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Daisy Shu, Frank J Lovicu; EGFR signaling promotes TGFβ-dependent epithelial-mesenchymal transition (EMT) in the lens. Invest. Ophthalmol. Vis. Sci. 2017;58(8):3188. doi: https://doi.org/.
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© ARVO (1962-2015); The Authors (2016-present)
Epithelial-mesenchymal transition (EMT) of lens epithelial cells (LECs) plays a crucial role in the pathogenesis of anterior subcapsular cataract (ASC) and posterior capsular opacification (PCO). While transforming growth factor-beta (TGFβ) is a potent inducer of EMT in the lens, studies in cancer research have shown that the effects of TGFβ can be augmented by the addition of epidermal growth factor (EGF). The present study aims to investigate the combined effect of TGFβ and EGF in the lens to elucidate the involvement of EGF receptor (EGFR)-signaling in TGFβ-induced EMT in LECs.
Lens epithelial explants from 21-day-old Wistar rats were cultured for up to five days with either 200 pg/ml TGFβ2, 5 ng/ml EGF, or a combination of these, with or without a 2-hour pre-treatment with 50 nM PD153035 (EGFR inhibitor). Expression of β-catenin, alpha-smooth muscle actin (α-SMA), tropomyosin (isoforms 1.6-1.9) and downstream signaling molecules, including phosphorylated EGFR, Smad2/3 and MAPK/ERK1/2 were determined using immunofluorescence and/or western blotting. Cell morphology was examined using phase-contrast microscopy.
Compared to TGFβ2 alone, the combined treatment of LECs with TGFβ2 and EGF resulted in a more dramatic morphological elongation and transdifferentiation into myofibroblastic cells. Cells treated with EGF alone retained their epithelial phenotype. The addition of EGF augmented the expression levels of α-SMA and tropomyosin compared to TGFβ2 alone. Treatment with TGFβ2 alone upregulated the phosphorylation of Smad2/3, ERK1/2 and the EGF receptor. Interestingly, pharmacologic inhibition of EGFR-signaling using PD153035 inhibited TGFβ2-induced upregulation of α-SMA and tropomyosin, with retention of β-catenin labeling to the cell membrane. PD153035 also prevented cell elongation, lens capsular wrinkling and cell loss associated with TGFβ2-induced EMT over the five-day culture period.
Our findings demonstrate that TGFβ2 cooperates synergistically with EGF to enhance EMT in LECs. Moreover, as TGFβ2 alone can transactivate EGFR signaling, and EGFR-signaling is required for TGFβ2 activity, we reveal a novel mechanism in the pathogenesis of ASC and PCO. Hence, by targeting both TGFβ2 and/or EGFR activity, this may be a more efficacious strategy in the pharmacologic treatment of cataract.
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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