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Saray Tabak, Elie Beit-Yannai, Sofia Schreiber-Avissar; Exosomes Dose Response Effects on non-Pigmented ciliary Epithelium & Trabecular Meshwork Communication. Invest. Ophthalmol. Vis. Sci. 2018;59(9):3508.
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Exosomes are a major constituent of the aqueous humor, which participate in the communication between the non-pigmented ciliary epithelium (NPCE) and the trabecular meshwork (TM) tissues. Exosomes are small lipid bi-layer membranous vesicles, 40-150nm in diameter, secreted by most cell types in culture, and play a role in intercellular communication, allowing exchange of nucleotides, proteins and lipids between the exosome-producing cells and target cells. Limited data was published regarding the role of exosomes in the ocular drainage system in health and glaucoma disease. The required amount of exosomes needed in order to mimic physiological condition is unclear.Our purpose was to investigate the effect of various exosomes concentrations derived from NPCE on the expression of signaling molecules participating in the communication between NPCE and TM cells, via the ocular aqueous humor, by tracking the Wnt signaling pathway in TM cells.
Exosomes were isolated by PEG 8,000 method and concentrations were determined by Tunable Resistive Pulse Sensing (TRPS) method. Various exosomes concentrations were incubated with TM cells for different times. Uptake of NPCE derived exosomes by TM was measured by Image Stream analysis. mRNA levels, protein expression and MMP’s activity were determined using RT-PCR, Western blot and Gel Zymography respectively.
NPCE exosomes exhibit a time dependent uptake by TM cells at high concentration and higher accumulation of NPCE exosomes, suggesting specific communication between the two cells. A tendency of decrease was found in β-catenin, Axin2 and LEF1 mRNA levels in low concentrations compared to high ones. In the protein level, low exosomes concentration was associated with significant decreased expression of pGSK3β, while high ones diminished this effect. Moreover, a bimodal effect was documented in response to higher concentrations. In addition, a dose dependent increase in the activity of proMMP-9, MMP-9 and MMP-2 was found in TM cells treated with NPCE exosomes.
Using different exosomes concentrations, led to a dose response effect in the level of protein, gene expression and MMP’s activity. Hence, our findings suggest that exosomes concentration plays a role in the NPCE-TM communication in-vitro.
This is an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.
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