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±11nm in the unprocessed AH. Data show that centrifugation does not significantly affect the mean particle size (121±11nm versus 124±21nm), 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.

Characteristics of exosomes andmicroparticles discovered in human tears

“…Экзосомы не только обеспечивают адресную доставку молекулярных сигналов клеткам-реципиентам, но и несут уникальные маркеры клетки-продуцента, что делает их многообещающим субстратом в диагностических исследованиях, в первую очередь, с использованием их малых РНК и белков [3,26]. Недавние исследования экзосом водянистой влаги глаза выявили около 10 различных микроРНК, связанных с патогенезом глаукомы [27]. Эти результаты позволяют предполагать, что детальное исследование экзосом СЖ будет полезным для понимания патогенеза офтальмологических заболеваний, в частности, связанных с передней поверхностью глаза.…”

Literature review of baseline information to support the risk assessment of RNAi‐based GM plants

“…In fact, the bulk of the circulating RNA literature concerns biomarker potential of circulating miRNAs, which is of high clinical relevance but outside the scope of this report. Extracellular miRNAs were identified in a broad range of biological fluids (Pegtel et al, 2011;Huang et al, 2013;Kropp et al, 2014;Baglio et al, 2015;Dismuke et al, 2015;Wu et al, 2015). Apart from plasma, circulating miRNAs in milk are also intensely studied (Izumi et al, 2012;Kuruppath et al, 2013;Sun et al, 2013;Alsaweed et al, 2016c, b, a;Benmoussa et al, 2016;Xi et al, 2016).…”

“…The same year, placental miRNAs were found circulating in maternal plasma , which was one of the discoveries leading to the notion that miRNAs could be a mobile regulating molecule (Iguchi et al, 2010) and that could even mediate transgenerational epigenetic heritance (Sharma, 2015) or be transmitted across species Buck et al, 2014). Since then, extracellular miRNAs were identified in a broad range of biological fluids, including plasma, aqueous humour, cerebrospinal fluid, nasal mucus, or milk (Pegtel et al, 2011;Huang et al, 2013b;Kropp et al, 2014;Baglio et al, 2015;Dismuke et al, 2015;Wu et al, 2015a). miRNAs were identified in the cargo of exosomes, membranous vesicles 40 to 100 nm in diameter, which are constitutively released by almost all cell type and are found essentially in every biological fluid (reviewed, for example, in (Rak, 2013;Yoon et al, 2014) However, extracellular miRNAs do not need to be necessarily encapsulated in extracellular vesicles, as two studies showed that 95-99% of extracellular miRNA are not in extracellular vesicles but associated with AGO proteins in serum and cell culture media (Arroyo et al, 2011;Turchinovich et al, 2011).…”

Therapeutic and diagnostic applications of extracellular vesicles

“…EVs have been isolated from e.g. urine [11], plasma [26], semen [25], nasal secretion [24], breast milk [221], the aqueous humor of eyes [222], cerebrospinal fluid [223], peritoneal fluid [224], bronchoalveolar lavage [225]. Depending on the respective disease for which the biomarker is being developed, an accessible biofluid should be considered in which the EVs of interest are likely the most concentrated and a liquid biopsy can be easily obtained.…”