Ralitsa T. Loewen

Bio: Ralitsa T. Loewen is an academic researcher from University of Würzburg. The author has an hindex of 2, co-authored 2 publications receiving 5 citations.

3D-Reconstruction of the human conventional outflow system by ribbon scanning confocal microscopy.

Abstract: Purpose The risk for glaucoma is driven by the microanatomy and function of the anterior segment. We performed a computation-intense, high-resolution, full-thickness ribbon-scanning confocal microscopy (RSCM) of the outflow tract of two human eyes. We hypothesized this would reveal important species differences when compared to existing data of porcine eyes, an animal that does not spontaneously develop glaucoma. Methods After perfusing two human octogenarian eyes with lectin-fluorophore conjugate and optical clearance with benzyl alcohol benzyl benzoate (BABB), anterior segments were scanned by RSCM and reconstructed in 3D for whole-specimen rendering. Morphometric analyses of the outflow tract were performed for the trabecular meshwork (TM), limbal, and perilimbal outflow structures and compared to existing porcine data. Results RSCM provided high-resolution data for IMARIS-based surface reconstruction of outflow tract structures in 3D. Different from porcine eyes with an abundance of highly interconnected, narrow, and short collector channels (CCs), human eyes demonstrated fewer CCs which had a 1.5x greater cross-sectional area (CSA) and 2.6x greater length. Proximal CC openings at the level of Schlemm's canal (SC) had a 1.3x larger CSA than distal openings into the scleral vascular plexus (SVP). CCs were 10.2x smaller in volume than the receiving SVP vessels. Axenfeld loops, projections of the long ciliary nerve, were also visualized. Conclusion In this high-resolution, volumetric RSCM analysis, human eyes had far fewer outflow tract vessels than porcine eyes. Human CCs spanned several clock-hours and were larger than in porcine eyes. These species differences may point to factors downstream of the TM that increase our vulnerability to glaucoma.

3D-Reconstruction of the Human Conventional Outflow System by Ribbon Scanning Confocal Microscopy

Abstract: Purpose: The risk for glaucoma is driven by the microanatomy and function of the anterior segment. We performed a computation-intense, high-resolution, full-thickness ribbon-scanning confocal microscopy (RSCM) of the outflow tract of two human eyes. We hypothesized this would reveal important species differences when compared to existing data of porcine eyes, an animal that does not spontaneously develop glaucoma. Methods: After perfusing two human octogenarian eyes with lectin-fluorophore conjugate and optical clearance with benzyl alcohol benzyl benzoate (BABB), anterior segments were scanned by RSCM and reconstructed in 3D for whole-specimen rendering. Morphometric analyses of the outflow tract were performed for the trabecular meshwork (TM), limbal, and perilimbal outflow structures and compared to existing porcine data. Results: RSCM provided high-resolution data for IMARIS-based surface reconstruction of outflow tract structures in 3D. Different from porcine eyes with an abundance of highly interconnected, narrow, and short collector channels (CCs), human eyes demonstrated fewer CCs which had a 1.5x greater cross-sectional area (CSA) and 2.6x greater length. Proximal CC openings at the level of Schlemm9s canal (SC) had a 1.3x larger CSA than distal openings into the scleral vascular plexus (SVP). CCs were 10.2x smaller in volume than the receiving SVP vessels. Axenfeld loops, projections of the long ciliary nerve, were also visualized. Conclusion: In this high-resolution, volumetric RSCM analysis, human eyes had far fewer outflow tract vessels than porcine eyes. Human CCs spanned several clock-hours and were larger than in porcine eyes. These species differences may point to factors downstream of the TM that increase our vulnerability to glaucoma.

Regionally Discrete Aqueous Humor Outflow Quantification using Fluorescein Canalograms

Abstract: Purpose To visualize and quantify conventional outflow directly in its anatomic location. Methods We obtained fluorescein canalograms in six porcine whole eyes and six porcine anterior segment cultures. Eyes were perfused with a constant pressure of 15 mmHg using media containing 0.017 mg/ml fluorescein. Flow patterns were visualized using a stereo dissecting microscope equipped for fluorescent imaging. Images were captured every 30 seconds for 20 minutes for time lapse analysis. Anterior chamber cultures were imaged again on day three of culture. Canalograms were first analyzed for filling time per quadrant. We then wrote a program to automatically compute focal flow fits for each macropixel and to detect convergent perilimbal flow patterns with macropixels grouped into 3 equal-radial width rings around the cornea. A generalized additive model was used to determine fluorescence changes of individual macropixels. Results The resulting imaging algorithm deployed 1024 macropixels that were fit to determine maximum intensity and time to fill. These individual fits highlighted the focal flow function. In whole eyes, significantly faster flow was seen in the inferonasal (IN) and superonasal (SN) quadrants compared to the superotemporal (ST) and inferotemporal (IT) ones (p<0.05). In anterior chamber cultures, reduced flow on day 1 increased in all quadrants on day 3 except in IT (p<0.05). Perilimbal ring analysis uncovered convergent perilimbal flow. Conclusions An algorithm was developed that analyzes regional and circumferential outflow patterns. This algorithm found flow patterns that changed over time and differ in whole eyes and anterior segment cultures.

Aqueous outflow imaging techniques and what they tell us about intraocular pressure regulation.

Abstract: Recent advances in the medical and surgical management of open-angle glaucoma have increased the number of treatment options available. Several new intraocular pressure (IOP)-lowering treatments target the conventional aqueous outflow (AO) system. However, success rates are variable and outcomes in individual patients are often difficult to predict. Variable treatment responses remain unexplained and highlight deficiencies in our current understanding of AO regulation and IOP homeostasis. Imaging is often relied upon to confirm diagnoses and monitor treatment responses in other ocular and systemic pathologies. As yet no suitable AO imaging tool has been developed to fulfil this role in glaucoma. A variety of imaging techniques have been used to study the AO tracts of humans and animals in ex vivo and in vivo eyes. In this review, results from novel imaging techniques that assess aqueous drainage through the episcleral venous system are considered and we argue these provide new insights into AO regulation. We suggest that the ability to objectively measure AO responses to interventions would be a significant clinical advance, and we have demonstrated that this can be achieved with direct visualisation of aqueous drainage. We predict that the evolution of AO imaging technology will continue to reveal critical components of AO and IOP regulation, and that personalised IOP-lowering treatment in glaucoma care may well become a reality in the near future.

A Predictive Test of Outflow Enhancement by Ab Interno Trabeculectomy

Abstract: Purpose: To investigate trabeculopuncture (TP) for predicting the outcome of ab interno trabeculectomy (AIT). AIT is an effective, low-risk procedure for open angle glaucoma. Despite widespread utilization, it fails in patients with an unidentified distal outflow resistance. Methods: We bisected 81 enucleated porcine eyes and perfused them for 72 hours. They were assigned to two groups: trial (n=42) and control (n=39). Intraocular pressure (IOP) was measured continuously. At 24 hours, four YAG-laser trabeculopunctures on the nasal trabecular meshwork were performed, followed by a 180° AIT at the same site at 48 hours. Eyes were divided into TP and AIT responders and non-responders; the proportion of TP responders between both AIT groups was compared. Results: Both post-TP and post-AIT IOPs were lower than baseline IOP (p=0.015 and p<0.01, respectively). The success rates of TP and AIT were 69% and 85.7%, respectively. The proportion of TP responders among AIT responders was greater than that of AIT non-responders (p<0.01). Sensitivity and specificity values of TP as predictive test for AIT success were 77.7% and 83.3%, respectively. The positive and negative predictive values were 96.6% and 38.5%, respectively. Conclusion: A 10% reduction in IOP after TP can be used as predictor for the success (>20% IOP decrease) of 180° AIT in porcine eyes.

Trabeculopuncture as a predictive test of distal outflow resistance in canal-based surgery

Abstract: We investigated whether trabeculopuncture (TP) could detect distal outflow resistance to predict the outcome of canal-based glaucoma surgery such as ab interno trabeculectomy (AIT). These procedures have a high utilization in open angle glaucoma, but fail in eyes with an unidentified distal outflow resistance. We assigned 81 porcine eyes to two groups: trial (n = 42) and control (n = 39). At 24 h, four YAG-laser trabeculopunctures were placed nasally, followed by a 180° AIT at the same site at 48 h. The proportion of TP responders between both AIT groups was compared. Histology and outflow canalograms were determined. Both post-TP and post-AIT IOPs were lower than baseline IOP (p = 0.015 and p < 0.01, respectively). The success rates of TP and AIT were 69% and 85.7%, respectively. Sensitivity and specificity values of TP as predictive test for AIT success were 77.7% and 83.3%, respectively. The positive and negative predictive values were 96.6% and 38.5%, respectively. We conclude that a 10% reduction in IOP after TP can be used as a predictor for the success (> 20% IOP decrease) of 180° AIT in porcine eyes.