June 2017
Volume 58, Issue 8
Open Access
ARVO Annual Meeting Abstract  |   June 2017
Endoplasmic reticulum stress at the intersection of inflammation and hyperglycemia in mediating tight junction alterations in diabetic retinopathy
Author Affiliations & Notes
  • RAJI RAJESH LENIN
    Department of Ophthalmology, University of Tennessee, Memphis, Tennessee, United States
  • Rajashekhar Gangaraju
    Department of Ophthalmology, University of Tennessee, Memphis, Tennessee, United States
  • Footnotes
    Commercial Relationships   RAJI RAJESH LENIN, None; Rajashekhar Gangaraju, None
  • Footnotes
    Support  EY023427
Investigative Ophthalmology & Visual Science June 2017, Vol.58, 4037. doi:
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      RAJI RAJESH LENIN, Rajashekhar Gangaraju; Endoplasmic reticulum stress at the intersection of inflammation and hyperglycemia in mediating tight junction alterations in diabetic retinopathy. Invest. Ophthalmol. Vis. Sci. 2017;58(8):4037.

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

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Abstract

Purpose : Endoplasmic Reticulum (ER) stress plays a major role in several metabolic disorders including diabetes. We hypothesized that ER stress pathways interact with inflammatory signaling leading to retinal barrier dysfunction in diabetic retinopathy (DR). Using a well characterized chronic proinflammatory tie2-TNF mouse model and retinal endothelial cells we assessed the connection between ER stress and hyperglycemic stress in mediating vascular permeability.

Methods : Tie2-TNF and age matched wildtype mice were made diabetic with intraperitoneal injections of Streptozotocin and evaluated for retinal vascular permeability by fluorescein labeled-BSA leakage assay. Human retinal microvascular endothelial cells (HREC) were treated with 10 ng/mL TNF-α and 30 mmol/L glucose for 3 hours with and without ER stress inhibitor (10µM TUDCA). Gene and protein expression of ER stress markers were analyzed by Taqman assay and Western blot analysis respectively. Alterations in endothelial junction proteins were evaluated by western blot.

Results : Frozen sections of retina from tie2-TNF transgenic mice demonstrated increased vascular leakage compared to wildtype mice which was further exacerbated with diabetes. Interestingly, whole mount retinal preparation from diabetic tie2-TNF mice demonstrated hyper permeable vessels with strong extravasation of FITC-BSA and extravasation of red blood cells. Retinal extracts from 6-month old tie2-TNF mice exhibited increased mRNA levels of ER stress markers (GRP78, PERK, IRE1α and CHOP) compared to age matched wild type mice (n=3 each, p<0.05). In HREC cells, ER stress was significantly upregulated with TNF and high glucose combination as evidenced by increased gene expression of GRP-78, PERK, IRE-1α, ATF-6, XBP-1, CHOP and increased protein expression of GRP78 and a decreased ZO-1 expression (p<0.05 compared to control). On the other hand, TUDCA treated cells reversed ER stress markers.

Conclusions : ER stress is a key mediator of vascular damage in the setting of increased inflammatory burden in retinal endothelial cells. Understanding the underlying ER stress mechanisms may shed new insights into novel therapeutic targets for DR.

This is an abstract that was submitted for the 2017 ARVO Annual Meeting, held in Baltimore, MD, May 7-11, 2017.

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