Nimesh Bhikhu Patel

Reproducibility of Measuring Lamina Cribrosa Pore Geometry in Human and Nonhuman Primates with In Vivo Adaptive Optics Imaging

PURPOSE The ability to consistently resolve lamina cribrosa pores in vivo has applications in the study of optic nerve head and retinal disease mechanisms. Repeatability was assessed in imaging laminar pores in normal living eyes with a confocal adaptive optics scanning laser ophthalmoscope (AOSLO). METHODS Reflectance images (840 nm) of the anterior lamina cribrosa were acquired using the AOSLO in four or more different sessions in two normal rhesus monkey eyes and three normal human eyes. Laminar pore areas, elongations (ratio of major to minor axes of the best-fit ellipse) and nearest neighbor distances were calculated for each session. Measurement repeatability was assessed across sessions. RESULTS Pore areas ranged from 90 to 4365 μm(2) in monkeys and 154 to 6637 μm(2) in humans. Mean variabilities in measuring pore area and elongation (i.e., mean of the standard deviation of measurements made across sessions for the same pores) were 50 μm(2) (6.1%) and 0.13 (6.7%), respectively, in monkeys and 113 μm(2) (8.3%) and 0.17 (7.7%), respectively, in humans. Mean variabilities in measuring nearest neighbor distances were 1.93 μm (5.2%) in monkeys and 2.79 μm (4.1%) in humans. There were no statistically significant differences in any pore parameters across sessions (ANOVA, P > 0.05). CONCLUSIONS The anterior lamina cribrosa was consistently imaged in vivo in normal monkey and human eyes. The small intersession variability in normal pore geometry suggests that AOSLO imaging could be used to measure and track changes in laminar pores in vivo during glaucomatous progression.

Retinal Nerve Fiber Layer Assessment: Area versus Thickness Measurements from Elliptical Scans Centered on the Optic Nerve

PURPOSE An evaluation of the retinal nerve fiber layer (RNFL) provides important information on the health of the optic nerve. Standard measurements of the RNFL consider only thickness, but an accurate assessment should also consider axial length, size of the optic nerve head (ONH), blood vessel contribution, and distance of the scan from the ONH margin. In addition, although most primate ONHs are elliptical, the circular scan centered on the ONH is the mainstay in both clinical and research analyses. The purpose of this study was to evaluate thickness and area measures of RNFL cross sections when axial length and ONH shape are included. METHODS Circular, raster, and radial scans of left eye optic nerves were acquired from 40 normal rhesus monkeys (Macaca mulatta) using spectral domain optical coherence tomography. The disc margin was identified by manually selecting the RPE/Bruchs membrane opening and ONH border tissue. With a pixel-to-micrometer conversion computed from a three-surface schematic eye, RNFL scans were interpolated at 300 to 600 μm (50-μm increments) from the edge of the ONH. The thickness and area of the RNFL at each distance were obtained by custom programs. Blood vessels in the RNFL were selected and removed from the overall RNFL measures. RESULTS The average RNFL thickness decreased systematically from 149 ± 12.0 μm for scans 300 μm from the disc margin to 113 ± 7.2 μm at an eccentricity of 600 μm (P < 0.05). In contrast, the cross-sectional areas of the RNFL did not vary with scan location from the disc margin (0.85 ± 0.07 mm(2) at 300 μm compared with 0.86 ± 0.06 mm(2) at 600 μm). Blood vessels accounted for 9.3% of total RNFL thickness or area, but varied with retinal location. On average, 17.6% of the superior and 14.2% of the inferior RNFL was vascular, whereas blood vessels accounted for only 2.3% of areas of the temporal and nasal RNFL regions. CONCLUSIONS In nonhuman primates, with appropriate transverse scaling and ONH shape analysis, the cross-sectional area of the RNFL is independent of scan distance, up to 600 μm from the rim margin, indicating that the axonal composition changes little over this range. The results suggest that, with incorporation of transverse scaling, the RNFL cross-sectional area, rather than RNFL thickness, provides an accurate assessment of the retinal ganglion cell axonal content within the eye.

Age-Associated Changes in the Retinal Nerve Fiber Layer and Optic Nerve Head

PURPOSE Optical coherence tomography (OCT) measures of the retinal nerve fiber layer (RNFL) thickness and neuroretinal rim (NRR) parameters are often used as a surrogate for retinal ganglion cell content. The purpose of this study was to investigate the relationship between these morphological measures and the aging effects on these structures. METHODS One hundred thirteen healthy individuals, aged 19 to 76 years, with no prior history of retinal of optic nerve head pathology were recruited. A circumpapillary and radial OCT scan centered on the optic nerve head (ONH) was used for data analysis. Transverse scaling was calculated for each subject using measures from optical biometry. Custom algorithms were used for morphological analysis of the ONH NRR and RNFL that included quantification of major retinal vascular contribution. RESULTS There was a significant age-related loss of RNFL thickness (-0.23 μm/y, R(2) = 0.24, P < 0.01), major retinal vascular contribution (-0.03 μm/y, R(2) = 0.07, P = 0.01, neural rim volume (NRV, -0.004 mm(3)/y, R(2) = 0.15, P < 0.01), and minimum rim width (MRW, -1.77 μm/y, R(2) = 0.23, P < 0.01) before, and after, incorporating the Bruchs membrane opening size (sMRW, -1.86 μm/y, R(2) = 0.22, P < 0.01). When normalized, the rates of change for ONH NRR parameters (NRV, 0.69%/y and sMRW, 0.50%/y) exceeded that of RNFL thickness (0.19%/y, P < 0.01). CONCLUSIONS Although both RNFL and ONH NRR parameters contain axons of retinal ganglion cells, there are differences in age-related changes in these measures that should be considered in clinical application.

Toll-Like Receptor Expression and Activation in Mice with Experimental Dry Eye

PURPOSE To investigate the expression and/or function of toll-like receptors (TLRs) and antimicrobial peptides (AMPs) in dry eye inflammation. METHODS Experimental dry eye (EDE) was induced in C57BL/6 mice and TLR mRNA and protein expression were determined at the ocular surface and lacrimal gland. TLR agonist cocktail was applied to the ocular surface in untreated (UT), corneal scratched, and EDE mice. The corneal expression of cathelin-related antimicrobial peptide (CRAMP; human LL-37 orthologue), and mouse beta defensin (mBD)-3 and -4 (human BD-2 orthologue) was compared. LL-37, hBD-2, TLR4, 5, and TLR9 mRNA expression was examined in patients with dysfunctional tear syndrome (DTS) via conjunctival impression cytology. Murine central corneal thickness (CCT) and inflammatory cell recruitment into the stroma was determined by in vivo imaging. RESULTS EDE upregulated TLR2-4 and 9 mRNA expression in the palpebral conjunctiva and with the exception of TLR4, a similar expression, occurred in the corneal epithelium. TLR2 and 5 were upregulated in lacrimal gland and overall, there was a corresponding change in TLR protein. EDE decreased CRAMP mRNA and protein. hBD-2 and TLR9 expression were modulated in DTS subjects. Topical TLR agonist increased inflammatory cells recruitment and CCT in mice with a cornea scratch. In EDE, TLR agonist treatment downregulated corneal mBD-4 protein caused corneal epithelial loss, and stromal ulceration resulting in decreased CCT. CONCLUSIONS DTS modulates the expression of TLR and CRAMP and topical application of TLR agonists in EDE mice resulted in corneal epithelial loss and thinning. These results suggest that TLRs are involved in DTS inflammation.

Loss of the low-frequency component of the global-flash multifocal electroretinogram in primate eyes with experimental glaucoma.

PURPOSE To study relationships between glaucoma-sensitive components identified with frequency-domain analysis of global-flash multifocal electroretinogram (mfERG), regional retinal nerve fiber layer thickness (RNFLT), and local visual field sensitivity (VS). METHODS Eleven macaque monkeys, including four controls and seven with unilateral laser-induced trabecular meshwork scarification and ocular hypertension, were observed with optical coherence tomography (OCT), full-field light-adapted flash ERG, 103-hexagon global-flash mfERG (MFOFO), and static perimetry. The effects of experimental glaucoma on mfERG were assessed in the frequency domain. Relations between root mean square (RMS) amplitude of a glaucoma-sensitive frequency range and peripapillary RNFLT (standard 12° OCT circular scan), and between RMS amplitude and VS were studied. RESULTS Experimental glaucoma led to a dramatic and consistent power loss in the low-frequency (<25 Hz) band of mfERG. The RMS of this low-frequency component (LFC) correlated significantly with the regional RNFLT. The r(2) of linear fits was 0.39 (P < 0.001) for cross-sectional group data and 0.60 after correction for intersubject variability. The r(2) of linear fits for longitudinal data from individual animals was as high as 0.78 (P < 0.001). Local LFC RMS amplitude also correlated significantly with interpolated VS for hexagons. The r(2) for exponential fits of hexagon LFC RMS amplitudes (inner three rings) versus VS (dB) was 0.29 to 0.52 (P < 0.001) for the group and up to 0.95 in individuals. CONCLUSIONS The significant correlations between regional measures of global-flash mfERG, RNFLT, and VS suggest that LFC RMS amplitude provides a useful index for objective quantification of local RGC function and monitoring of early changes in glaucoma.

Persistent Asymmetric Optic Disc Swelling After Long-Duration Space Flight: Implications for Pathogenesis.

Background: Several ophthalmic findings including optic disc swelling, globe flattening and choroidal folds have been observed in astronauts following long-duration space flight. The authors now report asymmetric choroidal expansion, disc swelling and optic disc morphologic changes in a 45-year-old astronaut which occurred during long-duration space flight and persisted following his space mission. Methods: Case study of ocular findings in an astronaut documented during and after a long-duration space flight of approximately 6 months. Before, during and after his spaceflight, he underwent complete eye examination, including fundus photography, ultrasound, and optical coherence tomography. Results: We documented asymmetric choroidal expansion inflight that largely resolved by 30 days postflight, asymmetric disc swelling observed inflight that persisted for over 180 days postflight, asymmetric optic disc morphologic changes documented inflight by OCT that persisted for 630 days postflight and asymmetric globe flattening that began inflight and continued 660 days postflight. Lumbar puncture opening pressures obtained at 7 and 365 days post-mission were 22 and 16 cm H20 respectively. Conclusion: The persistent asymmetric findings noted above, coupled with the lumbar puncture opening pressures, suggest that prolonged microgravity exposure may have produced asymmetric pressure changes within the perioptic subarachnoid space.

The relationship between retinal nerve fiber layer thickness and optic nerve head neuroretinal rim tissue in glaucoma.

PURPOSE The purpose of this study was to determine the relationship between optical coherence tomography (OCT) measures of retinal nerve fiber layer (RNFL) and neuroretinal rim (NRR) in a nonhuman primate experimental glaucoma model, and in a population of clinical patients. METHODS For nonhuman primates, normative data were collected from 44 healthy monkeys, and nine animals with unilateral experimental glaucoma that were followed longitudinally. Cross-sectional human subjects data were collected from 89 healthy, 74 glaucoma suspects, and 104 glaucoma patients. Individualized transverse scaling for OCT scans was calculated using a schematic eye that incorporated optical ocular biometry. Custom algorithms were used to quantify RNFL thickness with and without vessels removed, scaled minimum rim width (sMRW), and neural rim volume (NRV). RESULTS For the experimental glaucoma group, NRR parameters showed the first changes with increased cumulative IOP. The data for both NRR and RNFL measures were best fit by an exponential rise model (NRV, R2=0.79, P<0.01, sMRW, R2=0.74, P<0.01). The major retinal vascular thickness contribution to the RNFL decreased (0.03 μm/μm, P<0.01) with RNFL loss, but the percent vascular contribution increased (-0.1%/μm, P<0.01) with disease progression. Overall, the findings for the cross-sectional human data were similar to those of the experimental model. CONCLUSIONS The findings illustrate a nonlinear relationship between NRR and RNFL measures and provide support for the use of multiple OCT scaled morphological measures for the diagnosis and management of primary open angle glaucoma in humans.

Relationship Between Juxtapapillary Choroidal Volume and Beta-Zone Parapapillary Atrophy in Eyes With and Without Primary Open-Angle Glaucoma

PURPOSE To evaluate whether quantity of choroidal tissue directly adjacent to the optic nerve differs between eyes with and without glaucoma and whether beta-zone parapapillary atrophy influences this relationship. DESIGN Prospective cohort study. METHODS Subjects were enrolled in a longitudinal, observational study at our institution. We studied 1 eye of 63 primary open-angle glaucoma (POAG), 30 ocular hypertension (OH), and 48 control subjects. Using optical coherence tomography enhanced depth imaging, we acquired 12 radial scans centered on the optic nerve head with 15 degrees of separation between scans. After images were enhanced, segmented, and corrected for ocular magnification, juxtapapillary choroidal volumetric parameters were calculated using raw thickness measurements and standard interpolation techniques. Juxtapapillary choroidal volume was then compared by diagnosis and by beta-zone parapapillary atrophy status. RESULTS Total juxtapapillary choroidal volume was significantly reduced in POAG vs OH and control eyes (1.057 vs 1.228 vs 1.255 μL, P = .04) and it was reduced in eyes with vs without beta-zone parapapillary atrophy (1.076 μL, n = 80 vs 1.306 μL, n = 61, P < .001). Juxtapapillary choroidal volume did not differ between POAG, OH, and control eyes when beta-zone parapapillary atrophy was absent, but juxtapapillary choroidal volume was significantly reduced in POAG vs control eyes when beta-zone parapapillary atrophy was present (0.957 vs 1.196 μL, P = .02). Furthermore, POAG eyes with beta-zone parapapillary atrophy had substantially lower juxtapapillary choroidal volume compared to POAG eyes without beta-zone parapapillary atrophy (0.957 vs 1.356 μL, P < .001). CONCLUSIONS The volume of choroid adjacent to the optic nerve was significantly reduced in POAG eyes when beta-zone parapapillary atrophy was present, suggesting that beta-zone parapapillary atrophy may be a biomarker for juxtapapillary choroidal atrophy and associated vascular compromise in POAG.

In Vivo Changes in Lamina Cribrosa Microarchitecture and Optic Nerve Head Structure in Early Experimental Glaucoma

The lamina cribrosa likely plays an important role in retinal ganglion cell axon injury in glaucoma. We sought to (1) better understand optic nerve head (ONH) structure and anterior lamina cribrosa surface (ALCS) microarchitecture between fellow eyes of living, normal non-human primates and (2) characterize the time-course of in vivo structural changes in the ONH, ALCS microarchitecture, and retinal nerve fiber layer thickness (RNFLT) in non-human primate eyes with early experimental glaucoma (EG). Spectral domain optical coherence tomography (SDOCT) images of the ONH were acquired cross-sectionally in six bilaterally normal rhesus monkeys, and before and approximately every two weeks after inducing unilateral EG in seven rhesus monkeys. ONH parameters and RNFLT were quantified from segmented SDOCT images. Mean ALCS pore area, elongation and nearest neighbor distance (NND) were quantified globally, in sectors and regionally from adaptive optics scanning laser ophthalmoscope images. In bilaterally normal monkeys, ONH parameters were similar between fellow eyes with few inter-eye differences in ALCS pore parameters. In EG monkeys, an increase in mean ALCS Depth (ALCSD) was the first structural change measured in 6 of 7 EG eyes. A decrease in mean minimum rim width (MRW) simultaneously accompanied this early change in 4 of 6 EG eyes and was the first structural change in the 7th EG eye. Mean ALCS pore parameters were among the first or second changes measured in 4 EG eyes. Mean ALCS pore area and NND increased in superotemporal and temporal sectors and in central and peripheral regions at the first time-point of change in ALCS pore geometry. RNFLT and/or mean ALCS radius of curvature were typically the last parameters to initially change. Survival analyses found mean ALCSD was the only parameter to significantly show an initial change prior to the first measured loss in RNFLT across EG eyes.

The Adenosine Receptor Antagonist, 7-Methylxanthine, Alters Emmetropizing Responses in Infant Macaques

Purpose Previous studies suggest that the adenosine receptor antagonist, 7-methylxanthine (7-MX), retards myopia progression. Our aim was to determine whether 7-MX alters the compensating refractive changes produced by defocus in rhesus monkeys. Methods Starting at age 3 weeks, monkeys were reared with −3 diopter (D; n = 10; 7-MX −3D/pl) or +3D (n = 6; 7-MX +3D/pl) spectacles over their treated eyes and zero-powered lenses over their fellow eyes. In addition, they were given 100 mg/kg of 7-MX orally twice daily throughout the lens-rearing period (age 147 ± 4 days). Comparison data were obtained from lens-reared controls (−3D/pl, n = 17; +3D/pl, n = 9) and normal monkeys (n = 37) maintained on a standard diet. Refractive status, corneal power, and axial dimensions were assessed biweekly. Results The −3D/pl and +3D/pl lens-reared controls developed compensating myopic (−2.10 ± 1.07 D) and hyperopic anisometropias (+1.86 ± 0.54 D), respectively. While the 7-MX +3D/pl monkeys developed hyperopic anisometropias (+1.79 ± 1.11 D) that were similar to those observed in +3D/pl controls, the 7-MX −3D/pl animals did not consistently exhibit compensating myopia in their treated eyes and were on average isometropic (+0.35 ± 1.96 D). The median refractive errors for both eyes of the 7-MX −3D/pl (+5.47 D and +4.38 D) and 7-MX +3D/pl (+5.28 and +3.84 D) monkeys were significantly more hyperopic than that for normal monkeys (+2.47 D). These 7-MX–induced hyperopic ametropias were associated with shorter vitreous chambers and thicker choroids. Conclusions In primates, 7-MX reduced the axial myopia produced by hyperopic defocus, augmented hyperopic shifts in response to myopic defocus, and induced hyperopia in control eyes. The results suggest that 7-MX has therapeutic potential in efforts to slow myopia progression.