Macrophage-Derived Exosomes in TLR9(−/−) Mice Ameliorate Sepsis-Induced Mitochondrial Oxidative Stress and Apoptosis in Cardiomyocytes

OBJECTIVE: To explore the mechanisms of TLR9 from macrophages on mitochondrial apoptosis in cardiomyocytes at early stage of sepsis. METHODS: The in vivo and in vitro sepsis mice were bone marrow transplantation (BMT) with wild type (WT) or (toll-like receptor 9) TLR9 knockout ((−/−) or KO) myeloid cells and then constructed by cecum ligation and puncture (CLP) as vivo experiment and cardiomyocytes cocultured with WT or TLR9-deficient macrophages treated with LPS as vitro experiment, respectively. Sepsis model were performed by CLP. The expression levels of exosome, PI3K/AKT, and ERK1/2, inflammatory factors, and apoptotic proteins were tested by western blot in vivo. Besides, associated apoptotic proteins and JC-1 fluorescence assay were tested in vitro. RESULTS: The expressions of p-PI3K, p-AKT, exosome markers (CD9, CD63, and TSG101), p-ERK1/2, TNF-α, IFN-γ, IL-1β, and cleaved-caspase-3/-9 were significantly increased in septic mice vs. control mice, and these proteins were declined dramatically in TLR9(−/−) BMT mice vs. WT BMT mice in sepsis mice models. Meanwhile, the protein expression of cytochrome C, cleaved-caspase-3, and cleaved-caspase-9 increased significantly in primary mouse myocardial cells cocultured with TLR9(−/−) or WT macrophages stimulated with LPS, and these mitochondrial apoptotic proteins as well as the green 5,5',6,6'-tetrachloro-1,1',3,3'- tetraethylbenzimidazolcarbocyanine iodide (JC-1) fluorescence were dramatically lower in LPS-stimulated cardiomyocytes cocultured with TLR9(−/−) than with WT macrophages. CONCLUSION: TLR9(−/−) in macrophages suppressed the inflammatory reaction as well as the exosome secretion and resulted in the inhibition of apoptosis and oxidative stress in sepsis-induced cardiomyopathy.