June 2020
Volume 61, Issue 7
Free
ARVO Annual Meeting Abstract  |   June 2020
Circadian secretion of VEGF and PEDF in Polarized RPE
Author Affiliations & Notes
  • Sara Ann Sillik
    Ophthalmology and Vision Science, University of Arizona, Tucson, Arizona, United States
  • Rory Morrison-Colvin
    Ophthalmology and Vision Science, University of Arizona, Tucson, Arizona, United States
  • Nicole Congrove
    Ophthalmology and Vision Science, University of Arizona, Tucson, Arizona, United States
  • Brian S McKay
    Ophthalmology and Vision Science, University of Arizona, Tucson, Arizona, United States
  • Robert W Snyder
    SynderBiomedical, Tucson, Arizona, United States
  • Footnotes
    Commercial Relationships   Sara Sillik, None; Rory Morrison-Colvin, None; Nicole Congrove, None; Brian McKay, None; Robert Snyder, SnyderBiomedical (F)
  • Footnotes
    Support  NEI EY026544
Investigative Ophthalmology & Visual Science June 2020, Vol.61, 4135. doi:
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    • Get Citation

      Sara Ann Sillik, Rory Morrison-Colvin, Nicole Congrove, Brian S McKay, Robert W Snyder; Circadian secretion of VEGF and PEDF in Polarized RPE. Invest. Ophthalmol. Vis. Sci. 2020;61(7):4135.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract

Purpose : Most functions in the eye are under circadian control. A continuous perfusion system was used to establish a rhythm using circadian modulators, L-DOPA and dopamine, to evaluate whether VEGF and PEDF secretion from retinal pigment epithelium (RPE) change in a diurnal manner.

Methods : The perfusion system is designed to provide a continuous flow of media which alternates delivery of L-DOPA and dopamine in low tyrosine media for 12-hour intervals over a two-day period. Human RPE from induced pluripotent stem cells (iPSC) were plated onto transwell inserts. We used the transepithelial electrical resistance (TEER) measurements of the monolayers to assess monolayer integrity. Once the monolayers exhibited a tight barrier, they were placed into the perfusion system and exposed to alternating L-DOPA (1uM) then dopamine (1uM) in 12-hour intervals. Conditioned media samples from the effluent were collected every two hours throughout the run, and PEDF and VEGF were measured by ELISA. Cosinor analysis was used for statistical analysis.

Results : Examining the 48-hour time course shows that the secretion of PEDF apically had a periodicity of 26.0 hours (p = 0.03) whereas the basal periodicity was 20.0 hours (p = 0.07). The results demonstrated that apical PEDF rose in response to L-DOPA to maximum value (0.15 ng) at 24 hours and subsequently decreased to minimum value (0.06 ng) by hour 34 in response to dopamine. Basal PEDF started off increasing but dropped at hour 20 to its minimum value (0.01 ng) then began to increase again until hour 40 for its maximum (0.05 ng). The amount of VEGF expressed in the same 48-hour time course demonstrated unique periodicity of 26.0 hours for apical (p = 0.06). The release of VEGF from the basal side was below the limit of detection throughout the time course. Apical VEGF increased to max value (0.05 ng) at hour 26 and had a minimum value (0.006 ng) at hour 48. This variation of periodicity between apical and basal may be partially attributed to the difference in volumes of the apical and basal chambers of the perfusion.

Conclusions : The results suggest that VEGF and PEDF are expressed in a circadian manner in response to L-DOPA and dopamine. The RPE illustrate polarized release of both PEDF and VEGF in the time course. Future experiments will be centered around perfusing the iPSC cells with either only L-DOPA or dopamine.

This is a 2020 ARVO Annual Meeting abstract.

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