Feng Tian1, Jiazhao Yang1, Rui Xia2. 1. Department of Traumatic orthopedics, The First Affiliated Hospital of USTC, No.17 Lujiang Road, Luyang District, 230001, Hefei City, Anhui Province, China. 2. Department of Traumatic orthopedics, The First Affiliated Hospital of USTC, No.17 Lujiang Road, Luyang District, 230001, Hefei City, Anhui Province, China. tf13514993782@163.com.
Abstract
BACKGROUND: Spinal cord injury (SCI) is a devastating neurological event that leads to severe motor and sensory dysfunction. Exosome-mediated transfer of circular RNAs (circRNAs) was associated with SCI, and exosomes have been reported to be produced by mesenchymal stem cells (MSCs). This study is designed to explore the mechanism of exosomal circZFHX3 on LPS-induced MSCs injury in SCI. METHODS: Exosomes were detected by transmission electron microscope and nanoparticle tracking analysis. CD9, CD63, CD81, and TSC101, B-cell lymphoma-2 (Bcl-2), Bcl-2 related X protein (Bax), Cleaved caspase 3, and Insulin-like growth factor 1 (IGF-1) protein levels were measured by western blot assay. CircZFHX3, microRNA-16-5p (miR-16-5p), and IGF-1 level were detected by real-time quantitative polymerase chain reaction (RT-qPCR). Cell viability and apoptosis were detected by Cell Counting Kit-8 (CCK-8) and flow cytometry assay. Levels of IL-1β, IL-6, and TNF-α were assessed using Enzyme-linked immunosorbent assays (ELISA). ROS, LDH, and SOD levels were measured by the special kits. The binding between miR-16-5p and circZFHX3 or IGF-1 was predicted by Starbase and DianaTools and then verified by a dual-luciferase reporter and RNA Immunoprecipitation (RIP) assays. The biological role of exosomal circZFHX3 on SCI mice was examined in vivo. RESULTS: CircZFHX3 and IGF-1 were decreased, and miR-16-5p was increased in SCI mice. Also, exosomal circZFHX3 boosted cell viability and repress apoptosis, inflammation, and oxidative stress in LPS-treated BV-2 cells in vitro. Mechanically, circZFHX3 acted as a sponge of miR-16-5p to regulate IGF-1 expression. Exosomal circZFHX3 reduced cell injury of SCI in vivo. CONCLUSIONS: Exosomal circZFHX3 inhibited LPS-induced BV-2 cell injury partly by regulating the miR-16-5p/ IGF-1 axis, hinting at a promising therapeutic strategy for the SCI treatment.
BACKGROUND: Spinal cord injury (SCI) is a devastating neurological event that leads to severe motor and sensory dysfunction. Exosome-mediated transfer of circular RNAs (circRNAs) was associated with SCI, and exosomes have been reported to be produced by mesenchymal stem cells (MSCs). This study is designed to explore the mechanism of exosomal circZFHX3 on LPS-induced MSCs injury in SCI. METHODS: Exosomes were detected by transmission electron microscope and nanoparticle tracking analysis. CD9, CD63, CD81, and TSC101, B-cell lymphoma-2 (Bcl-2), Bcl-2 related X protein (Bax), Cleaved caspase 3, and Insulin-like growth factor 1 (IGF-1) protein levels were measured by western blot assay. CircZFHX3, microRNA-16-5p (miR-16-5p), and IGF-1 level were detected by real-time quantitative polymerase chain reaction (RT-qPCR). Cell viability and apoptosis were detected by Cell Counting Kit-8 (CCK-8) and flow cytometry assay. Levels of IL-1β, IL-6, and TNF-α were assessed using Enzyme-linked immunosorbent assays (ELISA). ROS, LDH, and SOD levels were measured by the special kits. The binding between miR-16-5p and circZFHX3 or IGF-1 was predicted by Starbase and DianaTools and then verified by a dual-luciferase reporter and RNA Immunoprecipitation (RIP) assays. The biological role of exosomal circZFHX3 on SCI mice was examined in vivo. RESULTS: CircZFHX3 and IGF-1 were decreased, and miR-16-5p was increased in SCI mice. Also, exosomal circZFHX3 boosted cell viability and repress apoptosis, inflammation, and oxidative stress in LPS-treated BV-2 cells in vitro. Mechanically, circZFHX3 acted as a sponge of miR-16-5p to regulate IGF-1 expression. Exosomal circZFHX3 reduced cell injury of SCI in vivo. CONCLUSIONS: Exosomal circZFHX3 inhibited LPS-induced BV-2 cell injury partly by regulating the miR-16-5p/ IGF-1 axis, hinting at a promising therapeutic strategy for the SCI treatment.
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