Protection Of IL-33 Against MRSA Infection In Immunosuppre-ssive Mice And Preliminary Discussion Of Its Mechanism

As one of the most commonly used antineoplastic chemotherapeutic drugs, cyclophosphamide strongly inhibits the immune function, including the sharp reduction of white blood cells. Purulent infection is mainly caused by methicillin-resistant staphylococcus aureus(MRSA), which is resistant to multiple antibiotic therapy and spread fast. Neutrophils are generated from hematopoietic stem cells in bone marrow and circulate in blood for a few hours. Once microorganisms have successfully overcome the physical barriers and gained access to tissues, neutrophil migrate rapidly to the site of infection, killing pathogenic microorganisms. IL-33, as an "alarmin" and proinflammatory cytokine, plays an important role in infection, arthritis, atherosclerosis and et al. In this study, we will establish the immunosuppressive mice model with cyclophosphamide, and observe the therapeutic effect and immunological mechanism of IL-33 in this model. This data may provide a new evidence for the therapy of cyclophosphamide-induced immune suppression. Purpose:We investigate the anti-infection effect of IL-33 in the immunosuppressive mice model. At the same time, we explore the effect of IL-33 on the differentiation of hematopoietic stem cells to neutrophils, and the effect of IL-33 on neutrophil bactericidal and apoptotic activities in vitro. Methods:We will establish the immunosuppressive mice model with cyclophosphamide. After received intraperitoneal injection of IL-33, the mice were infected with MRSA. We observed the survival rate of mice, the number of neutrophils in blood and peritoneal fluid and the load of bacteria. We acquired neutrophils with percoll separation, then discussed the effect of IL-33 on the functions of neutrophils, such as chemotaxis, phagocytosis and bactericidal action. Flow cytometry was used to analyze the STAT3 phosphorylation, ROS generation and the apoptosis of neutrophils. MACs Micro Beads was used to separate and enrich hematopoietic stem cells. Brd U was used to detect the effect of IL-33 on proliferation of hematopoietic stem cells. Colony forming experiment was used to test the directional differentiation of hematopoietic stem cells to neutrophils. IL-33-pretreated neutrophils transfusion was used to treat immunosuppressive mice and observed its antiinfection effect. Results: 1. Protective effect of IL-33 on MRSA abdominal infection in immunosuppressive miceThe model of immunosuppressive mice was established successfμlly. After treatment with IL-33, the survival rate of immunosuppressive mice significantly increased. The number of neutrophils in blood and peritoneal fluid are increased. The load of bacteria was reduced. The function of neutrophils such as chemotaxis phagocytosis, bactericidal action was suppressed in immunosuppressive mice, while IL-33 improved all these functions of neutrophils. 2. IL-33 promotes the differentiation of hematopoietic stem cells to neutrophilsIL-33 activated JAK2-STAT3 pathway in hematopoietic stem cells, as well as the C/EBPβ-c-myc signal, which located the downstream of STAT3. IL-33 promoted hematopoietic stem cell proliferation and directional differentiation to neutrophils by JAK2- STAT3- C/EBPβ-c-myc. 3. IL-33 enhances the antibacterial function of neutrophilsIL-33 regulated ROS generation of neutrophil, which was involved in its bactericidal action via JAK2-STAT3 pathway. IL-33 also raised the Bcl-2 while reducing Bax, thereby inhibiting neutrophil apoptosis. 4. IL-33-pretreated neutrophils transfusion protects immunosuppressive mice against infectionIL-33-pretreated neutrophils transfusion can significantly improve the survival rate and the antiinfection function of neutrophils in immunosuppressive mice. Conclusion:IL-33 promotes hematopoietic stem cell proliferation and directional differentiation to neutrophils so as to increase the number of neutrophils. Meanwhile, IL-33 enhances the antibacterial activities of neutrophils. Thereby, IL-33 improves the fuction of neutrophils against MRSA infection in immunosuppressive mice.