Diabetes is a systemic disease that affects many different tissues and organs, including the eye. In the retina, the corresponding disease is called diabetic retinopathy (DR), which can lead to blindness if untreated. Signs of DR are present in nearly all patients who have had type 1 diabetes for two decades1. The first stage of DR, non-proliferative DR (NPDR), is marked by gradual capillary dropout. Associated with NPDR are a set of clinically-observable changes in the microcirculation, which include microaneurysms (MAs), or small outpouchings of the capillary wall, and intraretinal microvascular abnormalities (IRMAs), which are focal disruptions of the capillary topology. These clinically-observable changes are noticeable mainly because they are larger in size than the surrounding capillaries. For example, fundus photography is a noninvasive method to detect MAs, visible as small dots. New dots appear and existing dots disappear over time2, presumably due to formation of new outpouchings and non-perfusion of existing outpouchings. However, using fundus photography, it is not possible to investigate the relationship of MAs to the surrounding capillaries. This is because it is difficult to image capillaries, due to their small size and low contrast. We have recently developed a technique based on an adaptive optics scanning laser ophthalmoscope (AOSLO) that can noninvasively generate detailed images of capillaries without the use of injected dyes; this method is based on the use of motion as an intrinsic contrast signal3-5 (Figure 1 and Supplemental Digital Content 1 (available online at [LWW insert link]), AOSLO video of capillaries in Figure 1, from visit 4, 300 out of 2400 frames shown in real time. There are currently several other methods for the noninvasive investigation of capillaries, based on fundus photography6, and phase variance optical coherence tomography7, 8. Although the AOSLO has been used to evaluate retinal blood flow9, 10, it is not yet established whether AOSLO imaging can detect capillary defects such as MAs or IRMAs. Figure 1 High resolution images of a live human retina, generated using an adaptive optics scanning laser ophthalmoscope (AOSLO). Both images were generated noninvasively, from the same AOSLO video. (A) This image was generated by averaging frames across the video. ... The gold standard for investigating human retinal capillaries is fluorescein angiography (FA). There are clear advantages for using FA to evaluate DR. These include the improved ability to detect MAs, as well as the ability to assess regions of capillary dropout or capillary remodeling. Studies using FA-based methods have shown increases in both the size of the FAZ as well as the areas of the perifoveal intercapillary areas11, 12. Fluorescein has also been used to investigate blood flow in humans; in diabetes, the speed of capillary blood flow is decreased13. FA has been combined with AOSLO imaging14, but thus far, this method has not been applied to study DR. Typically, FA is performed by injecting a bolus of fluorescein dye into a peripheral vein, and then imaging the passage of the dye using a fundus camera. However, administration of fluorescein dye is clinically indicated only in the later stages of DR. As with all invasive procedures, there is also a small risk of adverse side effects15. There have been many efforts to develop instruments to image the capillaries, in order to assess the early stages of DR, and to investigate other retinal diseases. In this paper, we establish that AOSLO can be used to detect both clinical and subclinical signs of DR in the retina, where clinical signs are retinal features which would be considered to be clinically significant (e.g. MAs and IRMAs), and subclinical signs are retinal features which would not be considered clinically significant by the current standards of care (e.g. capillary bends and cone photoreceptor spacing). We use the AOSLO to simultaneously assess photoreceptors, capillaries, and leukocytes.
