Abstract
Non-invasive reflectance imaging of the human RPE cell mosaic is demonstrated using a modified confocal adaptive optics scanning light ophthalmoscope (AOSLO). The confocal circular aperture in front of the imaging detector was replaced with a combination of a circular aperture 4 to 16 Airy disks in diameter and an opaque filament, 1 or 3 Airy disks thick. This arrangement reveals the RPE cell mosaic by dramatically attenuating the light backscattered by the photoreceptors. The RPE cell mosaic was visualized in all 7 recruited subjects at multiple retinal locations with varying degrees of contrast and cross-talk from the photoreceptors. Various experimental settings were explored for improving the visualization of the RPE cell boundaries including: pinhole diameter, filament thickness, illumination and imaging pupil apodization, unmatched imaging and illumination focus, wavelength and polarization. None of these offered an obvious path for enhancing image contrast. The demonstrated implementation of dark-field AOSLO imaging using 790 nm light requires low light exposures relative to light safety standards and it is more comfortable for the subject than the traditional autofluorescence RPE imaging with visible light. Both these factors make RPE dark-field imaging appealing for studying mechanisms of eye disease, as well as a clinical tool for screening and monitoring disease progression.
Highlights
The retinal pigment epithelium (RPE) lies directly posterior to the photoreceptor layer, with apical processes enveloping the outer segments of rods and cones [1]
This proximity allows the RPE to phagocytize the photoreceptor outer segments, and assist in the turnover of visual pigments [2]. This homeostatic role of the RPE is essential to normal health [3], and diseased RPE has been implicated in the pathogenesis of age-related macular degeneration [4], diabetic retinopathy [5], Stargardt’s disease [6], Best’s disease [7], Leber’s congenital amaurosis [8] and retinitis pigmentosa [9]
We show experimental evidence that this can be achieved in subjects with normal retinal architecture with moderate success, using near-infrared light in a modified adaptive optics scanning light ophthalmoscope (AOSLO)
Summary
The retinal pigment epithelium (RPE) lies directly posterior to the photoreceptor layer, with apical processes enveloping the outer segments of rods and cones [1]. The use of adaptive optics scanning light ophthalmoscopes (AOSLOs), has allowed in vivo imaging of individual RPE cells in non-human primates and human volunteers using the intrinsic fluorescence of the lipofuscin, allowing for analyses previously possible only with histology [15,16]. Various imaging parameters including: pinhole diameter, filter thickness, illumination and imaging pupil apodization, unmatched imaging and illumination focus, wavelength and polarization, were varied in order to improve the visualization of the RPE cell boundaries This dark-field imaging technique [27] is validated against AOSLO autofluorescence in a subject free from eye disease, as well as confocal AOSLO reflectance in a subject with central serous retinopathy
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