Faculty Bibliography

PURPOSE: To test the reproducibility of spectral-domain optical coherence tomography (SD-OCT) total retinal thickness (TRT) measurements in mice. METHODS: C57Bl/6 mice were anesthetized, and three repeated volumetric images were acquired in both eyes with SD-OCT (250 A-scans x 250 frames x 1024 samplings), centered on the optic nerve head (ONH). The mice were repositioned between scans. TRT was automatically measured within a sampling band of retinal thickness with radii of 55 to 70 pixels, centered on the ONH by using custom segmentation software. The first volumetric image acquired in a given eye was used to register the remaining two SD-OCT images by manually aligning the en face images with respect to rotation and linear translation. Linear mixed-effects models were fitted to global and quadrant thicknesses, taking into account the clustering between eyes, to assess imprecision (measurement reproducibility). RESULTS: Twenty-six eyes of 13 adult mice (age 13 weeks) were imaged. The mean global TRT across all eyes was 298.21 mum, with a mouse heterogeneity standard deviation (SD) of 4.88 mum (coefficient of variation [CV] = 0.016), an eye SD of 3.32 mum (CV = 0.011), and a device-related imprecision SD of 2.33 mum (CV = 0.008). The superior quadrant had the thickest mean TRT measurement (310.38 mum) and the highest (worst) imprecision SD (3.13 mum; CV = 0.010), and the inferior quadrant had the thinnest mean TRT (291.55 mum). The quadrant with the lowest (best) imprecision SD was in the nasal one (2.06 mum; CV = 0.007). CONCLUSIONS: Good reproducibility was observed for SD-OCT retinal thickness measurements in mice. SD-OCT may be useful for in vivo longitudinal studies in mice.

BACKGROUND AND OBJECTIVE: Digital pathology has thus far focused on producing digital images of glass microscope slides. Spectral domain optical coherence tomography (SD-OCT) can be used to directly scan tissue blocks to produce three-dimensional histology images, potentially bypassing glass slide workflow. MATERIALS AND METHODS: Formalin-fixed paraffin-embedded tissue blocks were scanned using SD-OCT and resulting images were compared with corresponding areas on microscope slides. RESULTS: Low-magnification features were recognizable, including tissue outlines, fat, vessels, and outlines of colonic mucosal crypts. Subtle textures that were suggestive of benign breast lobules and ovarian tumor features were also visible. Initial SD-OCT images lacked resolution and contrast relative to traditional microscopy, but the image content suggests that additional features of interest are present and may be revealed with improved SD-OCT resolution and more post-processing experience. Elucidation of three-dimensional histology and pathology are also future tasks. CONCLUSION: Eventual availability of diagnostic-quality three-dimensional histology would have a profound impact on anatomic pathology.

Three dimensional (3D) ophthalmic imaging using optical coherence tomography (OCT) has revolutionized assessment of the eye, the retina in particular. Recent technological improvements have made the acquisition of 3D-OCT datasets feasible. However, while volumetric data can improve disease diagnosis and follow-up, novel image analysis techniques are now necessary in order to process the dense 3D-OCT dataset. Fundamental software improvements include methods for correcting subject eye motion, segmenting structures or volumes of interest, extracting relevant data post hoc and signal averaging to improve delineation of retinal layers. In addition, innovative methods for image display, such as C-mode sectioning, provide a unique viewing perspective and may improve interpretation of OCT images of pathologic structures. While all of these methods are being developed, most remain in an immature state. This review describes the current status of 3D-OCT scanning and interpretation, and discusses the need for standardization of clinical protocols as well as the potential benefits of 3D-OCT scanning that could come when software methods for fully exploiting these rich datasets are available clinically. The implications of new image analysis approaches include improved reproducibility of measurements garnered from 3D-OCT, which may then help improve disease discrimination and progression detection. In addition, 3D-OCT offers the potential for preoperative surgical planning and intraoperative surgical guidance.

We demonstrate ultrahigh speed swept source/Fourier domain ophthalmic OCT imaging using a short cavity swept laser at 100,000 - 400,000 axial scan rates. Several design configurations illustrate tradeoffs in imaging speed, sensitivity, axial resolution, and imaging depth. Variable rate A/D optical clocking is used to acquire linear-in-k OCT fringe data at 100 kHz axial scan rate with 5.3 um axial resolution in tissue. Fixed rate sampling at 1 GSPS achieves a 7.5mm imaging range in tissue with 6.0 um axial resolution at 100 kHz axial scan rate. A 200 kHz axial scan rate with 5.3 um axial resolution over 4mm imaging range is achieved by buffering the laser sweep. Dual spot OCT using two parallel interferometers achieves 400 kHz axial scan rate, almost 2X faster than previous 1050 nm ophthalmic results and 20X faster than current commercial instruments. Superior sensitivity roll-off performance is shown. Imaging is demonstrated in the human retina and anterior segment. Wide field 12x12 mm data sets include the macula and optic nerve head. Small area, high density imaging shows individual cone photoreceptors. The 7.5 mm imaging range configuration can show the cornea, iris, and anterior lens in a single image. These improvements in imaging speed and depth range provide important advantages for ophthalmic imaging. The ability to rapidly acquire 3D-OCT data over a wide field of view promises to simplify examination protocols. The ability to image fine structures can provide detailed information on focal pathologies. The large imaging range and improved image penetration at 1050 m wavelengths promises to improve performance for instrumentation which images both the retina and anterior eye. These advantages suggest that swept source OCT at 1050 nm wavelengths will play an important role in future ophthalmic instrumentation.

PURPOSE: Measurements of human Schlemm's canal (SC) have been limited to histologic sections. The purpose of this study was to demonstrate noninvasive measurements of aqueous outflow (AO) structures in the human eye, examining regional variation in cross-sectional SC areas (on/off collector channel [CC] ostia [SC/CC] and nasal/temporal) in the eyes of living humans. METHODS: SC was imaged by spectral-domain optical coherence tomography with a 200-nm bandwidth light source. Both eyes of 21 healthy subjects and one glaucomatous eye of three subjects were imaged nasally and temporally. Contrast and magnification were adjusted to maximize visualization. Cross-sectional SC on and off SC/CC was traced three times by two independent masked observers using ImageJ (ImageJ 1.40g, http://rsb.info.nih.gov/ij/ Wayne Rasband, developer, National Institutes of Health, Bethesda, MD). The mean SC area was recorded. A linear mixed-effects model was used to analyze eye, nasal/temporal laterality, and SC area on or off SC/CC. RESULTS: SC area was significantly larger on SC/CCs than off (12,890 vs. 7,391 micorm(2), P < 0.0001) and was significantly larger on the nasal side than on the temporal (10,983 vs. 8,308 micorm(2), P = 0.009). SC areas were significantly smaller in glaucoma patients than in normal subjects, whether pooled (P = 0.0073) or grouped by on (P = 0.0215) or off (P = 0.0114) SC/CC. CONCLUSIONS: Aqueous outflow structures, including SC and CCs, can be noninvasively assessed in the human eye. These measurements will be useful in physiological studies of AO and will be clinically useful in the determination of the impact of glaucoma therapies on IOP as well as presurgical planning.

PURPOSE. To investigate the characteristics of a spectral-domain optical coherence tomography (SD-OCT) image phenomenon known as the mirror artifact, calculate its prevalence, analyze potential risk factors, measure severity, and correlate it to spherical equivalent and central visual acuity (VA). METHODS. OCT macular cube 512 x 128 scans taken between January 2008 and February 2009 at the New England Eye Center were analyzed for the presence of mirror artifacts. Artifact severity was determined by the degree of segmentation breakdown that it caused on the macular map. A retrospective review was conducted of the medical records of patients with artifacts and of a random control group without artifacts. RESULTS. Of 1592 patients, 9.3% (148 patients, 200 eyes) had scans that contained mirror artifacts. A significantly more myopic spherical equivalent (P < 0.001), worse VA (P < 0.001), longer axial lengths (P = 0.004), and higher proportions of moderate to high myopia (P < 0.001) were found in patients with mirror artifacts than in patients without artifacts. Worse VA was associated with increased artifact severity (P = 0.04). CONCLUSIONS. In all scans analyzed, a high prevalence of mirror artifacts was found. This image artifact was often associated with patients with moderate to high myopia. Improvements in instrumentation may be necessary to resolve this problem in moderately and highly myopic eyes. Operators should be advised to properly position the retina when scanning eyes. In cases in which peripheral abnormalities in topographic measurements of retinal thickness are found, corresponding OCT scans should be examined for the presence of mirror artifacts.

PURPOSE: Ethyl pyruvate (EP) has pharmacologic effects that remediate cellular stress. In the organ-cultured murine lens, EP ameliorates oxidative stress, and in a rat cataract model, it attenuates cataract formation. However, corneal responses to EP have not been elucidated. In this study, the potential of EP as a therapeutic agent in corneal wound healing was determined by examining its effects on the transition of quiescent corneal stromal keratocytes into contractile myofibroblasts. METHODS: Three independent preparations of cultured human keratocytes were treated with TGF-beta1, to elicit a phenotypic transition to myofibroblasts in the presence or absence of 10 or 15 mM EP. Gene expression profiles of the 12 samples (keratocytes +/- EP +/- TGF-beta1 for three preparations) were produced by using gene microarrays. RESULTS: TGF-beta1-driven twofold changes in at least two of three experiments defined a group of 1961 genes. Genes showing twofold modulation by EP in at least two experiments appeared exclusively in myofibroblasts (857 genes), exclusively in keratocytes (409 genes), or in both phenotypes (252 genes). Analysis of these three EP-modulated groups showed that EP (1) inhibited myofibroblast proliferation with concomitant modulation of some cell cycle genes, (2) augmented the NRF2-mediated antioxidant response in both keratocytes and myofibroblasts, and (3) modified the TGF-beta1-driven transition of keratocytes to myofibroblasts by inhibiting the upregulation of a subset of profibrotic genes. CONCLUSIONS: These EP-induced phenotypic changes in myofibroblasts indicate the potential of EP as a therapeutic agent in corneal wound healing.

PURPOSE: To study optic nerve head (ONH) topography parameters measured by Stratus optical coherence tomography (OCT) in normal subjects and to analyze ONH data for differences in relation to disc size, ethnicity, and age. METHODS: Three hundred sixty-seven normal subjects underwent Stratus optical coherence tomography ONH measurement using the fast optic disc scan protocol software package 3.0. Only ONH scans meeting specific qualification criteria were included for data analysis ensuring appropriate scan quality and reliability. ONH topographic parameters of qualified scans were analyzed for differences in regards to optic disc size, age, and ethnicity. RESULTS: Two hundred and twelve qualified ONH scans were included for data analysis. Mean disc area was 2.27+/-0.41 mm and optic cup area, rim area, and horizontal integrated rim width increased with disc size, whereas vertical integrated rim area did not. Vertical integrated rim area, horizontal integrated rim width, and rim area decreased and cup area increased with age. Mean optic disc area was larger in African-Americans as compared with Hispanics or Whites and this difference was statistically significant. CONCLUSIONS: Optic cup area, rim area, and horizontal integrated rim width correlated to disc size. Vertical integrated rim area, horizontal integrated rim width, rim area, and cup area, changed with age. African-American optic discs had larger disc area measurements as compared with Whites optic discs and this difference was statistically significant.

PURPOSE: To develop a fully automated algorithm (AP) to perform a volumetric measure of the optic disc using conventional stereoscopic optic nerve head (ONH) photographs, and to compare algorithm-produced parameters with manual photogrammetry (MP), scanning laser ophthalmoscope (SLO) and optical coherence tomography (OCT) measurements. METHODS: One hundred twenty-two stereoscopic optic disc photographs (61 subjects) were analyzed. Disc area, rim area, cup area, cup/disc area ratio, vertical cup/disc ratio, rim volume and cup volume were automatically computed by the algorithm. Latent variable measurement error models were used to assess measurement reproducibility for the four techniques. RESULTS: AP had better reproducibility for disc area and cup volume and worse reproducibility for cup/disc area ratio and vertical cup/disc ratio, when the measurements were compared to the MP, SLO and OCT methods. CONCLUSION: AP provides a useful technique for an objective quantitative assessment of 3D ONH structures.

PURPOSE: Time domain optical coherence tomography (TD-OCT) has been used commonly in clinical practice, producing a large inventory of circular scan data for retinal nerve fiber layer (RNFL) assessment. Spectral domain (SD)-OCT produces three-dimensional (3-D) data volumes. The purpose of this study was to create a robust technique that makes TD-OCT circular scan RNFL thickness measurements comparable with those from 3-D SD-OCT volumes. METHODS: Eleven eyes of 11 healthy subjects and 7 eyes of 7 subjects with glaucoma were enrolled. Each eye was scanned with one centered and eight displaced TD-OCT scanning circles. One 3-D SD-OCT cube scan was obtained at the same visit. The matching location of the TD-OCT scanning circle was automatically detected within the corresponding 3-D SD-OCT scan. Algorithm performance was assessed by estimating the difference between the detected scanning circle location on 3-D SD-OCT volume and the TD-OCT circle location. Global and sectoral RNFL thickness measurement errors between the two devices were also compared. RESULTS: The difference (95% confidence interval) in scanning circle center locations between TD- and SD-OCT was 2.3 (1.5-3.2) pixels (69.0 [45.0-96.0] microm on the retina) for healthy eyes and 3.1 (2.0-4.1) pixels (93.0 [60.0-123.0] microm on the retina) for glaucomatous eyes. The absolute RNFL thickness measurement difference was significantly smaller with the matched scanning circle. CONCLUSIONS: Scan location matching may bridge the gap in RNFL thickness measurements between TD-OCT circular scan data and 3-D SD-OCT scan data, providing follow-up comparability across the two generations of OCTs.