The Role Of Glycogen Synthase 1 In Macrophage LPS Tolerance During Sepsis In Mice

Immunosuppression caused by monocyte/macrophage endotoxin(LPS)tolerance is the main cause of death in patients with sepsis,but its mechanism is far from clear.Abnormal glucose metabolism is the main cause of immune cell dysfunction.Studies have shown that over expression of glycogen synthase 1(GYS1)damages the function of myeloid leukemia cells and inhibits the release of inflammatory cytokines in the brain of aged mice.However the role of GYS1 in monocyte/macrophage LPS tolerance has not been reported.The purpose of this study was to explore the role of GYS1 in triggering immunosuppression of sepsis in mice by inducing monocyte/macrophage LPS tolerance.Objective:1.To determine the expression of GYS1 in septic immunosuppressive mice and LPS tolerant macrophages.2.To elucidate the effect of GYS1 on immunosuppression of sepsis in mice.Methods:The immunosuppressive model of sepsis in C57BL/6J mice was established by cecal ligation and puncture(CLP)combined with Pseudomonas aeruginosa(P.a.)(5×10~7 CFU/20 g,i.v.).The P.a.load in peripheral blood and spleen and the secretion of pro-inflammatory factors in serum were observed to determine the successful replication of the model.The expression of GYS1 in monocytes was detected by flow cytometry.Taking the primary peritoneal macrophage of mice as the research object,the model of LPS tolerant macrophage was replicated by LPS(1μg/m L).The secretion level of proinflammatory factors and phagocytic function verification model were detected,and the changes of GYS1 protein and gene in LPS tolerant macrophages were observed.The Gys1 of primary mouse peritoneal macrophages was silenced by lentiviral vector,and the changes of inflammatory indexes of pro-IL-1βand TNF-αwere detected.Adeno-associated virus was injected intravenously to specifically silence macrophage Gys1.On this basis,the immunosuppressive mouse model of sepsis was established.The effect of macrophage silencing was detected by flow cytometry,the P.a.load in liver,lung,spleen and peripheral blood was detected,and the pathological injury of lung,liver and kidney was observed by HE staining.The secretion of pro-inflammatory factors in spleen,lung and liver was detected by ELISA.Results:The bacterial load in peripheral blood and spleen of CLP combined with P.a.increased significantly,and the secretion of pro-inflammatory factor TNF-αin serum decreased,that is,CLP combined with Pseudomonas aeruginosa could successfully replicate the immunosuppressive mouse model of sepsis.High expression of GYS1 in bone marrow monocytes of septic immunosuppressive mice.The second blow of LPS could decrease the secretion of pro-inflammatory factors and phagocytosis of macrophages,that is,the second blow of LPS could successfully replicate the model of LPS-tolerant macrophages and increase the expression of GYS1 protein and m RNA in LPS-tolerant macrophages,while silencing Gys1 could restore the expression of pro-inflammatory cytokines pro-IL-1β,TNF-αand phagocytosis.Whole-level silencing of macrophage Gys1 can reduce the bacterial load in peripheral blood and organs of septic immunosuppressive mice,reduce histopathological damage and restore the secretion of pro-inflammatory factors.Conclusion:1.High expression of GYS1 in mononuclear bone marrow monocytes/macrophages and LPS tolerant macrophages in septic immunosuppressive mice.2.GYS1 triggers immunosuppression of sepsis in mice by inducing monocyte/macrophage LPS tolerance.