摘要
目的为了证实miR-146b在小鼠M1/M2型巨噬细胞分化平衡中的作用和机制,我们通过分离小鼠骨髓来源巨噬细胞,在LPS和IFN-γ联合刺激下促进M1型细胞分化;并结合李斯特菌感染和内毒性休克,共同证实miR-146b对M1型巨噬细胞分化的作用。方法分离野生型(WT)和IL-10-/-小鼠骨髓细胞,采用GM-CSF诱导成为巨噬细胞。并在IL-10-/-M1型巨噬细胞中转染miR-146bmimic;评价miR-146b对M1型巨噬细胞分化的影响,以及致炎因子TNF-γ、IL-6和IL-12等表达;采用miR-146bmimic处理李斯特菌感染和内毒性休克动物模型,评价miR-146b mimic对此模型的作用。结果相对于scramble组,miR-146bmimic能明显抑制IL-10缺失小鼠骨髓来源巨噬细胞向M1型分化,并降低了TNFα、IL-6和IL-12等表达。miR-146bmimic也明显减弱了机体对李斯特感染的敏感性,导致肝脏和脾脏菌斑增多;此外,在高剂量LPS刺激下,miR-146b mimic延长了小鼠生存率,并且降低了致炎因子TNFα、IL-6和IL-12等表达。而且,IRF5可以作为miR-146b直接靶向因子。结论 miR-146b在小鼠M1型巨噬细胞激活过程作为一个负调控因子,起到了抑制炎症的作用。
Objective To investigate the function of miR-146b in the balance of M1/M2 type maerophage differenti- ation,we isolated the BMDMs and was stimulated by LPS and IFN-g,and confirmed the M1 type macrophage together with Lister infection and endoxin shock model. Methods BMDMs were isolated and stimulated by GM-CSF from WT and Ⅱ-10-/- mice. miR-146b mimic or scramble was transfected in IL-10-/- macrophage. The level of TNF-γ,IL-6 and IL-12 were accessed respectively by qPCR,ELISA or Flow cytometry analysis, miR-146b mimic also was used to treat the Lister infection and endoxin shock model. Results the results showed that miR-146b mimic could obviously inhibit the differentiation of M1 type macrophage, accompany with the decrease of TNFa, IL-6 and IL-12 expression, miR-146b mimic also could enhance the Lister infection by repressing the expression TNFa,IL-6 ,etc. but could prolong the sur- vival of endoxin shock murine model compared to scramble group. Further, IRF5 was verified to sever as the target gene of miR-146b. Conclusion miR-146b negatively regulated the inflammatory cytokine and inhibited the inflammation.
出处
《中国实验诊断学》
2017年第6期1076-1082,共7页
Chinese Journal of Laboratory Diagnosis
