Autophagy and apoptosis induced by OmpA of Acinetobacter baumannii via ROS/NLRP3 signaling pathway in THP-1 cells" />
鲍曼不动杆菌OmpA经ROS/NLRP3信号通路调控THP1细胞自噬和凋亡
Autophagy and apoptosis induced by OmpA of Acinetobacter baumannii via ROS/NLRP3 signaling pathway in THP-1 cells
DAI Xiao-yue1, WU Liang1, XIA Wen1, YIN Qing2, ZOU Zhi-qing1, ZHOU Ya-ling2, HE Lei2, DING Long-kun1, XI Yue1, ZHANG You-jiang3, XU Hua-xi1
(1. School of Medicine, Jiangsu University, Zhenjiang Jiangsu 212013; 2. Department of Laboratory, Affiliated Hospital of Jiangsu University, Zhenjiang Jiangsu 212001; 3. Center for Disease Control and Prevention, Guangling District, Yangzhou Jiangsu 225001, China)
Abstract：Objective: To investigate the effect of outer membrance protein A(OmpA) of Acinetobacter baumannii on autophagy and apoptosis of human monocyte macrophage THP-1 cells and its underlying mechanism. Methods: The OmpA/pET-28a vector was constructed and transformed into E.coli BL21(DE3) to express the recombinant OmpA (rOmpA), and the rabbits were immunized by rOmpA to prepare the polyclonal anti-OmpA antibody. The anti-OmpA antibody levels in serum of ventilatorassociated pneumonia(VAP) patients were detected by Western blotting. THP-1 cells were treated with different concentrations(1 μg/mL and 10 μg/mL) of rOmpA for 1 h, 3 h and 6 h; and the expression of autophagy-related proteins LC-3 and Beclin-1 were detected by Western blotting; the flow cytometry was used to detect the THP-1 cell apoptosis rate and ROS production; the mRNA expressions of NLRP3, Caspase-1 and IL-1β in THP-1 cells were detected by qRT-PCR. Results: The OmpA expression in clinical A. baumannii strains could be detected by polyclonal anti-OmpA antibody, and the anti-OmpA antibody (IgG) in serum of VAP patients serum could be detected before and after the onset of VAP. The levels of anti-OmpA antibody in VAP patients′ serum were significantly higher than that before using tracheal intubation (P<0.05). After treated with rOmpA, the expression of autophagy-related proteins LC3-Ⅱ and Beclin-1 in THP-1 cells increased significantly, and the apoptosis rate increased vastly in a time- and concentration-dependent manner (P<0.05). Compared with the control group, when cells were treated by the rOmpA for 1 h, the IL-1β mRNA expression and reactive oxygen species were significantly up-regulated (P<0.05); when treated for 3 h and 6 h, the three kinds of mRNA expression had a significant decrease compared with the control group (P<0.05). Conclusion: OmpA induces THP-1 cells autophagy and apoptosis by activating NLRP3 inflammatory cytokines and releasing inflammatory factors such as ROS and IL-1β.
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