Maintaining healthy aqueous humour outflow (AHO) is important for intraocular cellular health and stable vision. Impairment of AHO can lead to increased intraocular pressure, optic nerve damage and concomitant glaucoma. An improved understanding of AHO will lead to improved glaucoma surgeries that enhance native AHO as well as facilitate the development of AHO-targeted pharmaceuticals. Recent AHO imaging has evolved to live human assessment and has focused on the structural evaluation of AHO pathways and the functional documentation of fluid flow. Structural AHO evaluation is predominantly driven by optical coherence tomography, and functional evaluation of flow is performed using various methods, including aqueous angiography. Advances in structural and functional evaluation of AHO are reviewed with discussion of strengths, weaknesses and potential future directions.

Anterior segment glaucoma clinical care and research has recently gained new focus because of novel imaging modalities and the advent of angle-based surgical treatments. Traditional investigation drawn to the trabecular meshwork now emphasizes the entire conventional aqueous humor outflow (AHO) pathway from the anterior chamber to the episcleral vein. AHO investigation can be divided into structural and functional assessments using different methods. The historical basis for studying the anterior segment of the eye and AHO in glaucoma is discussed. Structural studies of AHO are reviewed and include traditional pathological approaches to modern tools such as multi-model two-photon microscopy and optical coherence tomography. Functional assessment focuses on visualizing AHO itself through a variety of non-real-time and real-time techniques such as aqueous angiography. Implications of distal outflow resistance and segmental AHO are discussed with an emphasis on melding bench-side research to viable clinical applications. Through the development of an improved structure: function relationship for AHO in the anterior segment of the normal and diseased eye, a better understanding of the eye with improved therapeutics may be developed.

Maintaining healthy aqueous humour outflow (AHO) is important for intraocular cellular health and stable vision. Impairment of AHO can lead to increased intraocular pressure, optic nerve damage and concomitant glaucoma. An improved understanding of AHO will lead to improved glaucoma surgeries that enhance native AHO as well as facilitate the development of AHO-targeted pharmaceuticals. Recent AHO imaging has evolved to live human assessment and has focused on the structural evaluation of AHO pathways and the functional documentation of fluid flow. Structural AHO evaluation is predominantly driven by optical coherence tomography, and functional evaluation of flow is performed using various methods, including aqueous angiography. Advances in structural and functional evaluation of AHO are reviewed with discussion of strengths, weaknesses and potential future directions.

Purpose

To study changes in aqueous humor outflow (AHO) patterns after trabecular micro-bypass (TMB) in glaucoma patients using intraoperative sequential aqueous angiography.

Design

Prospective comparative case series.

Subjects

Fifteen subjects (14 with glaucoma and 1 normal).

Methods

Sequential aqueous angiography (Spectralis HRA+OCT; Heidelberg Engineering) was performed on fourteen glaucoma patients undergoing routine TMB (iStent Inject; Glaukos Corporation) and cataract surgery and one normal patient undergoing cataract surgery alone. Indocyanine green (ICG) aqueous angiography established initial baseline nasal angiographic AHO patterns. Two TMB stents were placed in regions of baseline low or high angiographic AHO in each eye (n = 2 eyes with enough space to place two stents in both low angiographic regions; n = 8 eyes with two stents both placed in high angiographic regions; n = 4 eyes with enough space to place one stent in a low angiographic region and the other stent in a high angiographic region). Subsequent fluorescein aqueous angiography was utilized to query alterations to angiographic AHO patterns.

Main outcome measure

Angiographic signal and patterns before and after TMB.

Results

At baseline, all eyes showed segmental angiographic AHO patterns. Focused on the nasal hemisphere of each eye, for each stent TMB in initially low ICG angiographic signal regions showed transient or persistently improved fluorescein angiographic signal (11.2-fold; p = 0.014). TMB in initially high ICG signal regions led to faster development of fluorescein angiographic patterns (3.1-fold; p = 0.02).

Conclusion

TMB resulted in different patterns of aqueous angiographic AHO improvement whose further understanding may advance basic knowledge of AHO and possibly enhance intraocular pressure reduction after glaucoma surgery in the future.

Steroid-induced ocular hypertension can be seen even after trabecular meshwork (TM) bypass/ablation. Thus, the purpose was to investigate steroid-response in cells distal to the TM by using primary scleral fibroblasts. Primary scleral cell cultures were generated using mid-depth scleral wedges from human donor corneo-scleral rims (n = 5) after corneal transplantation. Cells were treated with dexamethasone (DEX; 100 nM) and compared to media (MED)/vehicle (DMSO) controls. Cell size, shape, and migration were studied using the IncuCyte Live-Cell Analysis System. Cytoskeleton was compared using Alexa Fluor-568 Phalloidin and senescence tested by evaluating beta-galactosidase. Western blot comparison was performed for α-SMA, FKBP-51, fibronectin, phospho-myosin light chain, and myocilin. Scleral fibroblasts upregulated FKBP-51 in response to DEX indicating the existence of steroid-responsive pathways. Compared to controls, DEX-treated cells proliferated slower (~50%; p < 0.01-0.02), grew larger (~1.3-fold; p < 0.001), and migrated less (p = 0.01-0.006). Alexa Fluor 568 Phalloidin actin stress fiber labeling was more diffuse in DEX-treated cells (p = 0.001-0.004). DEX-treated cells showed more senescence compared to controls (~1.7-fold; p = 0.01-0.02). However, DEX-treated cells did not show increased cross-linked actin network formation or elevated myocilin/fibronectin/α-SMA/phospho-myosin light chain protein expression. For all parameters, MED- and DMSO-treated control cells were not significantly different. Primary scleral fibroblasts, grown from tissue collected immediately distal to the TM, demonstrated scleral-response behaviors that were similar to, but not identical with, classic TM steroid-response. Further study is needed to understand how these scleral cellular alterations may contribute to steroid-response IOP elevation after TM bypass/ablation surgery.

Purpose

To study changes in aqueous humor outflow (AHO) patterns after trabecular micro-bypass (TMB) in glaucoma patients using intraoperative sequential aqueous angiography.

Design

Prospective comparative case series.

Subjects

Fifteen subjects (14 with glaucoma and 1 normal).

Methods

Sequential aqueous angiography (Spectralis HRA+OCT; Heidelberg Engineering) was performed on fourteen glaucoma patients undergoing routine TMB (iStent Inject; Glaukos Corporation) and cataract surgery and one normal patient undergoing cataract surgery alone. Indocyanine green (ICG) aqueous angiography established initial baseline nasal angiographic AHO patterns. Two TMB stents were placed in regions of baseline low or high angiographic AHO in each eye (n = 2 eyes with enough space to place two stents in both low angiographic regions; n = 8 eyes with two stents both placed in high angiographic regions; n = 4 eyes with enough space to place one stent in a low angiographic region and the other stent in a high angiographic region). Subsequent fluorescein aqueous angiography was utilized to query alterations to angiographic AHO patterns.

Main outcome measure

Angiographic signal and patterns before and after TMB.

Results

At baseline, all eyes showed segmental angiographic AHO patterns. Focused on the nasal hemisphere of each eye, for each stent TMB in initially low ICG angiographic signal regions showed transient or persistently improved fluorescein angiographic signal (11.2-fold; p = 0.014). TMB in initially high ICG signal regions led to faster development of fluorescein angiographic patterns (3.1-fold; p = 0.02).

Conclusion

TMB resulted in different patterns of aqueous angiographic AHO improvement whose further understanding may advance basic knowledge of AHO and possibly enhance intraocular pressure reduction after glaucoma surgery in the future.

The purpose was to create a three-dimensional (3-D) model of circumferential aqueous humor outflow (AHO) in a living human eye with an automated detection algorithm for Schlemm’s canal (SC) and first-order collector channels (CC) applied to spectral-domain optical coherence tomography (SD-OCT). Anterior segment SD-OCT scans from a subject were acquired circumferentially around the limbus. A Bayesian Ridge method was used to approximate the location of the SC on infrared confocal laser scanning ophthalmoscopic images with a cross multiplication tool developed to initiate SC/CC detection automated through a fuzzy hidden Markov Chain approach. Automatic segmentation of SC and initial CC’s was manually confirmed by two masked graders. Outflow pathways detected by the segmentation algorithm were reconstructed into a 3-D representation of AHO. Overall, only <1% of images (5114 total B-scans) were ungradable. Automatic segmentation algorithm performed well with SC detection 98.3% of the time and <0.1% false positive detection compared to expert grader consensus. CC was detected 84.2% of the time with 1.4% false positive detection. 3-D representation of AHO pathways demonstrated variably thicker and thinner SC with some clear CC roots. Circumferential (360 deg), automated, and validated AHO detection of angle structures in the living human eye with reconstruction was possible.

Purpose

To evaluate the feasibility of safely performing aqueous angiography in intact eyes of living nonhuman primates (NHPs) for evaluating aqueous humor outflow and segmental patterns.

Design

Cross-sectional, observational study.

Subjects

Six nonhuman primates.

Methods

Aqueous angiography was performed in 6 nonhuman primates. After anesthesia, an anterior chamber (AC) maintainer was placed through a temporal 1-mm side-port wound. Indocyanine green (ICG; 0.4%) or 2.5% fluorescein was introduced (individually or in sequence) into the eye with a gravity-driven constant-pressure system. Aqueous angiography images were obtained with a Spectralis HRA+OCT (Heidelberg Engineering GmbH, Heidelberg, Germany) suspended over the NHP eye using a custom-designed surgical boom arm. Concurrent anterior segment optical coherence tomography (OCT) was performed on distally angiographically positive and negative regions.

Main outcome measures

Angiographic patterns described by location, time-course, choice of tracer, and anterior-segment OCT.

Results

Aqueous angiography in the living NHP eye demonstrated mostly stable angiographic patterns. With multimodal imaging, angiographically positive signal co-localized with episcleral veins as identified by infrared imaging and intrascleral lumens, as demonstrated by anterior segment OCT. Sequential aqueous angiography in individual eyes with ICG followed by fluorescein showed similar angiographic patterns. A pulsatile nature of aqueous angiographic outflow was sometimes observed. Aqueous angiographic patterns could also dynamically change. In some instances, positive angiographic flow suddenly arose in regions previously without an angiographic signal. Alternatively, an angiographic signal could suddenly disappear from regions in which an angiographic signal was initially documented.

Conclusions

Aqueous angiography in living NHPs demonstrated segmental and pulsatile patterns with a newly described ability to dynamically shift. These characteristics further the understanding of live aqueous humor outflow biology and may be useful in improving glaucoma surgeries aimed at trabecular meshwork bypass.

The purpose was to create a three-dimensional (3-D) model of circumferential aqueous humor outflow (AHO) in a living human eye with an automated detection algorithm for Schlemm’s canal (SC) and first-order collector channels (CC) applied to spectral-domain optical coherence tomography (SD-OCT). Anterior segment SD-OCT scans from a subject were acquired circumferentially around the limbus. A Bayesian Ridge method was used to approximate the location of the SC on infrared confocal laser scanning ophthalmoscopic images with a cross multiplication tool developed to initiate SC/CC detection automated through a fuzzy hidden Markov Chain approach. Automatic segmentation of SC and initial CC’s was manually confirmed by two masked graders. Outflow pathways detected by the segmentation algorithm were reconstructed into a 3-D representation of AHO. Overall, only <1% of images (5114 total B-scans) were ungradable. Automatic segmentation algorithm performed well with SC detection 98.3% of the time and <0.1% false positive detection compared to expert grader consensus. CC was detected 84.2% of the time with 1.4% false positive detection. 3-D representation of AHO pathways demonstrated variably thicker and thinner SC with some clear CC roots. Circumferential (360 deg), automated, and validated AHO detection of angle structures in the living human eye with reconstruction was possible.