April 2014
Volume 55, Issue 13
Free
ARVO Annual Meeting Abstract  |   April 2014
Inhibition of αB Crystallin Induces Mesenchymal to Epithelial Transition in RPE cells Through Down-regulation of Snail and Slug
Author Affiliations & Notes
  • Keijiro Ishikawa
    Arnold and Mabel Beckman Macular Research Center, Doheny Eye Institute, Los Angeles, CA
    Ophthalmology and Pathology, University of Southern California, Los Angeles, CA
  • Ram Kannan
    Arnold and Mabel Beckman Macular Research Center, Doheny Eye Institute, Los Angeles, CA
  • Christine Spee
    Ophthalmology and Pathology, University of Southern California, Los Angeles, CA
  • Parameswaran G Sreekumar
    Arnold and Mabel Beckman Macular Research Center, Doheny Eye Institute, Los Angeles, CA
  • David R Hinton
    Ophthalmology and Pathology, University of Southern California, Los Angeles, CA
Investigative Ophthalmology & Visual Science April 2014, Vol.55, 363. doi:
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      Keijiro Ishikawa, Ram Kannan, Christine Spee, Parameswaran G Sreekumar, David R Hinton; Inhibition of αB Crystallin Induces Mesenchymal to Epithelial Transition in RPE cells Through Down-regulation of Snail and Slug. Invest. Ophthalmol. Vis. Sci. 2014;55(13):363.

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

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Abstract

Purpose: Epithelial-to-mesenchymal transition (EMT) of the retinal pigment epithelium (RPE) is a hallmark of proliferative vitreoretinopathy and age-related macular degeneration. The role of small heat shock proteins (sHSPs) in EMT has not been studied. The aim of this study was to investigate the role of αB crystallin, a prominent member of sHSP family, in EMT of RPE cells.

Methods: All studies were conducted in cultured human primary fetal RPE cells at two to four passages. We studied the expression changes of αB crystallin after stimulation with TGFβ2 (5ng/ml) by Western blotting and realtime RT-PCR analysis. To examine whether modulation of αB crystallin expression can alter EMT markers, we investigated the effects of αB crystallin silencing on the expression of E-cadherin, α-SMA and the transcriptional factors such as Snail and Slug with or without TGFβ2 by Western blotting and realtime RT-PCR. To examine whether αB crystallin is involved in TGFβ/SMAD signaling pathway, we also studied the effects of αB crystallin knockdown on phosphorylation of SMAD2/3 induced by TGFβ2 stimulation.

Results: The levels of mRNA and protein of αB crystallin were not significantly altered in RPE cells after TGFβ2 stimulation. Suppression of αB crystallin by siRNA induced significant up-regulation of E-cadherin and down-regulation of α-SMA, Snail and Slug in mRNA and protein expressions (p<0.05 vs controls). Knockdown of αB crystallin did not alter TGFβ2-induced phosphorylation of SMAD2/3.

Conclusions: Our results show that αB crystallin plays a critical role through modulation of Snail and Slug in EMT but not through TGFβ/SMAD signaling pathway in this process.

Keywords: 512 EMT (epithelial mesenchymal transition) • 701 retinal pigment epithelium • 655 proliferative vitreoretinopathy  
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