Zhiqiang Xue1, Wei Yuan2, Jing Li1, Hong Zhou1, Lihua Xu1, Jiayi Weng1, Xiaoyang Li1, Xinru Zhang1, Zhongqun Wang1, Jinchuan Yan3. 1. Department of Cardiology, Affiliated Hospital of Jiangsu University, Zhenjiang, Jiangsu Province, People's Republic of China. 2. Department of Cardiology, Affiliated Hospital of Jiangsu University, Zhenjiang, Jiangsu Province, People's Republic of China. Electronic address: yuanwei1@medmail.com.cn. 3. Department of Cardiology, Affiliated Hospital of Jiangsu University, Zhenjiang, Jiangsu Province, People's Republic of China. Electronic address: yanjinchuan@hotmail.com.
Abstract
BACKGROUND: Oxidized low-density lipoprotein (ox-LDL) is the most common inflammatory factor that mediates the activation and apoptosis of macrophages. Cyclophilin A (CyPA) is expressed following oxidative stress, hypoxia, and infection. However, the role of CyPA in the activation and apoptosis of macrophages is unclear. The aims of the study were to determine whether CyPA mediates the ox-LDL-induced activation and apoptosis in RAW264.7 cells and to analyze potential mechanisms. METHODS AND RESULTS: Through Western blot and ELISA test, the expression of CyPA induced by ox-LDL is time-dependent in RAW264.7 cells. Gene silencing of CyPA reduced the generation of lipid droplets in the cytoplasm and downregulated the expression of the surface markers of macrophage activation, namely, CD80, CD86, and major histocompatibility complex class 2 antigen. Cell apoptosis is significantly decreased and the level of anti-apoptosis protein bcl-2 is increased in CyPA silent cells compared with the control group. Finally, autophagy-related protein LC3-II/LC3-I ratio level significantly decreased in CyPA silent cells with less autophagosome formation while the blocked autophagy flux was recovered. The differences in the activation and apoptosis between CyPA silent cells and the control cells were inhibited by pre-treatment with class III PI 3-kinase inhibitor 3-MA. CONCLUSIONS: These results indicate that CyPA mediates the ox-LDL-induced activation and apoptosis in RAW264.7 cells by regulating autophagy.
BACKGROUND: Oxidized low-density lipoprotein (ox-LDL) is the most common inflammatory factor that mediates the activation and apoptosis of macrophages. Cyclophilin A (CyPA) is expressed following oxidative stress, hypoxia, and infection. However, the role of CyPA in the activation and apoptosis of macrophages is unclear. The aims of the study were to determine whether CyPA mediates the ox-LDL-induced activation and apoptosis in RAW264.7 cells and to analyze potential mechanisms. METHODS AND RESULTS: Through Western blot and ELISA test, the expression of CyPA induced by ox-LDL is time-dependent in RAW264.7 cells. Gene silencing of CyPA reduced the generation of lipid droplets in the cytoplasm and downregulated the expression of the surface markers of macrophage activation, namely, CD80, CD86, and major histocompatibility complex class 2 antigen. Cell apoptosis is significantly decreased and the level of anti-apoptosis protein bcl-2 is increased in CyPA silent cells compared with the control group. Finally, autophagy-related protein LC3-II/LC3-I ratio level significantly decreased in CyPA silent cells with less autophagosome formation while the blocked autophagy flux was recovered. The differences in the activation and apoptosis between CyPA silent cells and the control cells were inhibited by pre-treatment with class III PI 3-kinase inhibitor 3-MA. CONCLUSIONS: These results indicate that CyPA mediates the ox-LDL-induced activation and apoptosis in RAW264.7 cells by regulating autophagy.
Authors: Ignacio Hernandez; Laura Tesoro; Rafael Ramirez-Carracedo; Javier Diez-Mata; Sandra Sanchez; Marta Saura; Jose Luis Zamorano; Carlos Zaragoza; Laura Botana Journal: Int J Mol Sci Date: 2021-03-12 Impact factor: 5.923