James C. H. Tan

Aqueous Angiography: Real-Time and Physiologic Aqueous Humor Outflow Imaging.

Purpose Trabecular meshwork (TM) bypass surgeries attempt to enhance aqueous humor outflow (AHO) to lower intraocular pressure (IOP). While TM bypass results are promising, inconsistent success is seen. One hypothesis for this variability rests upon segmental (non-360 degrees uniform) AHO. We describe aqueous angiography as a real-time and physiologic AHO imaging technique in model eyes as a way to simulate live AHO imaging. Methods Pig (n = 46) and human (n = 6) enucleated eyes were obtained, orientated based upon inferior oblique insertion, and pre-perfused with balanced salt solution via a Lewicky AC maintainer through a 1mm side-port. Fluorescein (2.5%) was introduced intracamerally at 10 or 30 mm Hg. With an angiographer, infrared and fluorescent (486 nm) images were acquired. Image processing allowed for collection of pixel information based on intensity or location for statistical analyses. Concurrent OCT was performed, and fixable fluorescent dextrans were introduced into the eye for histological analysis of angiographically active areas. Results Aqueous angiography yielded high quality images with segmental patterns (p<0.0001; Kruskal-Wallis test). No single quadrant was consistently identified as the primary quadrant of angiographic signal (p = 0.06–0.86; Kruskal-Wallis test). Regions of high proximal signal did not necessarily correlate with regions of high distal signal. Angiographically positive but not negative areas demonstrated intrascleral lumens on OCT images. Aqueous angiography with fluorescent dextrans led to their trapping in AHO pathways. Conclusions Aqueous angiography is a real-time and physiologic AHO imaging technique in model eyes.

Aqueous Angiography-Mediated Guidance of Trabecular Bypass Improves Angiographic Outflow in Human Enucleated Eyes.

Purpose To assess the ability of trabecular micro-bypass stents to improve aqueous humor outflow (AHO) in regions initially devoid of AHO as assessed by aqueous angiography. Methods Enucleated human eyes (14 total from 7 males and 3 females [ages 52–84]) were obtained from an eye bank within 48 hours of death. Eyes were oriented by inferior oblique insertion, and aqueous angiography was performed with indocyanine green (ICG; 0.4%) or fluorescein (2.5%) at 10 mm Hg. With an angiographer, infrared and fluorescent images were acquired. Concurrent anterior segment optical coherence tomography (OCT) was performed, and fixable fluorescent dextrans were introduced into the eye for histologic analysis of angiographically positive and negative areas. Experimentally, some eyes (n = 11) first received ICG aqueous angiography to determine angiographic patterns. These eyes then underwent trabecular micro-bypass sham or stent placement in regions initially devoid of angiographic signal. This was followed by fluorescein aqueous angiography to query the effects. Results Aqueous angiography in human eyes yielded high-quality images with segmental patterns. Distally, angiographically positive but not negative areas demonstrated intrascleral lumens on OCT images. Aqueous angiography with fluorescent dextrans led to their trapping in AHO pathways. Trabecular bypass but not sham in regions initially devoid of ICG aqueous angiography led to increased aqueous angiography as assessed by fluorescein (P = 0.043). Conclusions Using sequential aqueous angiography in an enucleated human eye model system, regions initially without angiographic flow or signal could be recruited for AHO using a trabecular bypass stent.

Aqueous Angiography with Fluorescein and Indocyanine Green in Bovine Eyes

Purpose We characterize aqueous angiography as a real-time aqueous humor outflow imaging (AHO) modality in cow eyes with two tracers of different molecular characteristics. Methods Cow enucleated eyes (n = 31) were obtained and perfused with balanced salt solution via a Lewicky AC maintainer through a 1-mm side-port. Fluorescein (2.5%) or indocyanine green (ICG; 0.4%) were introduced intracamerally at 10 mm Hg individually or sequentially. With an angiographer, infrared and fluorescent images were acquired. Concurrent anterior segment optical coherence tomography (OCT) was performed, and fixable fluorescent dextrans were introduced into the eye for histologic analysis of angiographically positive and negative areas. Results Aqueous angiography in cow eyes with fluorescein and ICG yielded high-quality images with segmental patterns. Over time, ICG maintained a better intraluminal presence. Angiographically positive, but not negative, areas demonstrated intrascleral lumens with anterior segment OCT. Aqueous angiography with fluorescent dextrans led to their trapping in AHO pathways. Sequential aqueous angiography with ICG followed by fluorescein in cow eyes demonstrated similar patterns. Conclusions Aqueous angiography in model cow eyes demonstrated segmental angiographic outflow patterns with either fluorescein or ICG as a tracer. Translational Relevance Further characterization of segmental AHO with aqueous angiography may allow for intelligent placement of trabecular bypass minimally invasive glaucoma surgeries for improved surgical results.

Deep tissue analysis of distal aqueous drainage structures and contractile features

Outflow resistance in the aqueous drainage tract distal to trabecular meshwork is potentially an important determinant of intraocular pressure and success of trabecular bypass glaucoma surgeries. It is unclear how distal resistance is modulated. We sought to establish: (a) multimodal 2-photon deep tissue imaging and 3-dimensional analysis of the distal aqueous drainage tract (DT) in transgenic mice in vivo and ex vivo; (b) criteria for distinguishing the DT from blood and lymphatic vessels; and (c) presence of a DT wall organization capable of contractility. DT lumen appeared as scleral collagen second harmonic generation signal voids that could be traced back to Schlemm’s canal. DT endothelium was Prox1-positive, CD31-positive and LYVE-1-negative, bearing a different molecular signature from blood and true lymphatic vessels. DT walls showed prominent filamentous actin (F-actin) labeling reflecting cells in a contracted state. F-actin co-localized with mesenchymal smooth muscle epitopes of alpha-smooth muscle actin, caldesmon and calponin, which localized adjacent and external to the endothelium. Our findings support a DT wall organization resembling that of blood vessels. This reflects a capacity to contract and support dynamic alteration of DT caliber and resistance analogous to the role of blood vessel tone in regulating blood flow.

Toward in vivo two-photon analysis of mouse aqueous outflow structure and function

ABSTRACT The promise of revolutionary insights into intraocular pressure (IOP) and aqueous humor outflow homeostasis, IOP pathogenesis, and novel therapy offered by engineered mouse models has been hindered by a lack of appropriate tools for studying the aqueous drainage tissues in their original 3‐dimensional (3D) environment. Advances in 2‐photon excitation fluorescence imaging (TPEF) combined with availability of modalities such as transgenic reporter mice and intravital dyes have placed us on the cusp of unlocking the potential of the mouse model for unearthing insights into aqueous drainage structure and function. Multimodality 2‐photon imaging permits high‐resolution visualization not only of tissue structural organization but also cells and cellular function. It is possible to dig deeper into understanding the cellular basis of aqueous outflow regulation as the technique integrates analysis of tissue structure, cell biology and physiology in a way that could also lead to fresh insights into human glaucoma. We outline recent novel applications of two‐photon imaging to analyze the mouse conventional drainage system in vivo or in whole tissues: (1) collagen second harmonic generation (SHG) identifies the locations of episcleral vessels, intrascleral plexuses, collector channels, and Schlemms canal in the distal aqueous drainage tract; (2) the prospero homeobox protein 1‐green fluorescent protein (GFP) reporter helps locate the inner wall of Schlemms canal; (3) Calcein AM, siGLO™, the fluorescent reporters m‐Tomato and GFP, and coherent anti‐Stokes scattering (CARS), are adjuncts to TPEF to identify live cells by their membrane or cytosolic locations; (4) autofluorescence and sulforhodamine‐B to identify elastic fibers in the living eye. These tools greatly expand our options for analyzing physiological and pathological processes in the aqueous drainage tissues of live mice as a model of the analogous human system.

Implantation of a second glaucoma drainage device

PurposeTo evaluate success rates in controlling intraocular pressure (IOP) after implantation of a second glaucoma drainage device (GDD) with a Baerveldt glaucoma implant in patients with refractory glaucoma, with a secondary aim of reducing the need for postoperative glaucoma medications.Material and methodsThis retrospective, noncomparative, interventional study included patients undergoing a second GDD for uncontrolled glaucoma from a tertiary care glaucoma service. Data were obtained from the medical records for the preoperative period and after the 1st, 15th, and 30th day, 3, 6, and 12xa0months, and then yearly until the last postoperative visit. Visual acuity, IOP, and number of glaucoma medications (NGM) from the follow-up visits were compared to baseline. Success and failure criteria were analyzed based on IOP level or need of glaucoma medications.ResultsForty-nine patients were studied, with a mean follow-up time of 25u2009±u200921xa0months. The mean preoperative IOP was 23.7u2009±u20098.2xa0mmHg, and decreased to 14.8u2009±u20094.0xa0mmHg after 1xa0year, 14.4u2009±u20093.9xa0mmHg after 2xa0years, and 16.6u2009±u20098.5xa0mmHg after 3xa0years. The mean preoperative NGM was 3.4u2009±u20091.3, and decreased to 2.0u2009±u20091.8 after 1xa0year, 2.5u2009±u20091.6 after 2xa0years, and 2.8u2009±u20092.0 after 3xa0years. Absolute success was 9% after 1xa0year for a postoperative IOP between 5 and 18xa0mmHg, and 76% for a postoperative IOP between 5 and 21xa0mmHg. The qualified success was 88% at the first and second years and 83% at the third year.ConclusionWith up to 3xa0years of follow-up, a second glaucoma drainage device was successful in reducing IOP to below 21xa0mmHg, but not as successful below 18xa0mmHg. The success rate is improved with the use of glaucoma medications with up to 3xa0years of follow-up.

Endoscopic cyclophotocoagulation versus second glaucoma drainage device after prior aqueous tube shunt surgery.

To evaluate the efficacy in controlling intraocular pressure (IOP) with endoscopic cyclophotocoagulation (ECP) versus implantation of a second glaucoma drainage device (GDD‐2) in the treatment of uncontrolled glaucoma with a prior aqueous tube shunt.