Abstract：Objective To investigate the effects of 5-azacytidine on radiation-induced epithelial-mesenchymal transition in rat alveolar type Ⅱ epithelial cell line (RLE-6TN) and explore their working mechanisms, and to provide experimental evidence for the potential drug-based treatment of radiation-induced pulmonary fibrosis. Methods RLE-6TN cells were cultured in vitro and divided into four groups according to the experimental purposes:control group (C), radiation group (R), 5-azacytidine group (A), and radiation followed by 5-azacytidine group (R+A). The microstructural changes in cells were determined by transmission electron microscopy. Inverted phase-contrast microscopy revealed the morphological changes in cells. The mRNA expression levels of E-cadherin and α-SMA were measured by quantitative real-time polymerase chain reaction (qRT-PCR). The protein expression levels of E-cadherin, GSK3β, and p-GSK3β(Ser9) were measured by Western blot. The one-way analysis of variance was used for pairwise comparison. Results The cells in group R became spindle-like. Similar morphological changes were not observed in cells in group R+A. Osmiophilic lamellar bodies disappeared at last in cells in group R. RT-PCR results showed that compared with group C, group R had a significantly lower mRNA expression level of E-cadherin ((0.23±0.06) vs.(1.00±0.00),P=0.002)) and a significantly higher mRNA expression level of α-SMA ((2.91±0.01) vs.(1.00±0.00),P=0.000)). However, compared with group R, group R+A had a significantly higher mRNA expression level of E-cadherin ((0.47±0.05) vs. (1.00±0.00),P=0.024)) but a significantly lower mRNA expression level of α-SMA ((2.50±0.02) vs.(1.00±0.00),P=0.037)). The results of Western blot showed that the protein expression level of E-cadherin was significantly reduced ((0.07±0.01) vs.(0.48±0.02),P=0.028)), while the proteinexpression level of p-GSK3β was significantly increased in Group R than in Group C ((0.85±0.04) vs.(0.23±0.03),P=0.031)). However, compared with group R, group R+A had a significantly lower protein expression level of E-cadherin ((0.25±0.00) vs.(0.07±0.01),P=0.024)) and significantly less up-regulation of the protein expression level of p-GSK3β((0.39±0.03) vs.(0.85±0.04),P=0.014)). Conclusions X-ray radiation can induce the epithelial-mesenchymal transition in epithelial cells. 5-azacytidine suppresses radiation-induced epithelial-mesenchymal transition by inhibition of the activity of p-GSK3β in RLE-6TN cells.
Wang Yacheng,Li Xin,Chen Jing et al. Effects of 5-azacytidine on radiation-induced epithelial-mesenchymal transition in rat alveolar type Ⅱ epithelial cell line[J]. Chinese Journal of Radiation Oncology, 2015, 24(3): 349-352.
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