title: Human aqueous humor exosomes.
abstract: Aqueous humor (AH) is a dynamic intraocular fluid that supports the vitality of tissues that regulate intraocular pressure. We recently discovered that extracellular nanovesicles called exosomes are a major constituent of AH. Exosomes function in extracellular communication and contain proteins and small RNA. Our goal was to characterize the physical properties of AH exosomes and their exosomal RNA (esRNA) content. We isolated exosomes from human AH collected during cataract surgery from five patients using serial ultracentrifugation. We measured the size and concentration of AH exosomes in solution using nanoparticle tracking analysis. We found a single population of vesicles having a mean size of 121 ± 11 nm in the unprocessed AH. Data show that centrifugation does not significantly affect the mean particle size (121 ± 11 nm versus 124 ± 21 nm), but does impact the final number of exosomes in solution (87% loss from the unprocessed AH; n = 5). We extracted esRNA from the pooled human AH samples using miRCURY RNA isolation kit from Exiqon. The quality of extracted esRNA was evaluated using Agilent Bioanalyzer 2100 and was used to generate a sequencing library for small RNA sequencing with Illumina MiSeq sequencer. More than 10 different miRNAs were identified; abundant species included miR-486-5p, miR-204, and miR-184. We found that the majority of extracellular vesicles in the AH were in the exosome size range, suggesting that miRNAs housed within exosomes may function in communication between AH inflow and outflow tissues.
authors.affiliation: Department of Ophthalmology, Duke University Medical Center, Durham, NC, USA.
authors.forename: W Michael
authors.initials: WM
authors.lastname: Dismuke
authors.affiliation: Department of Ophthalmology, Duke University Medical Center, Durham, NC, USA.
authors.forename: Pratap
authors.initials: P
authors.lastname: Challa
authors.affiliation: Department of Ophthalmology, Duke University Medical Center, Durham, NC, USA.
authors.forename: Iris
authors.initials: I
authors.lastname: Navarro
authors.affiliation: Department of Ophthalmology, Duke University Medical Center, Durham, NC, USA; Department of Biomedical Engineering, Duke University Medical Center, Durham, NC, USA.
authors.forename: W Daniel
authors.initials: WD
authors.lastname: Stamer
authors.affiliation: Department of Cellular Biology and Anatomy, Medical College of Georgia, Georgia Regents University, Augusta, GA, USA; James and Jean Culver Vision Discovery Institute, Georgia Regents University, Augusta, GA, USA. Electronic address: yutliu@gru.edu.
authors.forename: Yutao
authors.initials: Y
authors.lastname: Liu
mesh_terms: D001082:Aqueous Humor / Q000166:cytology*; D055354:Exosomes / Q000235:genetics* / Q000648:ultrastructure*; D005260:Female; D006801:Humans; D008297:Male; D035683:MicroRNAs / Q000302:isolation & purification*; D014461:Ultracentrifugation
chemical_list: D035683:MicroRNAs
keywords: Aqueous humor; Exosomal RNA; Exosome; Glaucoma; miRNA