ABSTRACT
Objective To investigate the expression of microRNA-146a-3p(miR-146a-3p) and interleukin 17 (IL-17) in peripheral blood CD4+ T cells of neonates with sepsis (NS) and its regulation mechanism. Methods The expression of miR-146a-3p and IL-17 mRNA in CD4+ T cells of 66 children with sepsis (septicemia group), 40 children with infectious diseases (infection group), and 40 healthy newborns (control group) were detected by real-time quantitative PCR, and the level of IL-17 in peripheral blood was detected by ELISA. The targeting effect of the miR-146a-3p on IL-17 was verified by the dual luciferase reporter assay. After isolation of the CD4+ T cells, the expression of miR-146a-3p in CD4+ T cells was promoted or inhibited by miR-146a-3p mimic or miR-146a-3p inhibitor. The proportion of Th17 cells was detected by flow cytometry, and the methylation of miR-146a-3p gene was detected by methylation specific PCR. The changes of Th17 cell proportion and IL-17 expression were observed after adding methylation inhibitor 5-Aza-2'-deoxycytidine (5-Aza-dC) to CD4+ T cells. Results Compared with those in the infection group and the control group, the expression of miR-146a-3p in peripheral blood CD4+ T cells decreased, while the expression of IL-17 mRNA and the level of IL-17 in peripheral blood increased in septicemia group. Transfection of miR-146a-3p mimic in CD4+ T cells significantly inhibited the activity of wild-type luciferase in the 3'UTR of IL-17. After transfection of miR-146a-3p mimic, the expression of IL-17 mRNA in CD4+ T cells, and the level of IL-17 and the proportion of Th17 cells in the supernatant were significantly decreased. After transfection of miR-146a-3p inhibitor, the expression of IL-17 mRNA, and the level of IL-17 and the proportion of Th17 cells in the supernatant were increased. The methylation rate of miR-146a-3p gene promoter in the peripheral blood of the septicemia group was significantly higher than those of the control group and the infection group. After the addition of 5-Aza-dC in CD4+ T cells, the expression of miR-146a-3p was increased and the expression of IL-17 mRNA was decreased, and in the supernatant, the level of IL-17 was decreased and the proportion of Th17 cells was significantly reduced. Conclusion The expression of miR-146a-3p is down-regulated and the expression of IL-17 is up-regulated in peripheral blood CD4+ T cells of children with neonatal sepsis.
