Wenyuan Wang1, Yufeng Lu1, Yulin Wang2, Yan Zhang3, Beilei Xia1, Juan Cao4. 1. Department of Gastroenterology, Affiliated Hospital of Jiangnan University, Wuxi, China. 2. Clinical Medicine, Xuzhou Medical University, Xuzhou, China. 3. Department of Hematology, Affiliated Hospital of Jiangnan University, Wuxi, China. 4. Department of Infectious Diseases, the Affiliated Wuxi People's Hospital of Nanjing Medical University, Wuxi, China.
Abstract
BACKGROUND: Siderophores are major virulent factors of K. pneumoniae, and their roles are iron chelators in the host. Several studies have shown that iron chelation could result in mitochondrial dysfunction and increase the production of reactive oxygen species (ROS), which further induces cell mitophagy and apoptosis. However, the impacts of siderophores on platelets are still unknown. METHODS: We obtained platelets of healthy volunteers to perform in vitro experiments in our study and treated platelets with different siderophores. Mitophagy related proteins (TOMM20, TIMM23, LC3, and p62), signal proteins (PINK1/Parkin and BNIP3), and apoptosis protein (caspase3) in platelets were analyzed by western blot. The co-localization of mitotracker with LC3-II was analyzed by immunofluorescence assays. The flow cytometer was used to evaluate ROS levels. RESULTS: All four kinds of siderophores (10 μM) secreted by K. pneumoniae increased the expression of LC3 II and reduced the expression of mitochondrial membrane protein, TOMM20, and TIMM23. Immunofluorescence assays revealed that the treatment of enterobactin significantly increased the co-localization of mitotracker with LC3-II. All four kinds of siderophores increased the ROS level in platelets. Mitophagy of platelets was activated through several pathways, including PINK1/Parkin- and BNIP3-dependent pathways. We also proved that siderophores increased the expression of caspase3 in platelets, and the expression of caspase3 significantly decreased after the pathways of mitophagy were blocked. CONCLUSIONS: K. pneumoniae siderophores lead to mitophagy in platelets, and mitophagy further induces apoptosis, which may be a potential treatment of thrombocytopenia in infections. 2020 Annals of Translational Medicine. All rights reserved.
BACKGROUND: Siderophores are major virulent factors of K. pneumoniae, and their roles are iron chelators in the host. Several studies have shown that iron chelation could result in mitochondrial dysfunction and increase the production of reactive oxygen species (ROS), which further induces cell mitophagy and apoptosis. However, the impacts of siderophores on platelets are still unknown. METHODS: We obtained platelets of healthy volunteers to perform in vitro experiments in our study and treated platelets with different siderophores. Mitophagy related proteins (TOMM20, TIMM23, LC3, and p62), signal proteins (PINK1/Parkin and BNIP3), and apoptosis protein (caspase3) in platelets were analyzed by western blot. The co-localization of mitotracker with LC3-II was analyzed by immunofluorescence assays. The flow cytometer was used to evaluate ROS levels. RESULTS: All four kinds of siderophores (10 μM) secreted by K. pneumoniae increased the expression of LC3 II and reduced the expression of mitochondrial membrane protein, TOMM20, and TIMM23. Immunofluorescence assays revealed that the treatment of enterobactin significantly increased the co-localization of mitotracker with LC3-II. All four kinds of siderophores increased the ROS level in platelets. Mitophagy of platelets was activated through several pathways, including PINK1/Parkin- and BNIP3-dependent pathways. We also proved that siderophores increased the expression of caspase3 in platelets, and the expression of caspase3 significantly decreased after the pathways of mitophagy were blocked. CONCLUSIONS: K. pneumoniae siderophores lead to mitophagy in platelets, and mitophagy further induces apoptosis, which may be a potential treatment of thrombocytopenia in infections. 2020 Annals of Translational Medicine. All rights reserved.
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