and has significant potential to monitor TM function and thus positively affect the clinical care of glaucoma, the leading cause of irreversible blindness worldwide.
Wu P-J, Kabovka I, Ruberti J, et al., 2018, Water content, not stiffness, dominates Brillouin spectroscopy measurements in hydrated materials, Nature Methods, Vol: 15, Pages: 561-562, ISSN: 1548-7091
O'Callaghan J, Cassidy PS, Reina-Torres E, et al., 2018, INDUCIBLE MMP-3 EXPRESSION IN A STEROID-INDUCED MURINE MODEL OF GLAUCOMA, Annual Meeting of the Association-for-Research-in-Vision-and-Ophthalmology (ARVO), Publisher: ASSOC RESEARCH VISION OPHTHALMOLOGY INC, ISSN: 0146-0404
Overby DR, Madekurozwa M, Stamer WD, et al., 2018, Modulation of outflow facility by the ocular pulse, Annual Meeting of the Association-for-Research-in-Vision-and-Ophthalmology (ARVO), Publisher: ASSOC RESEARCH VISION OPHTHALMOLOGY INC, ISSN: 0146-0404
Feola A, Sherwood J, Overby DR, et al., 2018, Impact of Estrogen Deficiency on Outflow Facility and Ocular Biomechanics in Rats, Annual Meeting of the Association-for-Research-in-Vision-and-Ophthalmology (ARVO), Publisher: ASSOC RESEARCH VISION OPHTHALMOLOGY INC, ISSN: 0146-0404
Kaliviotis E, Sherwood JM, Balabani S, 2018, Local viscosity distribution in bifurcating microfluidic blood flows, Physics of Fluids, Vol: 30, ISSN: 1070-6631
The red blood cell (RBC) aggregation phenomenon is majorly responsible for the non-Newtonian nature of blood, influencing the blood flow characteristics in the microvasculature. Of considerable interest is the behaviour of the fluid at the bifurcating regions. In vitro experiments, using microchannels, have shown that RBC aggregation, at certain flow conditions, affects the bluntness and skewness of the velocity profile, the local RBC concentration, and the cell-depleted layer at the channel walls. In addition, the developed RBC aggregates appear unevenly distributed in the outlets of these channels depending on their spatial distribution in the feeding branch, and on the flow conditions in the outlet branches. In the present work, constitutive equations of blood viscosity, from earlier work of the authors, are applied to flows in a T-type bifurcating microchannel to examine the local viscosity characteristics. Viscosity maps are derived for various flow distributions in the outlet branches of the channel, and the location of maximum viscosity magnitude is obtained. The viscosity does not appear significantly elevated in the branches of lower flow rate as would be expected on the basis of the low shear therein, and the maximum magnitude appears in the vicinity of the junction, and towards the side of the outlet branch with the higher flow rate. The study demonstrates that in the branches of lower flow rate, the local viscosity is also low, helping us to explain why the effects of physiological red blood cell aggregation have no adverse effects in terms of in vivo vascular resistance.
arora H, nila A, Vitharana K, et al., 2017, Microstructural consequences of blast lung injury characterised with digital volume correlation, Frontiers in Materials, Vol: 4, ISSN: 2296-8016
This study focuses on microstructural changes that occur within the mammalian lung when subject to blast and how these changes influence strain distributions within the tissue. Shock tube experiments were performed to generate the blast injured specimens (cadaveric Sprague-Dawley rats). Blast overpressures of 100 and 180 kPa were studied. Synchrotron tomography imaging was used to capture volumetric image data of lungs. Specimens were ventilated using a custom-built system to study multiple inflation pressures during each tomography scan. These data enabled the first digital volume correlation (DVC) measurements in lung tissue to be performed. Quantitative analysis was performed to describe the damaged architecture of the lung. No clear changes in the microstructure of the tissue morphology were observed due to controlled low- to moderate-level blast exposure. However, significant focal sites of injury were observed using DVC, which allowed the detection of bias and concentration in the patterns of strain level. Morphological analysis corroborated the findings, illustrating that the focal damage caused by a blast can give rise to diffuse influence across the tissue. It is important to characterize the non-instantly fatal doses of blast, given the transient nature of blast lung in the clinical setting. This research has highlighted the need for better understanding of focal injury and its zone of influence (alveolar interdependency and neighboring tissue burden as a result of focal injury). DVC techniques show great promise as a tool to advance this endeavor, providing a new perspective on lung mechanics after blast.
Campbell IC, Sherwood JM, Overby DR, et al., 2017, Quantification of Scleral Biomechanics and Collagen Fiber Alignment., Glaucoma. Methods in Molecular Biology., Editors: Jakobs, Publisher: Humana Press, Pages: 135-159
The stiffness of the sclera is important in several ocular disorders, and there is hence a need to quantify the biomechanical properties of this tissue. Here, we present two methods for measuring the stiffness of scleral ocular tissues: ocular compliance testing and digital image correlation strain mapping. In tandem with these approaches, we provide two methods to spatially quantify the anisotropic alignment of collagen fibers making up the sclera, using second harmonic generation microscopy and small-angle light scattering. Together, these approaches allow specimen-specific measurement of tissue stiffness and collagen alignment, which are key factors in determining how the eye responds to mechanical loads.
Wu P-J, Kabakova I, Ruberti J, et al., 2017, Brillouin microscopy, what is it really measuring?
Brillouin microscopy measures compressibility, but is being increasingly usedto assess stiffness of cells and tissues. Using hydrogels with tunableproperties, we demonstrate that Brillouin microscopy is insensitive tostiffness of hydrated materials, but depends strongly on water content,consistent with a theoretical model of biphasic compressibility. Empiricalcorrelations between Brillouin measurements and stiffness arise due to theirmutual dependence on water content, but correlations vanish once hydration iscontrolled.
Roy Chowdhury U, Rinkoski TA, Bahler CK, et al., 2017, Effect of Cromakalim Prodrug 1 (CKLP1) on Aqueous Humor Dynamics and Feasibility of Combination Therapy With Existing Ocular Hypotensive Agents., Investigative Ophthalmology & Visual Science, Vol: 58, Pages: 5731-5742, ISSN: 1552-5783
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.
Madekurozwa M, Reina-Torres E, Overby DR, et al., 2017, Direct measurement of pressure-independent aqueous humour flow using iPerfusion, Experimental Eye Research, Vol: 162, Pages: 129-138, ISSN: 0014-4835
Reduction of intraocular pressure is the sole therapeutic target for glaucoma. Intraocular pressure is determined by the dynamics of aqueous humour secretion and outflow, which comprise several pressure-dependent and pressure-independent mechanisms. Accurately quantifying the components of aqueous humour dynamics is essential in understanding the pathology of glaucoma and the development of new treatments. To better characterise aqueous humour dynamics, we propose a method to directly measure pressure-independent aqueous humour flow. Using the iPerfusion system, we directly measure the flow into the eye when the pressure drop across the pressure-dependent pathways is eliminated. Using this approach we address i) the magnitude of pressure-independent flow in ex vivo eyes, ii) whether we can accurately measure an artificially imposed pressure-independent flow, and iii) whether the presence of a pressure-independent flow affects our ability to measure outflow facility. These studies are conducted in mice, which are a common animal model for aqueous humour dynamics. In eyes perfused with a single cannula, the average pressure-independent flow was 1 [-3, 5] nl/min (mean [95% confidence interval]) (N = 6). Paired ex vivo eyes were then cannulated with two needles, connecting the eye to both iPerfusion and a syringe pump, which was used to impose a known pressure-independent flow of 120 nl/min into the experimental eye only. The measured pressure-independent flow was then 121 [117, 125] nl/min (N = 7), indicating that the method could measure pressure-independent flow with high accuracy. Finally, we showed that the artificially imposed pressure-independent flow did not affect our ability to measure facility, provided that the pressure-dependence of facility and the true pressure-independent flow were accounted for. The present study provides a robust method for measurement of pressure-independent flow, and demonstrates the importance of accurately quantifying this paramete
Sherwood JM, Starner WD, Overby DR, 2017, Factors affecting oscillatory shear stress in Schlemm's canal, Annual Meeting of the Association-for-Research-in-Vision-and-Ophthalmology (ARVO) - Imaging in the Eye, Publisher: ASSOC RESEARCH VISION OPHTHALMOLOGY INC, ISSN: 0146-0404
Fu J, Allen RS, Campbell I, et al., 2017, Menopause alters Ocular Biomechanics and Increases Visual Impairment in a Rat Model of Glaucoma, Annual Meeting of the Association-for-Research-in-Vision-and-Ophthalmology (ARVO), Publisher: ASSOC RESEARCH VISION OPHTHALMOLOGY INC, ISSN: 0146-0404
Madekurozwa M, Feola A, Ethier CR, et al., 2017, Comparison of Pressure-Dependent Facility in Rodent Eyes, Annual Meeting of the Association-for-Research-in-Vision-and-Ophthalmology (ARVO) - Imaging in the Eye, Publisher: ASSOC RESEARCH VISION OPHTHALMOLOGY INC, ISSN: 0146-0404
Kaliviotis E, Pasias D, Sherwood JM, et al., 2017, Red blood cell aggregate flux in a bifurcating microchannel., Medical Engineering and Physics, Vol: 48, Pages: 23-30, ISSN: 1350-4533
Red blood cell aggregation plays a key role in microcirculatory flows, however, little is known about the transport characteristics of red blood cell aggregates in branching geometries. This work reports on the fluxes of red blood cell aggregates of various sizes in a T-shaped microchannel, aiming to clarify the effects of different flow conditions in the outlet branches of the channel. Image analysis techniques, were utilised, and moderately aggregating human red blood cell suspensions were tested in symmetric (∼50-50%) and asymmetric flow splits through the two outlet (daughter) branches. The results revealed that the flux decreases with aggregate size in the inlet (parent) and daughter branches, mainly due to the fact that the number of larger structures is significantly smaller than that of smaller structures. However, when the flux in the daughter branches is examined relative to the aggregate size flux in the parent branch an increase with aggregate size is observed for a range of asymmetric flow splits. This increase is attributed to size distribution and local concentration changes in the daughter branches. The results show that the flow of larger aggregates is not suppressed downstream of a bifurcation, and that blood flow is maintained, for physiological levels of red blood cell aggregation.
Wen JC, Reina-Torres E, Sherwood JM, 2017, Intravitreal Anti-VEGF Injections Reduce Aqueous Outflow Facility in Patients With Neovascular Age-Related Macular Degeneration' (vol 58, pg 1893, 2017), INVESTIGATIVE OPHTHALMOLOGY & VISUAL SCIENCE, Vol: 58, Pages: 2816-2816, ISSN: 0146-0404
Chandrawati R, Chang J, Reina-Torres E, et al., 2017, Localized and controlled delivery of nitric oxide to the conventional outflow pathway via enzyme biocatalysis: towards therapy for Glaucoma, Advanced Materials, Vol: 29, ISSN: 1521-4095
Nitric oxide (NO) has been shown to lower intraocular pressure (IOP), however its therapeutic effects on outflow physiology are location- and dose-dependent. Here, a NO delivery platform that directly targets the resistance-generating region of the conventional outflow pathway and locally liberates a controlled dose of NO is reported. An increase in outflow facility (decrease in IOP) is demonstrated in mouse model.
Campbell IC, Hannon BG, Read AT, et al., 2017, Quantification of the efficacy of collagen cross-linking agents to induce stiffening of rat sclera, Journal of the Royal Society Interface, Vol: 14, ISSN: 1742-5662
The concept of scleral stiffening therapies has emerged as a novel theoretical approach for treating the ocular disorders glaucoma and myopia. Deformation of specific regions of the posterior eye is innately involved in the pathophysiology of these diseases, and thus targeted scleral stiffening could resist these changes and slow or prevent progression of these diseases. Here, we present the first systematic screen and direct comparison of the stiffening effect of small molecule collagen cross-linking agents in the posterior globe, namely using glyceraldehyde, genipin and methylglyoxal (also called pyruvaldehyde). To establish a dose–response relationship, we used inflation testing to simulate the effects of increasing intraocular pressure in freshly harvested rat eyes stiffened with multiple concentrations of each agent. We used digital image correlation to compute the mechanical strain in the tissue as a metric of stiffness, using a novel treatment paradigm for screening relative stiffening by incubating half of each eye in cross-linker and using the opposite half as an internal control. We identified the doses necessary to increase stiffness by approximately 100%, namely 30 mM for glyceraldehyde, 1 mM for genipin and 7 mM for methylglyoxal, and we also identified the range of stiffening it was possible to achieve with such agents. Such findings will inform development of in vivo studies of scleral stiffening to treat glaucoma and myopia.
Kaliviotis E, Sherwood JM, Balabani S, 2017, Partitioning of red blood cell aggregates in bifurcating microscale flows, SCIENTIFIC REPORTS, Vol: 7, ISSN: 2045-2322
Microvascular flows are often considered to be free of red blood cell aggregates, however, recent studies have demonstrated that aggregates are present throughout the microvasculature, affecting cell distribution and blood perfusion. This work reports on the spatial distribution of red blood cell aggregates in a T-shaped bifurcation on the scale of a large microvessel. Non-aggregating and aggregating human red blood cell suspensions were studied for a range of flow splits in the daughter branches of the bifurcation. Aggregate sizes were determined using image processing. The mean aggregate size was marginally increased in the daughter branches for a range of flow rates, mainly due to the lower shear conditions and the close cell and aggregate proximity therein. A counterintuitive decrease in the mean aggregate size was apparent in the lower flow rate branches. This was attributed to the existence of regions depleted by aggregates of certain sizes in the parent branch, and to the change in the exact flow split location in the T-junction with flow ratio. The findings of the present investigation may have significant implications for microvascular flows and may help explain why the effects of physiological RBC aggregation are not deleterious in terms of in vivo vascular resistance.
Overby DR, 2017, VEGF as a paracrine regulator of conventional outflow facility, Investigative Ophthalmology & Visual Science, Vol: 58, Pages: 1899-1908, ISSN: 1552-5783
Purpose: Vascular endothelial growth factor (VEGF) regulates microvascular endothelial permeability, and the permeability of Schlemm's canal (SC) endothelium influences conventional aqueous humor outflow. We hypothesize that VEGF signaling regulates outflow facility.Methods: We measured outflow facility (C) in enucleated mouse eyes perfused with VEGF-A164a, VEGF-A165b, VEGF-D, or inhibitors to VEGF receptor 2 (VEGFR-2). We monitored VEGF-A secretion from human trabecular meshwork (TM) cells by ELISA after 24 hours of static culture or cyclic stretch. We used immunofluorescence microscopy to localize VEGF-A protein within the TM of mice.Results: VEGF-A164a increased C in enucleated mouse eyes. Cyclic stretch increased VEGF-A secretion by human TM cells, which corresponded to VEGF-A localization in the TM of mice. Blockade of VEGFR-2 decreased C, using either of the inhibitors SU5416 or Ki8751 or the inactive splice variant VEGF-A165b. VEGF-D increased C, which could be blocked by Ki8751.Conclusions: VEGF is a paracrine regulator of conventional outflow facility that is secreted by TM cells in response to mechanical stress. VEGF affects facility via VEGFR-2 likely at the level of SC endothelium. Disruption of VEGF signaling in the TM may explain why anti-VEGF therapy is associated with decreased outflow facility and sustained ocular hypertension.
Wen JC, Reina-Torres E, Sherwood JM, et al., 2017, Intravitreal anti-VEGF injections reduce aqueous outflow facility in patients with neovascular age-related macular degeneration, Investigative Ophthalmology & Visual Science, Vol: 58, Pages: 1893-1898, ISSN: 0146-0404
Purpose: We assess the effect of intravitreal anti-VEGF injections on tonographic outflow facility.Methods: Patients with age-related macular degeneration who had received unilateral intravitreal anti-VEGF injections were recruited into two groups, those with ≤10 and those with ≥20 total anti-VEGF injections. Intraocular pressure and tonographic outflow facility of injected and uninjected fellow eyes were measured and compared between groups. Risk factors for development of reduced outflow facility also were assessed.Results: Outflow facility was 12% lower in the injected eyes of patients who received ≥20 anti-VEGF injections, compared to contralateral uninjected eyes (P = 0.02). In contrast, there was no facility reduction for patients with ≤10 anti-VEGF injections (P = 0.4). In patients with ocular hypertension in the uninjected eye (IOP > 21 mm Hg, n = 5), the outflow facility of injected eyes was on average 46% lower (P = 0.01) than in the uninjected fellow eyes. This was significantly greater than the difference observed in patients with IOP ≤ 21 mm Hg in the uninjected eye (P = 2 × 10−4). In patients with ocular hypertension in the injected eye (n = 6) the differences in facility and IOP between contralateral eyes were significantly greater than in patients with IOP ≤ 21 mm Hg in the injected eye (P = 2 × 10−4 and P = 7 × 10−4, respectively).Conclusions: Chronic anti-VEGF injections significantly reduce outflow facility in patients with AMD. The greatest facility reduction is observed in patients with baseline ocular hypertension. Ophthalmologists who administer anti-VEGF injections should be aware of these findings and monitor patients closely for changes in IOP or evidence of glaucoma, especially in those with pre-existing ocular hypertension.
O'Callaghan J, Crosbie DE, Cassidy PS, et al., 2017, Therapeutic potential of AAV-mediated MMP-3 secretion from corneal endothelium in treating glaucoma, HUMAN MOLECULAR GENETICS, Vol: 26, Pages: 1230-1246, ISSN: 0964-6906
Intraocular pressure (IOP) is maintained as a result of the balance between production of aqueous humour (AH) by the ciliary processes and hydrodynamic resistance to its outflow through the conventional outflow pathway comprising the trabecular meshwork (TM) and Schlemm’s canal (SC). Elevated IOP, which can be caused by increased resistance to AH outflow, is a major risk factor for open-angle glaucoma. Matrix metalloproteinases (MMPs) contribute to conventional aqueous outflow homeostasis in their capacity to remodel extracellular matrices, which has a direct impact on aqueous outflow resistance and IOP. We observed decreased MMP-3 activity in human glaucomatous AH compared to age-matched normotensive control AH. Treatment with glaucomatous AH resulted in significantly increased transendothelial resistance of SC endothelial and TM cell monolayers and reduced monolayer permeability when compared to control AH, or supplemented treatment with exogenous MMP-3.Intracameral inoculation of AAV-2/9 containing a CMV-driven MMP-3 gene (AAV-MMP-3) into wild type mice resulted in efficient transduction of corneal endothelium and an increase in aqueous concentration and activity of MMP-3. Most importantly, AAV-mediated expression of MMP-3 increased outflow facility and decreased IOP, and controlled expression using an inducible promoter activated by topical administration of doxycycline achieved the same effect. Ultrastructural analysis of MMP-3 treated matrices by transmission electron microscopy revealed remodelling and degradation of core extracellular matrix components. These results indicate that periodic induction, via use of an eye drop, of AAV-mediated secretion of MMP-3 into AH could have therapeutic potential for those cases of glaucoma that are sub-optimally responsive to conventional pressure-reducing medications.
Tam LC, Reina-Torres E, Sherwood JM, et al., 2017, Enhancement of outflow facility in the murine eye by targeting selected tight-junctions of Schlemm's canal endothelia, Scientific Reports, Vol: 7, ISSN: 2045-2322
The juxtacanalicular connective tissue of the trabecular meshwork together with inner wall endothelium of Schlemm’s canal (SC) provide the bulk of resistance to aqueous outflow from the anterior chamber. Endothelial cells lining SC elaborate tight junctions (TJs), down-regulation of which may widen paracellular spaces between cells, allowing greater fluid outflow. We observed significant increase in paracellular permeability following siRNA-mediated suppression of TJ transcripts, claudin-11, zonula-occludens-1 (ZO-1) and tricellulin in human SC endothelial monolayers. In mice claudin-11 was not detected, but intracameral injection of siRNAs targeting ZO-1 and tricellulin increased outflow facility significantly. Structural qualitative and quantitative analysis of SC inner wall by transmission electron microscopy revealed significantly more open clefts between endothelial cells treated with targeting, as opposed to non-targeting siRNA. These data substantiate the concept that the continuity of SC endothelium is an important determinant of outflow resistance, and suggest that SC endothelial TJs represent a specific target for enhancement of aqueous movement through the conventional outflow system.
Kaliviotis E, Dusting J, Sherwood JM, et al., 2016, Quantifying local characteristics of velocity, aggregation and hematocrit of human erythrocytes in a microchannel flow, Clinical Hemorheology and Microcirculation, Vol: 63, Pages: 123-148, ISSN: 1875-8622
The effect of erythrocyte aggregation on blood viscosity and microcirculatory flow is a poorly understood area of haemodynamics, especially with relevance to serious pathological conditions. Advances in microfluidics have made it possible to study the details of blood flow in the microscale, however, important issues such as the relationship between the local microstructure and local flow characteristics have not been investigated extensively. In the present study an experimental system involving simple brightfield microscopy has been successfully developed for simultaneous, time-resolved quantification of velocity fields and local aggregation of human red blood cells (RBC) in microchannels. RBCs were suspended in Dextran and phosphate buffer saline solutions for the control of aggregation. Local aggregation characteristics were investigated at bulk and local levels using statistical and edge-detection image processing techniques. A special case of aggregating flow in a microchannel, in which hematocrit gradients were present, was studied as a function of flowrate and time. The level of aggregation was found to strongly correlate with local variations in velocity in both the bulk flow and wall regions. The edge detection based analysis showed that near the side wall large aggregates are associated with regions corresponding to high local velocities and low local shear. On the contrary, in the bulk flow region large aggregates occurred in regions of low velocity and high erythrocyte concentration suggesting a combined effect of haematocrit and velocity distributions on local aggregation characteristics. The results of this study showed that using multiple methods for aggregation quantification, albeit empirical, could help towards a robust characterisation of the structural properties of the fluid.
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