September 2016
Volume 57, Issue 12
Open Access
ARVO Annual Meeting Abstract  |   September 2016
Alleviation of inflammation in human RPE cells by bromodomain inhibitors is independent of SIRT1
Author Affiliations & Notes
  • Maria Hytti
    Ophthalmology, University of Eastern Finland, Kuopio, Finland
    School of Pharmacy, University of Eastern Finland, Kuopio, Finland
  • Niina Piippo
    Ophthalmology, University of Eastern Finland, Kuopio, Finland
    School of Pharmacy, University of Eastern Finland, Kuopio, Finland
  • Eveliina Korhonen
    Ophthalmology, University of Eastern Finland, Kuopio, Finland
    School of Pharmacy, University of Eastern Finland, Kuopio, Finland
  • Paavo Honkakoski
    School of Pharmacy, University of Eastern Finland, Kuopio, Finland
  • Kai Kaarniranta
    Ophthalmology, University of Eastern Finland, Kuopio, Finland
    Ophthalmology, Kuopio University Hospital, Kuopio, Finland
  • Anu Kauppinen
    School of Pharmacy, University of Eastern Finland, Kuopio, Finland
    Ophthalmology, Kuopio University Hospital, Kuopio, Finland
  • Footnotes
    Commercial Relationships   Maria Hytti, None; Niina Piippo, None; Eveliina Korhonen, None; Paavo Honkakoski, None; Kai Kaarniranta, None; Anu Kauppinen, None
  • Footnotes
    Support  Finnish Cultural Foundation - Central and North Savo Regional Funds, Silmä- ja Kudospankkisäätiö, Sokeain ystävät ry, Alfred Kordelin Foundation, Academy of Finland, Kuopio University Hospital, Silmäsäätiö, and Emil Aaltonen Foundation
Investigative Ophthalmology & Visual Science September 2016, Vol.57, 5787. doi:
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    • Get Citation

      Maria Hytti, Niina Piippo, Eveliina Korhonen, Paavo Honkakoski, Kai Kaarniranta, Anu Kauppinen; Alleviation of inflammation in human RPE cells by bromodomain inhibitors is independent of SIRT1. Invest. Ophthalmol. Vis. Sci. 2016;57(12):5787.

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

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Abstract

Purpose : Inflammation is a key player in the development of age-related macular degeneration (AMD) and the regulation of inflammatory reactions in RPE cells is a potential treatment strategy. Recently, specific inhibitors of the bromodomains in human bromodomain and extracellular domain containing (BET) proteins have been shown to reduce NF-kB mediated inflammation. Here, we assess the effect of BET bromodomain inhibition on DNA damage-induced inflammation and cell death in ARPE-19 cells, a cellular model of the human RPE, and determine the role of SIRT1 upregulation in these effects.

Methods : ARPE-19 cells were cultured to full confluency in DMEM:F-12 (1:1) growth medium. Experiments were performed under serum-free conditions. Cells were pretreated with three different bromodomain-inhibitors, JQ-1, PFI-1 and iBET-151 before DNA damage was induced by treatment with etoposide. Levels of pro-inflammatory cytokines IL-6 and IL-8 were measured by ELISA. At the same time, cellular toxicity was assessed using both the lactase dehydrogenase (LDH) and the MTT assay. SIRT1 levels and acetylation status of p53 were determined by western blotting. SIRT1 was knocked-down with SIRT1-specific human siRNA and pro-inflammatory cytokine levels were determined after SIRT1 knockdown as well.

Results : DNA damage reduced cell viability as measured by the MTT assay to 65% of control values and increased the release LDH to the medium. The BET bromodomain inhibitors did not increase metabolic activity but did lower the release of LDH. Etoposide treatment also increased the IL-6 and IL-8 levels 3 – 20 fold over the control values. Inhibition of BET bromodomains with either of the studied inhibitors reduced the release of IL-6 and IL-8 significantly (p<0.001). BET inhibition also reduced p53 acetylation but did not increase the SIRT1 levels in etoposide-treated cells. Concordantly, SIRT1 knock-down by siRNA did not affect the BET bromodomain inhibitors’ ability to decrease inflammation.

Conclusions : BET bromodomain inhibition shows strong anti-inflammatory potential in ARPE-19 cells, which might make it a valuable strategy for the treatment of early AMD. These effects were not mediated by SIRT1. Further studies will be necessary to determine the precise mode of action of the bromodomain inhibitors and the extent of their beneficial effect as a treatment option for AMD.

This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.

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