Characterization of Leukocyte Crawling in the Living Human Retina Using Adaptive Optics Scanning Light Ophthalmoscopy

Purpose: Direct visualization of leukocytes within the retinal microvasculature is essential for investigating local immune responses. In this study, we characterize leukocyte crawling dynamics in the retinal vasculature of healthy volunteers and three patients with systemic diseases using adaptive optics scanning light ophthalmoscopy (AOSLO). Methods: Twenty-three healthy volunteers (age range, 20-75 years) and three patients with systemic diseases (diabetes, sickle cell disease, and syphilis) were imaged using a commercial AOSLO device. Consecutive phase-contrast AOSLO images of the temporal retina were acquired over 5 minutes at two to four preidentified regions of interest. Leukocyte crawling events were tracked manually in the 5-minute time-lapse video; vessel type, lumen diameter, speed, and pattern were assessed. Results: A total of 232 intraluminal leukocyte crawling events were identified in healthy volunteers. Crawling events were more frequently observed in venules (89.66%) than arterioles (9.92%). Lumen diameter with leukocyte crawling ranged from 10.2 to 67.8 µm; mean ± SD crawling speed was 0.3 ± 0.1 µm/s. Four distinct crawling patterns, with respect to direction of blood flow, were identified: with, against, backtrack, and across. No significant difference in crawling speed was found between venules and arterioles (P = 0.67) or among crawling patterns (P = 0.197). In addition to crawling, various leukocyte dynamics, including adhesions, reverse transmigration, and extravascular cell motilities, were observed in patients with systemic diseases. Conclusions: This study demonstrates the feasibility of utilizing AOSLO to noninvasively visualize leukocyte crawling in the living human eye. The ability to characterize leukocyte dynamics in their natural environment offers invaluable insight into the localized retinal immune status.