September 2016
Volume 57, Issue 12
Open Access
ARVO Annual Meeting Abstract  |   September 2016
TGFβ2-mediated Epithelial to Mesenchymal Transition of lens epithelial cells: Role of AGE-RAGE interaction
Author Affiliations & Notes
  • Cibin T Raghavan
    Ophthalmology , University of Colorado, Aurora, Colorado, United States
  • Ram H Nagaraj
    Ophthalmology , University of Colorado, Aurora, Colorado, United States
  • Footnotes
    Commercial Relationships   Cibin Raghavan, None; Ram Nagaraj, None
  • Footnotes
    Support  EY022061
Investigative Ophthalmology & Visual Science September 2016, Vol.57, 2017. doi:
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      Cibin T Raghavan, Ram H Nagaraj; TGFβ2-mediated Epithelial to Mesenchymal Transition of lens epithelial cells: Role of AGE-RAGE interaction. Invest. Ophthalmol. Vis. Sci. 2016;57(12):2017.

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

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Abstract

Purpose : Proteins in basement membrane accumulate chemical modifications with age. One such modification is glycation, which results in the formation of advanced glycation endproducts (AGEs). Our previous study has shown that AGEs in human lens capsule promote TGFβ2- mediated epithelial to mesenchymal transition (EMT) of lens epithelial cells. In this study we have investigated the role of a receptor for advanced glycation endproducts (RAGE), a multi-ligand receptor, in TGFβ2-mediated EMT of lens epithelial cells.

Methods : Tissue culture plates were coated with a basement membrane extract (BME; 50 mg/ml) overnight and AGE-modified with a glycating mixture (2 mM ascorbate, 25 mM glucose and 250 μM methylglyoxal) for 7 days at 37°C. FHL 124 cells were cultured on unmodified or AGE-modified BME and treated with 10 ng/ml of TGFβ2 for 24 h at 37°C. RNA was isolated, cDNA generated and real time PCR analysis was carried for EMT-associated proteins. RAGE overexpression studies were carried out by transient transfection of GFP-RAGE. Effect of RAGE blockade on TGFβ2-mediated EMT response was studied using a RAGE antibody (10 μg/ml) and EN-RAGE (10 μg/ml), an endogenous ligand for RAGE.

Results : Similar to our previous observation in primary human lens epithelial cells, FHL124 cells also showed an enhanced EMT response upon TGFβ2 treatment when cultured on AGE-modified BME than on unmodified BME. RAGE was detected in FHL124 cells and its levels were unaltered in cells grown on either native or AGE-modified BME or upon treatment with TGFβ2. However, RAGE overexpression in these cells amplified the TGFβ2-mediated EMT response. Moreover, blockade of RAGE with an antibody or EN-RAGE followed by TGFβ2 treatment resulted a significant reduction in the EMT response.

Conclusions : These results imply that the interaction of matrix AGEs with RAGE plays an important role in TGFβ2-mediated EMT of lens epithelial cells and suggest that blockade of RAGE could be a strategy to prevent posterior capsule opacification.

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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