The role of Hsp70 and Hsp90 in TGF-β-inducedepithelial-to-mesenchymal transition in rat lens epithelial explants

[摘要] Purpose: This study investigates the effects of heat shock treatmentand the role of Hsp70 and Hsp90 on tranforming growth factor beta 2(TGF-β2)-induced epithelial-to-mesenchymal transition (EMT) in ratlens epithelial explants.Methods: Rat lens epithelial explants from 7-10 day-old Wistar ratswere dissected and incubated for 24 h before treatment. The explantswere divided into eight treatment groups: control (culture medium),fibroblast growth factor-2 (FGF-2), TGF-β2, and TGF-β2+FGF-2under normal culture conditions and heat shocked conditions. Theexplants were heat shocked at 45 °C before treatment with therespective media. H&E staining was performed on whole-mountepithelial explants from each group. Immunofluorescence staining forα-smooth muscle actin (α-SMA), F-actin, and E-cadherin wasalso used to determine EMT and fibrotic plaque formation in the lensepithelial explants. Apoptotic cell death was determined using the TUNEL(terminal deoxynucleotidyl transferase mediated dUTP nick end labeling)assay. Confocal microscopy was used to visualize immunoreactivity in thewhole-mount epithelial explants. Western blot analysis of α-SMA,E-cadherin, Hsp70, and Hsp90 were also performed.Results: TGF-β2-induced EMT and plaque formation in the lensepithelial explants. The simultaneous treatment of epithelial explantswith TGF-β2+FGF-2-induced the most significant morphologicalchanges and EMT. Heat shock treatment of lens epithelial explants beforeTGF-β2 treatment did not inhibit plaque formation, but there wassignificant reduction of α-SMA expression and greater E-cadherinexpression when compared to the non-heat shocked TGF-β2-treatedexplants. Interestingly, TGF-β-induced apoptotic cell death wassignificantly lower in the heat shocked explants compared to thenon-heat shock lens explants. Heat-induced accumulation of Hsp70 andHsp90 expression was reduced in the heat shocked groups at day 4 oftreatment.Conclusions: TGF-β2-induced EMT was significantly reduced inthe heat shocked TGF-β2 lens epithelial explants. After four daysof culture, there is a reduction in expression of Hsp70 and Hsp90 in theheat-shocked groups, indicating that the lens epithelial cells are undera less stressful condition than the non-heat shocked groups. Inconclusion, molecular chaperones can play a protective role againstTGF-β2-induced EMT and enhance cell survival.

[学科分类] 生物化学/生物物理