Richard A. Bilonick

Characterization of Outer Retinal Morphology with High-Speed, Ultrahigh-Resolution Optical Coherence Tomography

PURPOSE To visualize, quantitatively assess, and interpret outer retinal morphology by using high-speed, ultrahigh-resolution (UHR) OCT. METHODS Retinal imaging was performed in the ophthalmic clinic in a cross-section of 43 normal subjects with a 3.5-microm, axial-resolution, high-speed, UHR OCT prototype instrument, using a radial scan pattern (24 images, 1500 axial scans). Outer retinal layers were automatically segmented and measured. High-definition imaging was performed with a 2.8-microm axial-resolution, high-speed, UHR OCT research prototype instrument, to visualize the finer features in the outer retina. RESULTS Quantitative maps of outer retinal layers showed clear differences between the cone-dominated fovea and the rod-dominated parafovea and perifovea, indicating that photoreceptor morphology can explain the appearance of the outer retina in high-speed, UHR OCT images. Finer, scattering bands were visualized in the outer retina using high-definition imaging and were interpreted by comparison to known anatomy. CONCLUSIONS High-speed UHR OCT enables quantification of scattering layers in the outer retina. An interpretation of these features is presented and supported by quantitative measurements in normal subjects and comparison with known anatomy. The thick scattering region of the outer retina previously attributed to the retinal pigment epithelium (RPE) is shown to consist of distinct scattering bands corresponding to the photoreceptor outer segment tips, RPE, and Bruchs membrane. These results may advance understanding of the outer retinal appearance in OCT images. The normative measurements may also aid in future investigations of outer retinal changes in age-related macular degeneration and other diseases.

Effects of Age on Optical Coherence Tomography Measurements of Healthy Retinal Nerve Fiber Layer, Macula, and Optic Nerve Head

PURPOSE To determine the effects of age on global and sectoral peripapillary retinal nerve fiber layer (RNFL), macular thicknesses, and optic nerve head (ONH) parameters in healthy subjects using optical coherence tomography (OCT). DESIGN Retrospective, cross-sectional observational study. PARTICIPANTS A total of 226 eyes from 124 healthy subjects were included. METHODS Healthy subjects were scanned using the Fast RNFL, Fast Macula, and Fast ONH scan patterns on a Stratus OCT (Carl Zeiss Meditec, Dublin, CA). All global and sectoral RNFL and macular parameters and global ONH parameters were modeled in terms of age using linear mixed effects models. Normalized slopes were also calculated by dividing the slopes by the mean value of the OCT parameter for interparameter comparison. MAIN OUTCOME MEASURES Slope of each OCT parameter across age. RESULTS All global and sectoral RNFL thickness parameters statistically significantly decreased with increasing age, except for the temporal quadrant and clock hours 8 to 10, which were not statistically different from a slope of zero. Highest absolute slopes were in the inferior and superior quadrant RNFL and clock hour 1 (superior nasal). Normalized slopes showed a similar rate in all sectors except for the temporal clock hours (8-10). All macular thickness parameters statistically significantly decreased with increasing age, except for the central fovea sector, which had a slight positive slope that was not statistically significant. The nasal outer sector had the greatest absolute slope. Normalized macular slope in the outer ring was similar to the normalized slopes in the RNFL. Normalized inner ring had shallower slope than the outer ring with a similar rate in all quadrants. Disc area remained nearly constant across the ages, but cup area increased and rim area decreased with age, both of which were statistically significant. CONCLUSIONS Global and regional changes caused by the effects of age on RNFL, macula, and ONH OCT measurements should be considered when assessing eyes over time. FINANCIAL DISCLOSURE(S) Proprietary or commercial disclosure may be found after the references.

Retinal nerve fibre layer thickness measurement reproducibility improved with spectral domain optical coherence tomography

Background/aims: To investigate retinal nerve fibre layer (RNFL) thickness measurement reproducibility using conventional time-domain optical coherence tomography (TD-OCT) and spectral-domain OCT (SD-OCT), and to evaluate two methods defining the optic nerve head (ONH) centring: Centred Each Time (CET) vs Centred Once (CO), in terms of RNFL thickness measurement variability on SD-OCT. Methods: Twenty-seven eyes (14 healthy subjects) had three circumpapillary scans with TD-OCT and three raster scans (three-dimensional or 3D image data) around ONH with SD-OCT. SD-OCT images were analysed in two ways: (1) CET: ONH centre was defined on each image separately and (2) CO: ONH centre was defined on one image and exported to other images after scan registration. After defining the ONH centre, a 3.4 mm diameter virtual circular OCT was resampled on SD-OCT images to mimic the conventional circumpapillary RNFL thickness measurements taken with TD-OCT. Results: CET and CO showed statistically significantly better reproducibility than TD-OCT except for 11:00 with CET. CET and CO methods showed similar reproducibility. Conclusions: SD-OCT 3D cube data generally showed better RNFL measurement reproducibility than TD-OCT. The choice of ONH centring methods did not affect RNFL measurement reproducibility.

Identification and Assessment of Schlemm's Canal by Spectral-Domain Optical Coherence Tomography

PURPOSE Measurements of human Schlemms 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.

Optical Coherence Tomography Scan Circle Location and Mean Retinal Nerve Fiber Layer Measurement Variability

PURPOSE To investigate the effect on optical coherence tomography (OCT) retinal nerve fiber layer (RNFL) thickness measurements of varying the standard 3.4-mm-diameter circle location. METHODS The optic nerve head (ONH) region of 17 eyes of 17 healthy subjects was imaged with high-speed, ultrahigh-resolution OCT (hsUHR-OCT; 501 x 180 axial scans covering a 6 x 6-mm area; scan time, 3.84 seconds) for a comprehensive sampling. This method allows for systematic simulation of the variable circle placement effect. RNFL thickness was measured on this three-dimensional dataset by using a custom-designed software program. RNFL thickness was resampled along a 3.4-mm-diameter circle centered on the ONH, then along 3.4-mm circles shifted horizontally (x-shift), vertically (y-shift) and diagonally up to +/-500 microm (at 100-microm intervals). Linear mixed-effects models were used to determine RNFL thickness as a function of the scan circle shift. A model for the distance between the two thickest measurements along the RNFL thickness circular profile (peak distance) was also calculated. RESULTS RNFL thickness tended to decrease with both positive and negative x- and y-shifts. The range of shifts that caused a decrease greater than the variability inherent to the commercial device was greater in both nasal and temporal quadrants than in the superior and inferior ones. The model for peak distance demonstrated that as the scan moves nasally, the RNFL peak distance increases, and as the circle moves temporally, the distance decreases. Vertical shifts had a minimal effect on peak distance. CONCLUSIONS The location of the OCT scan circle affects RNFL thickness measurements. Accurate registration of OCT scans is essential for measurement reproducibility and longitudinal examination (ClinicalTrials.gov number, NCT00286637).

Glaucoma discrimination of segmented cirrus spectral domain optical coherence tomography (SD-OCT) macular scans

Aims To evaluate the glaucoma discriminating ability of macular retinal layers as measured by spectral domain optical coherence tomography (SD-OCT). Methods Healthy, glaucoma suspect and glaucomatous subjects had a comprehensive ocular examination, visual field testing and SD-OCT imaging (Cirrus HD-OCT; Carl Zeiss Meditec, Dublin, California, USA) in the macular and optic nerve head regions. OCT macular scans were segmented into macular nerve fibre layer (mNFL), ganglion cell layer with inner plexiform layer (GCIP), ganglion cell complex (GCC) (composed of mNFL and GCIP), outer retinal complex and total retina. Glaucoma discriminating ability was assessed using the area under the receiver operator characteristic curve (AUC) for all macular parameters and mean circumpapillary retinal nerve fibre layer (cpRNFL). Results Analysis was performed on 51 healthy, 49 glaucoma suspect and 63 glaucomatous eyes. The median visual field MD was −2.21 dB (IQR: −6.92 to −0.35) for the glaucoma group, −0.32 dB (IQR: −1.22 to 0.73) for the suspect group and −0.18 dB (IQR: −0.92 to 0.71) for the healthy group. Highest age adjusted AUCs were found for average GCC and GCIP (AUC=0.901 and 0.900, respectively) and their sectoral measurements: infero-temporal (0.922 and 0.913), inferior (0.904 and 0.912) and supero-temporal (0.910 and 0.897). These values were similar to the discriminating ability of the mean cpRNFL (AUC=0.913). Comparison of these AUCs did not yield any statistically significant difference (all p>0.05). Conclusions SD-OCT GCIP and GCC measurements showed similar glaucoma diagnostic ability and were comparable with that of cpRNFL.

Modeling hepatic fibrosis in African American and Caucasian American patients with chronic hepatitis C virus infection

Assessment of histological stage is an integral part of disease management in patients infected with the hepatitis C virus (HCV). The aim of this study was to develop a model incorporating objective clinical and laboratory parameters to estimate the probability of severe fibrosis (i.e., Ishak fibrosis ≥ 3) in previously untreated African American (AA) and Caucasian American (CA) patients with HCV genotype 1. The Ishak fibrosis scores of 205 CA and 194 AA patients enrolled in the Viral Resistance to Antiviral Therapy of Chronic Hepatitis C study (Virahep‐C) were modeled using simple and multiple logistic regression. The model was then validated in an independent cohort of 461 previously untreated patients with HCV. The distribution of fibrosis scores was similar in the AA and CA patients as was the proportion of patients with severe fibrosis (35% vs. 39%, P = .47). After accounting for the number of portal areas in the biopsy, patient age, serum aspartate aminotransferase, alkaline phosphatase, and platelet count were independently associated with severe fibrosis in the overall cohort, and the relationship with fibrosis was similar in both the AA and CA subgroups. The area under the receiver operating characteristic curve (AUROC) of the Virahep‐C model (0.837) was significantly better than in other published models (P = .0003). The AUROC of the Virahep‐C model was 0.851 in the validation population. In conclusion, a model consisting of widely available clinical and laboratory features predicted severe hepatic fibrosis equally well in AA and CA patients with HCV genotype 1 and was superior to other published models. The excellent performance of the Virahep‐C model in an external validation cohort suggests the findings are replicable and potentially generalizable. (HEPATOLOGY 2006;44:925–935.)

Delayed processing of blood increases the frequency of activated CD11b+ CD15+ granulocytes which inhibit T cell function☆

We tested whether granulocytes, which contaminate PBMC isolates after prolonged blood storage at room temperature, are responsible for inhibited T cell function in aged blood. We extend previous observations by characterizing these contaminating granulocytes as CD11b+ CD15+ cells comparable to activated CD11b+ CD15+ granulocytes induced by incubation of blood with FMLP. Granulocyte contamination of PBMC was observed within 6-8 h after venipuncture and room temperature storage (2.3 fold increase), and increased 11.3-fold by 24-26 h in comparison to PBMC from fresh blood. Refrigerated 22-26 hour storage of blood exacerbated granulocyte contamination (84-fold increase). In contrast, granulocyte contamination was markedly reduced if blood was diluted in RPMI-1640 medium (3.9-fold increase) or PBS (1.8-fold increase) prior to 22-26 hour room temperature storage. Granulocyte contamination significantly correlated with reduced CD3zeta chain expression, a marker of T cell dysfunction. Correspondingly, T cell proliferation following PHA stimulation was significantly decreased in PBMC with contaminating granulocytes from aged blood (77% of control) or FMLP treated blood (44% of control). Minimizing granulocyte contamination in PBMC of aged blood by cell sorting, or by reducing granulocyte activation by diluting blood in PBS prior to storage, increased CD3zeta chain expression and increased T cell proliferation following stimulation. These data indicate that granulocytes inhibit T cell function in aged blood. Therefore, preventing granulocyte activation in blood specimens is critical to maintain optimal T cell function. This may be accomplished by limiting the time from venipuncture to PBMC isolation to <8 h and may be extended to 26 h by simply diluting blood in PBS prior to room temperature storage.

In vivo lamina cribrosa micro-architecture in healthy and glaucomatous eyes as assessed by optical coherence tomography.

PURPOSE The lamina cribrosa (LC) is a prime location of glaucomatous damage. The purpose of this study was to compare LC 3-dimensional micro-architecture between healthy and glaucomatous eyes in vivo by using optical coherence tomography (OCT). METHODS Sixty-eight eyes (19 healthy and 49 glaucomatous) from 47 subjects were scanned in a 3.5 × 3.5 × 3.64-mm volume (400 × 400 × 896 pixels) at the optic nerve head by using swept-source OCT. The LC micro-architecture parameters were measured on the visible LC by an automated segmentation algorithm. The LC parameters were compared to diagnosis and visual field mean deviation (VF MD) by using a linear mixed effects model accounting for age. RESULTS The average VF MD for the healthy and glaucomatous eyes was -0.50 ± 0.80 dB and -7.84 ± 8.75 dB, respectively. Beam thickness to pore diameter ratio (P = 0.04) and pore diameter standard deviation (P < 0.01) were increased in glaucomatous eyes. With worse MD, beam thickness to pore diameter ratio (P < 0.01), pore diameter standard deviation (P = 0.05), and beam thickness (P < 0.01) showed a statistically significant increase while pore diameter (P = 0.02) showed a significant decrease. There were no significant interactions between any of the parameters and age (all P > 0.05). CONCLUSIONS Glaucomatous micro-architecture changes in the LC, detected by OCT analysis, reflect beams remodeling and axonal loss leading to reduction in pore size and increased pore size variability.

Retinal Nerve Fiber Layer Thickness Measurement Comparability between Time Domain Optical Coherence Tomography (OCT) and Spectral Domain OCT

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.