The Role Of MiR-150 In Regulation Of NKT Cells In Bacterial Infection And Autoimmune Diseases

Objectives To investigate the effect of miR-150 knockout on bacterial infection and rheumatoid arthritis through NKT cells.Methods 1 Using flow cytometry to sort NKT cells from different stages of mouse thymus and extract total RNA.After reverse transcription and pre-amplification,TaqMan low-density microRNA expression profiling array was used to detect expression changes,further verified using real-time PCR.2 The infection model of miR-150 KO mice and C57BL/6J mice were established by E.col K12.Survival and count of bacteria in lung and liver investigate the effect of miR-150 knockdown on bacterial resistance.Wright's staining and flow cytometry of peripheral blood neutrophils to identify the count of NKT cells and neutrophil.3 mi R-150 KO mice and C57BL/6J mice were immunized with type II collagen in complete Freund's adjuvant(CFA)to induce arthritis.The severity of arthritis was assessed on clinical score and ankle thickness.Using flow cytometry to measure CD4~+T cells,CD8~+T cells,B cells and IFN-?,IL-4,IL-17 in NKT cells.Besides,real-time PCR detected TNF-?,IFN-?,IL-17,IL-4 mRNA from cells in Joint cavity.Results 1 MicroRNA expressional profile analysis revealed that there are 36 microRNAs expression increased and 12 microRNAs whose expression decreased continuously from stage 1 to stage 3 NKT cells.The expression changes of let-7f,miR-150,miR-24,miR-29 a,in the process of NKT development were increased,and miR-223 and miR-155 were decreased.2 Compared with C57BL/6J mice,survival of mi R-150 KO mice was significant higher,and count of bacteria is lower.NKT cells had no significant changes in peripheral blood of two mice with health and infection,but neutrophils were increased significantly in peripheral blood of miR-150 KO mice compared with C57BL/6J mice.3The incidence of arthritis and disease severity in miR-150 KO mice was markedly lower than in C57BL/6J mice.We measured CD4~+T cells,CD8~+T cells,B cells and IFN-?,IL-4,IL-17 in NKT cells in spleen and inguinal lymph node,finded no significant change in miR-150 KO mice and C57BL/6J mice.We obversed that IFN-?,IL-4 were significant increased and IL-17 was significant decreased in NKT from spleen.The expression changes of TNF-?,IFN-?,IL-4,IL-17 mRNA in joint cavity were decreased compared with C57BL/6J mice.Conclusions 1 The expression changes of a number of specific microRNAs in NKT cells suggests that specific microRNAs regulate the development and function of NKT cells,and miR-150 was significant increased in the development of NKT cells.2 MiR-150 knockout was significant increased resistance of bacteria,it may be related to the increase of neutrophils mainly rather than NKT cells.3 miR-150 knockout inhibited the occurrence and development of arthritis,it may be related to the increase of IFN-?and IL-4 and decrease of IL-17 producted through NKT cells.