ABSTRACT
Objective: To investigate the role of X-box binding protein 1 (XBP1) in the treatment of human osteosarcoma HOS cells by pyropheophorbide-α methyl ester-mediated photodynamic therapy (MPPα-PDT), and to explore the possible mechanism. Methods: HOS cells were treated by MPPα-PDT for 6, 12 and 24 h, and the expression levels of inositol-requiring enzyme 1 alpha (IRE1α)-XBP1 pathway-related protein IRE1α and XBP1 were detected by Western blotting. The specific siRNA targeting XBP1 gene was transfected into HOS cells by lipofection, and treated by MPPα-PDT. The cells were divided into the blank group, siRNA-negative control (NC) group, siRNA-XBP1 group, MPPα-PDT group and MPPα-PDT+siRNA-XBP1 group. The expressions of XBP1 mRNA and protein were detected by real-time fluorescent quantitative PCR and Western blotting, respectively. The proliferation of HOS cells was detected by CCK-8 assay. The apoptotic rate was analyzed by flow cytometry (FCM). The expression levels of cleaved-caspase 3 and cleaved-poly ADP-ribose polymerase (cleaved-PARP) were determined by Western blotting. The intracellular level of reactive oxygen species (ROS) was detected by DCFH-DA probe staining, then the expression levels of Catalase and superoxide dismutase 1 (SOD1) proteins were detected by Western blotting. Results: The expression levels of IRE1α-XBP1 pathway-related proteins IRE1α and XBP1 were increased after the treatment of MPPα-PDT (both P < 0.05 ). siRNA-XBP1 inhibited the expression levels of XBP1 mRNA and protein (both P < 0.01). Silencing siRNA-XBP1 expression inhibited the proliferation activity of HOS cells (P < 0.01), up-regulated the apoptosis rate and the expression level of apoptosis-related protein cleaved-caspase 3 (both P < 0.05), and down-regulated the expression levels of antioxidant enzyme-related proteins Catalase and SOD1 (both P < 0.05). Compared with the MPPα-PDT group, the proliferation activity of HOS cells treated with MPPα-PDT+siRNA-XBP1 was decreased (P < 0.01), the apoptosis rate and the expression levels of cleaved-caspase 3 and cleaved-PARP were increased (all P < 0.05), the intracellular ROS level was up-regulated (P < 0.01), and the expression levels of Catalase and SOD1 were decreased (both P < 0.01). Conclusion: MPPα-PDT can induce the activation of IRE1α-XBP1 pathway in HOS cells. Silencing XBP1 can inhibit the proliferation activity of HOS cells, up-regulate the apoptosis rate, and increase the sensitivity of HOS cells to MPPα-PDT. The mechanism may be related to the up-regulation of intracellular ROS level and the down-regulation of anti-oxidation molecules.