June 2013
Volume 54, Issue 15
Free
ARVO Annual Meeting Abstract  |   June 2013
Epigenetic Regulation of Eotaxin Expression in Human Retinal Pigment Epithelium
Author Affiliations & Notes
  • Mark Kleinman
    Ophthalmology & Visual Sci, Univ of Kentucky, Lexington, KY
  • Andre Berner
    Ophthalmology & Visual Sci, Univ of Kentucky, Lexington, KY
  • Dingyuan Lou
    Ophthalmology & Visual Sci, Univ of Kentucky, Lexington, KY
  • Kabhilan Mohan
    Ophthalmology & Visual Sci, Univ of Kentucky, Lexington, KY
  • Chad Jackson
    Ophthalmology & Visual Sci, Univ of Kentucky, Lexington, KY
  • Jennifer Brown
    Ophthalmology & Visual Sci, Univ of Kentucky, Lexington, KY
  • Jayakrishna Ambati
    Ophthalmology & Visual Sci, Univ of Kentucky, Lexington, KY
  • Footnotes
    Commercial Relationships Mark Kleinman, None; Andre Berner, None; Dingyuan Lou, None; Kabhilan Mohan, None; Chad Jackson, None; Jennifer Brown, University of Ky (E); Jayakrishna Ambati, None
  • Footnotes
    Support None
Investigative Ophthalmology & Visual Science June 2013, Vol.54, 344. doi:
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      Mark Kleinman, Andre Berner, Dingyuan Lou, Kabhilan Mohan, Chad Jackson, Jennifer Brown, Jayakrishna Ambati; Epigenetic Regulation of Eotaxin Expression in Human Retinal Pigment Epithelium. Invest. Ophthalmol. Vis. Sci. 2013;54(15):344.

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

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Abstract

Purpose: Age-related macular degeneration (AMD) is the most common cause of irreversible blindness in developed countries. Previous reports have identified increased eotaxin expression in retinal pigment epithelium (RPE) in both dry and neovascular AMD. New insights into epigenetic derangements and chromatin remodeling have garnered great interest as a potent biologic regulatory system that may serve as a critical aging mechanism. Histone deacetylases (HDAC) are integral components of this process and exhibit a unique control of the chromatin remodeling process. HDACs have become a novel target for the epigenetic modulation of important pathways in aging and neurodegenerative diseases including AMD. Here, we show that eotaxin expression is regulated by HDAC-1 and demonstrate its interaction with the promoter region of eotaxin-3.

Methods: Primary human RPE (hRPE) isolates (n=3, single donor) were treated with either class I/II HDAC inhibitors (valproic acid (VPA), trichostatin A (TSA) and suberoylanilide hydroxamic acid (SAHA) or a class III HDAC inhibitor, nicotinamide (NIC). Eotaxin 1/2/3 expression levels were measured at 24 hours via real-time PCR. Culture supernatant was analyzed with multiplex bead arrays (Luminex) at 48 and 72 hours. HDAC-1 binding of the eotaxin-3 promoter was confirmed using chromatin immunoprecipitation with appropriate target and control antibodies (Cell Signaling) followed by PCR. Statistical significance was determined by P<0.05 with the Mann-Whitney U test.

Results: Exposure of primary hRPE isolates to class I/II HDAC inhibitors uniformly led to upregulation of eotaxin-3 (P<0.05) at 24 hours compared to vehicle alone or class III HDAC inhibition. While TSA led to elevated pan-eotaxin expression, VPA and SAHA resulted in specifically increased eotaxin-3 mRNA. At 48 hours, supernatant from VPA treated hRPE cells revealed a nearly 20% greater eotaxin-3 concentration compared to vehicle (P<0.05). ChIP demonstrated that HDAC-1 binds to the eotaxin-3 promoter in genomic DNA isolated from human RPE.

Conclusions: In this study, we observed HDAC-1 regulation of the eotaxin-3 gene with multiple class I/II inhibitors leading to elevated expressioin primary hRPE isolates. Ongoing studies in our laboratory are focused on whether epigenetic derangements that occur with aging may cause increased expression of pro-inflammatory gene cassettes in AMD via HDAC-1 modulation.

Keywords: 412 age-related macular degeneration • 536 gene modifiers • 557 inflammation  
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