Faculty Bibliography

Optic nerve head (ONH) tissue properties and biomechanics remain mostly unmeasurable in the experiment. We hypothesized that these can be estimated numerically from ocular parameters measurable in vivo with optical coherence tomography (OCT). Using parametric models representing human ONHs we simulated acute intraocular pressure (IOP) increases (10 mmHg). Statistical models were fit to predict, from OCT-measurable parameters, 15 outputs, including ONH tissue properties, stresses, and deformations. The calculations were repeated adding parameters that have recently been proposed as potentially measurable with OCT. We evaluated the sensitivity of the predictions to variations in the experimental parameters. Excellent fits were obtained to predict all outputs from the experimental parameters, with cross-validated R2s between 0.957 and 0.998. Incorporating the potentially measurable parameters improved fits significantly. Predictions of tissue stiffness were accurate to within 0.66 MPa for the sclera and 0.24 MPa for the lamina cribrosa. Predictions of strains and stresses were accurate to within 0.62% and 4.9 kPa, respectively. Estimates of ONH biomechanics and tissue properties can be obtained quickly from OCT measurements using an applet that we make freely available. These estimates may improve understanding of the eye sensitivity to IOP and assessment of patient risk for development or progression of glaucoma.

Primary open-angle glaucoma (POAG) is a leading cause of blindness worldwide. Using genome-wide association single-nucleotide polymorphism data from the Glaucoma Genes and Environment study and National Eye Institute Glaucoma Human Genetics Collaboration comprising 3,108 cases and 3,430 controls, we assessed biologic pathways as annotated in the KEGG database for association with risk of POAG. After correction for genic overlap among pathways, we found 4 pathways, butanoate metabolism (hsa00650), hematopoietic cell lineage (hsa04640), lysine degradation (hsa00310) and basal transcription factors (hsa03022) related to POAG with permuted p < 0.001. In addition, the human leukocyte antigen (HLA) gene family was significantly associated with POAG (p < 0.001). In the POAG subset with normal-pressure glaucoma (NPG), the butanoate metabolism pathway was also significantly associated (p < 0.001) as well as the MAPK and Hedgehog signaling pathways (hsa04010 and hsa04340), glycosaminoglycan biosynthesis-heparan sulfate pathway (hsa00534) and the phenylalanine, tyrosine and tryptophan biosynthesis pathway (hsa0400). The butanoate metabolism pathway overall, and specifically the aspects of the pathway that contribute to GABA and acetyl-CoA metabolism, was the only pathway significantly associated with both POAG and NPG. Collectively these results implicate GABA and acetyl-CoA metabolism in glaucoma pathogenesis, and suggest new potential therapeutic targets.

PURPOSE: To investigate gold nanorods (GNRs) as a contrast agent to enhance Doppler optical coherence tomography (OCT) imaging of the intrascleral aqueous humor outflow. METHODS: A serial dilution of GNRs was scanned with a spectral-domain OCT device (Bioptigen, Durham, NC) to visualize Doppler signal. Doppler measurements using GNRs were validated using a controlled flow system. To demonstrate an application of GNR enhanced Doppler, porcine eyes were perfused at constant pressure with mock aqueous alone or 1.0x10(12) GNR/mL mixed with mock aqueous. Twelve Doppler and volumetric SD-OCT scans were obtained from the limbus in a radial fashion incremented by 30 degrees , forming a circular scan pattern. Volumetric flow was computed by integrating flow inside non-connected vessels throughout all 12 scans around the limbus. RESULTS: At the GNR concentration of 0.7x10(12) GNRs/mL, Doppler signal was present through the entire depth of the testing tube without substantial attenuation. A well-defined laminar flow profile was observed for Doppler images of GNRs flowing through the glass capillary tube. The Doppler OCT measured flow profile was not statistically different from the expected flow profile based upon an autoregressive moving average model, with an error of -0.025 to 0.037 mm/s (p = 0.6435). Cross-sectional slices demonstrated the ability to view anterior chamber outflow ex-vivo using GNR-enhanced Doppler OCT. Doppler volumetric flow measurements were comparable to flow recorded by the perfusion system. CONCLUSIONS: GNRs created a measureable Doppler signal within otherwise silent flow fields in OCT Doppler scans. Practical application of this technique was confirmed in a constant pressure ex-vivo aqueous humor outflow model in porcine eyes.

PURPOSE: To determine the reproducibility of automated segmentation of the three-dimensional (3D) lamina cribrosa (LC) microarchitecture scanned in-vivo using optical coherence tomography (OCT). METHODS: Thirty-nine eyes (8 healthy, 19 glaucoma suspects and 12 glaucoma) from 49 subjects were scanned twice using swept-source (SS-) OCT in a 3.5x3.5x3.64 mm (400x400x896 pixels) volume centered on the optic nerve head, with the focus readjusted after each scan. The LC was automatically segmented and analyzed for microarchitectural parameters, including pore diameter, pore diameter standard deviation (SD), pore aspect ratio, pore area, beam thickness, beam thickness SD, and beam thickness to pore diameter ratio. Reproducibility of the parameters was assessed by computing the imprecision of the parameters between the scans. RESULTS: The automated segmentation demonstrated excellent reproducibility. All LC microarchitecture parameters had an imprecision of less or equal to 4.2%. There was little variability in imprecision with respect to diagnostic category, although the method tends to show higher imprecision amongst healthy subjects. CONCLUSION: The proposed automated segmentation of the LC demonstrated high reproducibility for 3D LC parameters. This segmentation analysis tool will be useful for in-vivo studies of the LC.

PURPOSE: To develop and characterize a mouse model with intraocular pressure (IOP) elevation after laser photocoagulation on the trabecular meshwork (TM), which may serve as a model to investigate the potential of stem cell-based therapies for glaucoma. METHODS: IOP was measured in 281 adult C57BL/6 mice to determine normal IOP range. IOP elevation was induced unilaterally in 50 adult mice, by targeting the TM through the limbus with a 532-nm diode laser. IOP was measured up to 24 weeks post-treatment. The optic nerve damage was detected by electroretinography and assessed by semiautomatic counting of optic nerve axons. Effects of laser treatment on the TM were evaluated by histology, immunofluorescence staining, optical coherence tomography (OCT) and transmission electron microscopy (TEM). RESULTS: The average IOP of C57BL/6 mice was 14.5 +/- 2.6 mmHg (Mean +/- SD). After laser treatment, IOP averaged above 20 mmHg throughout the follow-up period of 24 weeks. At 24 weeks, 57% of treated eyes had elevated IOP with the mean IOP of 22.5 +/- 2.5 mmHg (Mean +/- SED). The difference of average axon count (59.0%) between laser treated and untreated eyes was statistically significant. Photopic negative response (PhNR) by electroretinography was significantly decreased. CD45+ inflammatory cells invaded the TM within 1 week. The expression of SPARC was increased in the TM from 1 to 12 weeks. Histology showed the anterior chamber angle open after laser treatment. OCT indicated that most of the eyes with laser treatment had no synechia in the anterior chamber angles. TEM demonstrated disorganized and compacted extracellular matrix in the TM. CONCLUSIONS: An experimental murine ocular hypertension model with an open angle and optic nerve axon loss was produced with laser photocoagulation, which could be used to investigate stem cell-based therapies for restoration of the outflow pathway integrity for ocular hypertension or glaucoma.

PURPOSE: The CAV1/CAV2 (caveolin 1 and caveolin 2) genomic region previously was associated with primary open-angle glaucoma (POAG), although replication among independent studies has been variable. The aim of this study was to assess the association between CAV1/CAV2 single nucleotide polymorphisms (SNPs) and POAG in a large case-control dataset and to explore associations by gender and pattern of visual field (VF) loss further. DESIGN: Case-control study. PARTICIPANTS: We analyzed 2 large POAG data sets: the Glaucoma Genes and Environment (GLAUGEN) study (976 cases, 1140 controls) and the National Eye Institute Glaucoma Human Genetics Collaboration (NEIGHBOR) consortium (2132 cases, 2290 controls). METHODS: We studied the association between 70 SNPs located within the CAV1/CAV2 genomic region in the GLAUGEN and NEIGHBOR studies, both genotyped on the Illumina Human 660WQuadv1C BeadChip array and imputed with the Markov Chain Haplotyping algorithm using the HapMap 3 reference panel. We used logistic regression models of POAG in the overall population and separated by gender, as well as by POAG subtypes defined by type of VF defect (peripheral or paracentral). Results from GLAUGEN and NEIGHBOR were meta-analyzed, and a Bonferroni-corrected significance level of 7.7 x 10(-4) was used to account for multiple comparisons. MAIN OUTCOME MEASURES: Overall POAG, overall POAG by gender, and POAG subtypes defined by pattern of early VF loss. RESULTS: We found significant associations between 10 CAV1/CAV2 SNPs and POAG (top SNP, rs4236601; pooled P = 2.61 x 10(-7)). Of these, 9 were significant only in women (top SNP, rs4236601; pooled P = 1.59 x 10(-5)). Five of the 10 CAV1/CAV2 SNPs were associated with POAG with early paracentral VF (top SNP, rs17588172; pooled P = 1.07 x 10(-4)), and none of the 10 were associated with POAG with peripheral VF loss only or POAG among men. CONCLUSIONS: CAV1/CAV2 SNPs were associated significantly with POAG overall, particularly among women. Furthermore, we found an association between CAV1/CAV2 SNPs and POAG with paracentral VF defects. These data support a role for caveolin 1, caveolin 2, or both in POAG and suggest that the caveolins particularly may affect POAG pathogenesis in women and in patients with early paracentral VF defects.

PURPOSE: To create an in vivo model of vector-mediated trabecular meshwork (TM) ablation and replacement. METHODS: We generated a conditionally cytotoxic, trackable vector, HSVtkiG, that expressed herpes simplex virus 1 thymidine kinase (HSVtk) and enhanced green fluorescent protein (eGFP). We optimized HSVtkiG ablation in vitro with ganciclovir (GCV) in comparison to eGFP control vector GINSIN and investigated the mechanism. Right eyes of 24 rats were then injected intracamerally with either HSVtkiG or GINSIN, before intraperitoneal GCV was administered 1 week later. Intraocular pressure, central corneal thickness (CCT), and slit-lamp exams were assessed for 8 weeks. Transduction and ablation were followed by gonioscopic visualization of eGFP. Histology was obtained with TM cell counts and immunohistochemistry markers of inflammation. RESULTS: Transduction and ablation parameters were established in vitro. Apoptosis was the cause of cell death. In vivo, transduction was seen gonioscopically to be targeted to the TM, followed by disappearance of eGFP marker fluorescence in HSVtkiG-transduced cells after injection of GCV. Ablation resulted in an IOP decrease of 25% in HSVtkiG-injected eyes 2 days after GCV but not in GINSIN or noninjected control eyes (P < 0.05). Trabecular meshwork cellularity was decreased at the time of lowest IOP and recovered thereafter, while CCT remained unchanged. Inflammation was absent. CONCLUSIONS: A vector-based system for inducible ablation of cells of the outflow tract was developed. Trabecular meshwork ablation lowered IOP, and recovery of cellularity and IOP followed. This model may be useful to study pressure regulation by the TM, its stem cells, and migration patterns.

IMPORTANCE: Monitoring young children with optic pathway gliomas (OPGs) for visual deterioration can be difficult owing to age-related noncompliance. Optical coherence tomography (OCT) measures of retinal nerve fiber layer (RNFL) thickness have been proposed as a surrogate marker of vision but this technique is also limited by patient cooperation. OBJECTIVE: To determine whether measures of circumpapillary RNFL thickness, acquired with handheld OCT (HH-OCT) during sedation, can differentiate between young children with and without vision loss from OPGs. DESIGN, SETTING, AND PARTICIPANTS: This cross-sectional analysis of a prospective observational study was conducted at a tertiary-care children's hospital. Children with an OPG (sporadic or secondary to neurofibromatosis type 1) who were cooperative for visual acuity testing, but required sedation to complete magnetic resonance imaging, underwent HH-OCT imaging of the circumpapillary RNFL while sedated. MAIN OUTCOMES AND MEASURES: Area under the curve of the receiver operating characteristic, sensitivity, specificity, positive predictive value, and negative predictive value of the average and quadrant-specific RNFL thicknesses. RESULTS: Thirty-three children (64 eyes) met inclusion criteria (median age, 4.8 years; range, 1.8-12.6 years). In children with vision loss (abnormal visual acuity and/or visual field), RNFL thickness was decreased in all quadrants compared with the normal-vision group (P < .001 for all comparisons). Using abnormal criteria of less than 5% and less than 1%, the area under the curve was highest for the average RNFL thickness (0.96 and 0.97, respectively) compared with specific anatomic quadrants. The highest discrimination and predictive values were demonstrated for participants with 2 or more quadrants meeting less than 5% (sensitivity = 93.3; specificity = 97.9; positive predictive value = 93.3; and negative predictive value = 97.9) and less than 1% (sensitivity = 93.3; specificity = 100; positive predictive value = 100; and negative predictive value = 98.0) criteria. CONCLUSIONS AND RELEVANCE: Measures of RNFL thickness acquired with HH-OCT during sedation can differentiate between young children with and without vision loss from OPGs. For young children who do not cooperate with vision testing, HH-OCT measures may be a surrogate marker of vision. Longitudinal studies are needed to delineate the temporal relationship between RNFL decline and vision loss.

PURPOSE: To improve the diagnosis of glaucoma by combining time-domain optical coherence tomography (TD-OCT) measurements of the optic disc, circumpapillary retinal nerve fiber layer (RNFL), and macular retinal thickness. PATIENTS AND METHODS: Ninety-six age-matched normal and 96 perimetric glaucoma participants were included in this observational, cross-sectional study. Or-logic, support vector machine, relevance vector machine, and linear discrimination function were used to analyze the performances of combined TD-OCT diagnostic variables. RESULTS: The area under the receiver-operating curve (AROC) was used to evaluate the diagnostic accuracy and to compare the diagnostic performance of single and combined anatomic variables. The best RNFL thickness variables were the inferior (AROC=0.900), overall (AROC=0.892), and superior quadrants (AROC=0.850). The best optic disc variables were horizontal integrated rim width (AROC=0.909), vertical integrated rim area (AROC=0.908), and cup/disc vertical ratio (AROC=0.890). All macular retinal thickness variables had AROCs of 0.829 or less. Combining the top 3 RNFL and optic disc variables in optimizing glaucoma diagnosis, support vector machine had the highest AROC, 0.954, followed by or-logic (AROC=0.946), linear discrimination function (AROC=0.946), and relevance vector machine (AROC=0.943). All combination diagnostic variables had significantly larger AROCs than any single diagnostic variable. There are no significant differences among the combination diagnostic indices. CONCLUSIONS: With TD-OCT, RNFL and optic disc variables had better diagnostic accuracy than macular retinal variables. Combining top RNFL and optic disc variables significantly improved diagnostic performance. Clinically, or-logic classification was the most practical analytical tool with sufficient accuracy to diagnose early glaucoma.

PURPOSE: To compare three methods of Schlemm's canal (SC) cross-sectional area (CSA) measurement. METHODS: Ten eyes (10 healthy volunteers) were imaged three times using spectral-domain optical coherence tomography (Cirrus HD-OCT, Zeiss, Dublin, California, USA). Aqueous outflow vascular structures and SC collector channel ostia were used as landmarks to identify a reference location within the limbus. SC CSA was assessed within a 1 mm segment (+/-15 frames of the reference, 31 frames in all) by three techniques. (1) Using a random number table, SC CSA in five random frames from the set of 31 surrounding the reference were measured and averaged. (2) The most easily visualised SC location (subjective) was measured, and (3) SC CSA was measured in all 31 consecutive B-scans, and averaged. (comprehensive average, gold standard). Subjective and random CSAs were compared with the comprehensive by general estimating equation modelling, and structural equation modelling quantified agreement. RESULTS: The average from five random locations (4175+/-1045 microm(2)) was not significantly different than that obtained from the gold standard comprehensive assessment (4064+/-1308 microm(2), p=0.6537). Subjectively located SC CSA (7614+/-2162 microm(2)) was significantly larger than the comprehensive gold standard SC CSA (p<0.0001). The average of five random frames produced significantly less bias than did subjective location, yielding a calibration line crossing the 'no-bias' line. DISCUSSION: Subjectively located SC CSA measurements produce high estimates of SC CSA. SC assessed by measuring five random locations estimate CSA was similar to the gold standard estimate.