c-Myc mediates Epithelial-to-Mesenchymal Transition in Lens Epithelial Cells

Abstract

Purpose : Transforming growth factor beta (TGF-β2) has been well characterized as a potent inducer of epithelial-to-mesenchymal transition (EMT) in lens epithelial cells (LECs) during the formation of secondary cataract. We find that c-Myc is upregulated in this process. In this study, we investigated the role and mechanism of c-Myc on EMT in human LECs

Methods : The human lens epithelial cells (HLECs), SRA01/04, were treated with different doses of TGF-β2 (5 or 10ng/mL) for 48 hours to induce the EMT response. The abundance of c-Myc and downstream target genes were measured. To investigate the direct effect of c-Myc on EMT in HLECs, SRA01/04 cells were transfected with small interfering RNAs (siRNA) of c-Myc to inhibit its expression, or treated with 10058-F4 (50 μM), which inhibit c-Myc-Max interaction, to suppress the function of c-Myc. After transfection of siRNA or treatment with 10058-F4, HLECs were cultured in the presence or absence of TGF-β2 (5 ng/mL) for 24-48 hours. The experssion of EMT markers, such as fibronectin (FN), collagen IV (Col IV) and SNAI1/SNAI2 were determined using real-time PCR or western blot.

Results : Treatment of HLECs with TGF-β2 for 24 or 48 hours induced a significant upregulation of well-established ECM markers, FN and Col IV. We also observed a remarkable dose-dependent induction of c-Myc mRNA and protein levels treated with TGF-β2 at 5 and 10 ng/mL for 48 hours. The abundance of c-Myc transcriptional target gene NCL is also increased. Knockdown c-MYC in HLECs using siRNA diminished FN and Col IV induction by TGF-β2 suggesting EMT induced by TGF-β2 may be inhibited by c-Myc knockdown. Blockade of c-Myc reduced the core EMT transcription factor SNAI1 with or without TGF-β2 presence. Furthermore, 10058-F4, a c-Myc inhibitor which specifically block the c-Myc-Max interaction, showed similar effects.

Conclusions : Our findings demonstrate c-Myc is activated and might be required for TGF-β2 induced EMT in HLECs. Both genetically and pharmacologically Inhibition of c-Myc expression could suppress EMT indicating that 10058-F4 might be a potent therapeutic option in the treatment of cataract.

This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.