[Inhibition of Wnt/β-catenin signal and NOX4 impairs repair of silica-induced lung epithelial cell injury].

Objective To investigate the effects of the interaction between Wnt/β-catenin signal and NADPH oxidase 4 (NOX4) on the proliferation of pulmonary epithelial cells in response to silica exposure. Methods The airway instillation of silica to C57BL/6 mice was used to produce mouse silicosis models. Immunohistochemistry was used to determine NOX4 expression in the lungs of silicosis mice. Human lung epithelial cells (BEAS-2B) were exposed to silica to generate an oxidative injury epithelial cell model in vitro. Wnt signal conditioned medium (Wnt3a-CM) and Wnt signal inhibitor XAV939 were used to alter the activity of Wnt signal. An infection adenoviral vector expressing short hairpin RNA to NOX4 (NOX4 shRNA) was used to knock down NOX4 expression in BEAS-2B cells. Western blotting was performed to access the expression of Wnt3a, active-β-catenin (ABC), transcription factor 4 (TCF4), cyclin D1 and NOX4 proteins in lung tissues and human lung epithelial cells. CCK-8 assay was used to determine the effects of silica of different concentrations on cell viability as well as the effects of NOX4 expression knockdown on cell proliferation in human lung epithelial cells. CellROX fluorescent probe loading assay was used to detect the release of reactive oxygen species (ROS). Results Mouse silicosis model and BEAS-2B cell model of oxidative damage were successfully generated. The stimulation of silica significantly activated Wnt/β-catenin signal and induced NOX4 expression, sequentially resulting in ROS production. While ROS scavenger N-acetyl-L-cysteine (NAC) inhibited the silica-induced release of ROS, and then inhibited the expression of ABC protein and Wnt/β-catenin signal activity. An activation of Wnt signaling induced by Wnt3a-conditioned medium (Wnt3a-CM) increased the expression of NOX4, whereas the Wnt signal inhibitor XAV939 inhibited the expression of NOX4. The expression of Wnt/β-catenin signal ABC and cyclin D1 and cell proliferation were significantly inhibited by the shRNA-mediated suppression of NOX4 expression. Conclusion Blocking Wnt/-catenin signal and down-regulating NOX4 expression inhibit the proliferation of lung epithelial cells and the damage repair of lung epithelial cells induced by the silica exposure.