Potential measurement error from vessel reflex and multiple light paths in dual-wavelength retinal oximetry.

PURPOSE: This study aims to characterize the dependence of measured retinal arterial and venous saturation on vessel diameter and central reflex in retinal oximetry, with an ultimate goal of identifying potential causes and suggesting approaches to improve measurement accuracy. METHODS: In 10 subjects, oxygen saturation, vessel diameter and optical density are obtained using Oxymap Analyzer software without diameter correction. Diameter dependence of saturation is characterized using linear regression between measured values of saturation and diameter. Occurrences of negative values of vessel optical densities (ODs) associated with central vessel reflex are acquired from Oxymap Analyzer. A conceptual model is used to calculate the ratio of optical densities (ODRs) according to retinal reflectance properties and single and double-pass light transmission across fixed path lengths. Model-predicted values are compared with measured oximetry values at different vessel diameters. RESULTS: Venous saturation shows an inverse relationship with vessel diameter (D) across subjects, with a mean slope of -0.180 (SE = 0.022) %/µm (20 < D < 180 µm) and a more rapid saturation increase at small vessel diameters reaching to over 80%. Arterial saturation yields smaller positive and negative slopes in individual subjects, with an average of -0.007 (SE = 0.021) %/µm (20 < D < 200 µm) across all subjects. Measurements where vessel brightness exceeds that of the retinal background result in negative values of optical density, causing an artifactual increase in saturation. Optimization of model reflectance values produces a good fit of the conceptual model to measured ODRs. CONCLUSION: Measurement artefacts in retinal oximetry are caused by strong central vessel reflections, and apparent diameter sensitivity may result from single and double-pass transmission in vessels. Improvement in correction for vessel diameter is indicated for arteries however further study is necessary for venous corrections.

Topical Review: Studies of Ocular Function and Disease Using Hyperspectral Imaging.

SIGNIFICANCE: Advances in imaging technology over the last two decades have produced significant innovations in medical imaging. Hyperspectral imaging (HSI) is one of these innovations, enabling powerful new imaging tools for clinical use and greater understanding of tissue optical properties and mechanisms underlying eye disease.Hyperspectral imaging is an important and rapidly growing area in medical imaging, making possible the concurrent collection of spectroscopic and spatial information that is usually obtained from separate optical recordings. In this review, we describe several mainstream techniques used in HSI, along with noteworthy advances in optical technology that enabled modern HSI techniques. Presented also are recent applications of HSI for basic and applied eye research, which include a novel method for assessing dry eye syndrome, clinical slit-lamp examination of corneal injury, measurement of blood oxygen saturation in retinal disease, molecular changes in macular degeneration, and detection of early stages of Alzheimer disease. The review also highlights work resulting from integration of HSI with other imaging tools such as optical coherence tomography and autofluorescence microscopy and discusses the adaptation of HSI for clinical work where eye motion is present. Here, we present the background and main findings from each of these reports along with specific references for additional details.

In Vivo Assessment of Retinal Biomarkers by Hyperspectral Imaging: Early Detection of Alzheimer's Disease.

A noninvasive and cost-effective means to detect preclinical Alzheimer's disease (AD) and monitor disease progression would be invaluable. The retina is a developmental extension of the brain and has been viewed as a window to evaluate AD-related pathology. Cross-sectional studies have shown structural changes in the retina of AD patients that include thinning of the retinal nerve-fiber layer and changes in retinal vasculature. However, such changes do not manifest in early stages of the disease nor are they specific biomarkers for AD. Described herein is the utilization of our retinal hyperspectral imaging (rHSI) technique as a biomarker for identification of AD-related early pathological changes in the retina. Specifically, this account concerns the translation of our rHSI technique from animal models to human AD subjects. The underlying principle is Rayleigh light scattering, which is expected from low-order Aß aggregates present in early pathology. Recruitment was restricted to AD subjects (N = 19) and age-matched controls, with no family history of AD (N = 16). To limit the influence of skin pigmentation, subjects were restricted to those with skin pigmentation values of 2-3 on the Fitzpatrick scale. The largest spectral deviation from control subjects, rHSI signature, was obtained at the MCI stage with MMSE scores ⩾22, suggesting higher sensitivity of this technique in early disease stages. The rHSI signature observed is unaffected by eye pathologies such as glaucoma and cataract. Age of the subjects minimally influenced the spectral signatures. The rHSI technique shows promise for detection of preclinical AD; it is conducted in a truly noninvasive manner, without application of an exogenous label, and is thus potentially suitable for population screening.

Early Detection of Amyloidopathy in Alzheimer's Mice by Hyperspectral Endoscopy.

PURPOSE: To describe a spectral imaging system for small animal studies based on noninvasive endoscopy of the retina, and to present time-resolved spectral changes from live Alzheimer's mice prior to cognitive decline, corroborating our previous in vitro findings. METHODS: Topical endoscope fundus imaging was modified to use a machine vision camera and tunable wavelength system for acquiring monochromatic images across the visible to near-infrared spectral range. Alzheimer's APP/PS1 mice and age-matched, wild-type mice were imaged monthly from months 3 through 8 to assess changes in the fundus reflection spectrum. Optical changes were fit to Rayleigh light scatter models as measures of amyloid aggregation. RESULTS: Good quality spectral images of the central retina were obtained. Short-wavelength reflectance from Alzheimer's mice retinae showed significant reduction over time compared to wild-type mice. Optical changes were consistent with an increase in Rayleigh light scattering in neural retina due to soluble Aß1-42 aggregates. The changes in light scatter showed a monotonic increase in soluble amyloid aggregates over a 6-month period, with significant build up occurring at 7 months. CONCLUSIONS: Hyperspectral imaging technique can be brought inexpensively to the study of retinal changes caused by Alzheimer's disease progression in live small animals. A similar previous finding of reduction in the light reflection over a range of wavelengths in isolated Alzheimer's mice retinae, was reproducible in the living Alzheimer's mice. The technique presented here has a potential for development as an early Alzheimer's retinal diagnostic test in humans, which will support the treatment outcome.

Hyperspectral interferometry: Sizing microscale surface features in the pine bark beetle.

A new method of interferometry employing a Fabry-Perot etalon model was used to locate and size microscale features on the surface of the pine bark beetle. Oscillations in the reflected light spectrum, caused by self-interference of light reflecting from surfaces of foreleg setae and spores on the elytrum, were recorded using white light hyperspectral microscopy. By making the assumption that pairs of reflecting surfaces produce an etalon effect, the distance between surfaces could be determined from the oscillation frequency. Low frequencies of less than 0.08 nm(-1) were observed in the spectrum below 700 nm while higher frequencies generally occupied wavelengths from 600 to 850 nm. In many cases, two frequencies appeared separately or in combination across the spectrum. The etalon model gave a mean spore size of 3.04 ± 1.27 µm and a seta diameter of 5.44 ± 2.88 µm. The tapering near the setae tip was detected as a lowering of frequency. Spatial fringes were observed together with spectral oscillations from surfaces on the exoskeleton at higher magnification. These signals were consistent with embedded multi-layer reflecting surfaces. Possible applications for hyperspectral interferometry include medical imaging, detection of spore loads in insects and other fungal carriers, wafer surface and subsurface inspection, nanoscale materials, biological surface analysis, and spectroscopy calibration. This is, to our knowledge, the first report of oscillations directly observed by microscopy in the reflected light spectra from Coleoptera, and the first demonstration of broadband hyperspectral interferometry using microscopy that does not employ an internal interferometer.

Glaucoma filtration surgery and retinal oxygen saturation.

PURPOSE: Glaucoma may involve disturbances in retinal oxygenation and blood flow. The purpose of this study was to measure the effect of glaucoma filtration surgery on retinal vessel oxygen saturation. METHODS: A noninvasive spectrophotometric retinal oximeter was used to measure hemoglobin oxygen saturation in retinal arterioles and venules before and after glaucoma filtration surgery. Twenty-five consecutive patients were recruited, and 19 had adequate image quality. Fourteen underwent trabeculectomy and five glaucoma tube surgery. Twelve had primary open-angle glaucoma and seven had exfoliative glaucoma. IOP decreased from 23 +/- 7 to 10 +/- 4 mm Hg (mean +/- SD, P = 0.0001). RESULTS: Oxygen saturation increased in retinal arterioles from 97% +/- 4% to 99% +/- 6% (n = 19; P = 0.046) after surgery and was unchanged in venules (63% +/- 5% before surgery and 64% +/- 6% after, P = 0.76). There were no significant changes in saturation in the fellow eyes (P > 0.60). The arteriovenous difference was 34% before and 36% after surgery (P = 0.35). CONCLUSIONS: Glaucoma filtration surgery had almost no effect on retinal vessel oxygen saturation.

Improved detection of the central reflex in retinal vessels using a generalized dual-gaussian model and robust hypothesis testing.

This updates an earlier publication by the authors describing a robust framework for detecting vasculature in noisy retinal fundus images. We improved the handling of the "central reflex" phenomenon in which a vessel has a "hollow" appearance. This is particularly pronounced in dual-wavelength images acquired at 570 and 600 nm for retinal oximetry. It is prominent in the 600 nm images that are sensitive to the blood oxygen content. Improved segmentation of these vessels is needed to improve oximetry. We show that the use of a generalized dual-Gaussian model for the vessel intensity profile instead of the Gaussian yields a significant improvement. Our method can account for variations in the strength of the central reflex, the relative contrast, width, orientation, scale, and imaging noise. It also enables the classification of regular and central reflex vessels. The proposed method yielded a sensitivity of 72% compared to 38% by the algorithm of Can et al., and 60% by the robust detection based on a single-Gaussian model. The specificity for the methods were 95%, 97%, and 98%, respectively.

Automatic identification of retinal arteries and veins from dual-wavelength images using structural and functional features.

This paper presents an automated method to identify arteries and veins in dual-wavelength retinal fundus images recorded at 570 and 600 nm. Dual-wavelength imaging provides both structural and functional features that can be exploited for identification. The processing begins with automated tracing of the vessels from the 570-nm image. The 600-nm image is registered to this image, and structural and functional features are computed for each vessel segment. We use the relative strength of the vessel central reflex as the structural feature. The central reflex phenomenon, caused by light reflection from vessel surfaces that are parallel to the incident light, is especially pronounced at longer wavelengths for arteries compared to veins. We use a dual-Gaussian to model the cross-sectional intensity profile of vessels. The model parameters are estimated using a robust M-estimator, and the relative strength of the central reflex is computed from these parameters. The functional feature exploits the fact that arterial blood is more oxygenated relative to that in veins. This motivates use of the ratio of the vessel optical densities (ODs) from images at oxygen-sensitive and oxygen-insensitive wavelengths (ODR = OD600/OD570) as a functional indicator. Finally, the structural and functional features are combined in a classifier to identify the type of the vessel. We experimented with four different classifiers and the best result was given by a support vector machine (SVM) classifier. With the SVM classifier, the proposed algorithm achieved true positive rates of 97% for the arteries and 90% for the veins, when applied to a set of 251 vessel segments obtained from 25 dual wavelength images. The ability to identify the vessel type is useful in applications such as automated retinal vessel oximetry and automated analysis of vascular changes without manual intervention.

Algorithms for automated oximetry along the retinal vascular tree from dual-wavelength fundus images.

We present an automated method to perform accurate, rapid, and objective measurement of the blood oxygen saturation over each segment of the retinal vascular hierarchy from dual-wavelength fundus images. Its speed and automation (2 s per entire image versus 20 s per segment for manual methods) enables detailed level-by-level measurements over wider areas. An automated tracing algorithm is used to estimate vessel centerlines, thickness, directions, and locations of landmarks such as bifurcations and crossover points. The hierarchical structure of the vascular network is recovered from the trace fragments and landmarks by a novel algorithm. Optical densities (OD) are measured from vascular segments using the minimum reflected intensities inside and outside the vessel. The OD ratio (ODR=OD600/OD570) bears an inverse relationship to systemic HbO2 saturation (SO2). The sensitivity for detecting saturation change when breathing air versus pure oxygen was calculated from the measurements made on six subjects and was found to be 0.0226 ODR units, which is in good agreement with previous manual measurements by the dual-wavelength technique, indicating the validity of the automation. A fully automated system for retinal vessel oximetry would prove useful to achieve early assessments of risk for progression of disease conditions associated with oxygen utilization.

Hyperspectral imaging for measurement of oxygen saturation in the optic nerve head.

PURPOSE: To evaluate a hyperspectral imaging technique for monitoring relative spatial changes in retinal oxygen saturation. METHODS: The optic nerve head (ONH) and overlying vessels in cynomolgus monkey eyes were imaged with a fundus camera attached to a hyperspectral imaging system. Images were acquired with inspiration of room air and pure oxygen and at controlled intraocular pressures (IOP) of 15 mm Hg (normal) and 60 mm Hg (sustained for up to 5 minutes). Changes in relative blood oxygen saturation in the vessels and ONH were assessed from reflectance spectra. Saturation maps were derived from contributions of oxygenated and deoxygenated hemoglobin spectral signatures extracted from hyperspectral images. The results obtained with hyperspectral imaging were compared with known experimental outcomes. RESULTS: Pure oxygen markedly increased oxygen saturation in veins. Increases in arteries and the ONH were smaller. The results obtained with hyperspectral image analysis agreed with known changes in oxygen saturation from breathing experiments. Raising IOP reduced saturation in all structures and resulted in profound desaturation of arteries. During sustained high IOP, a rebound in saturation was observed in the ONH. Spatial maps clearly showed the saturation changes in arteries, veins, and surrounding tissues. CONCLUSIONS: Hyperspectral imaging can be adapted to measure and map relative oxygen saturation in retinal structures and the ONH in nonhuman primate eyes.