Abstract
Purpose :
Exosomes are small extracellular vesicles (EVs) that carry proteins, RNA, and DNA serving as a cellular communication system. Human tear film (TF) EVs have the potential to allow for further investigation of ocular diseases. We sought to determine the presence and properties of tear EVs in healthy volunteers to test whether tears contain EVs, and to determine the basic properties of tear EVs in healthy subjects.
Methods :
We collected tears from 6 healthy adults whose only ocular conditions were refractive error or a history of cataract surgery at least 3 months prior. Tear samples (right eye from each patient and paired left eye samples taken for direct comparison) were collected with Schirmer strips, submerged into 1mL PBS containing 0.1% azide, and stored frozen at -18C. For EV isolation, samples were thawed, and EVs were eluted from the strip overnight at 4oC in PBS. EVs were isolated by differential ultracentrifugation (1000xg/4oC/10min, 25000xg/4oC/30min) with a final wash and isolation step at 115,000xg/4oC/90min. The EVs were analyzed with ZetaView® ParticleMetrix nanoparticle tracking analysis, yielding the physical properties of the particles (size and concentration). Parameters were then compared between the right and left eyes using intraclass correlation coefficients (ICC).
Results :
The mean EV size was 179.0 +/- 21.7nm (range 152-201.3nm). The mean concentration was 5.78e7 +/- 4.16e7 nanoparticles/mL, with a range of 7.8e6 – 1.10e8 nanoparticles/mL. Data from the left eyes were not significantly different from that from the right eyes, obtained independently. There was a strong correlation for mean EV size (ICC=0.87) and EV concentration (ICC=0.77).
Conclusions :
Using a rigorous method of differential ultracentrifugation, we specifically isolated EVs from tears. The tears from the healthy individuals were varied amongst individuals but were in general agreement. The comparison of tear EVs from left and right eyes of an individual were similar, further suggesting the rigor and usefulness of our approach. Our data will serve as control values for further expansion of our investigation to clinical populations, where tear EVs parameters and composition may provide insights into ocular disease.
This abstract was presented at the 2023 ARVO Annual Meeting, held in New Orleans, LA, April 23-27, 2023.