Regulatory effects of miR-138 and RUNX3 on Th1/Th2 balance in peripheral blood of children with cough variant asthma.

Abstract

To study the expression levels of microRNA-138 (miR-138) and Runt-related transcription factor 3 (RUNX3) in peripheral blood of children with cough variant asthma (CVA) and their regulatory effects on Th1/Th2 balance. Sixty-five children with CVA (CVA group) and 30 healthy children (control group) were enrolled. Peripheral venous blood samples were collected for both groups, and CD4+ T cells were isolated and cultured. Enzyme-linked immunosorbent assay was used to measure the levels of interferon (IFN)-γ, interleukin (IL)-2, IL-4, IL-5, and IL-13 that were secreted by CD4+ T cells. Flow cytometry was used to determine the percentages of Th1 and Th2 cells. Quantitative real-time PCR was used to measure the level of RUNX3 mRNA in CD4+ T cells and the level of miR-138 in peripheral blood. Western blot was used to determine the protein expression of RUNX3 in CD4+ T cells. The dual-luciferase reporter assay was used to determine the targeting effects of miR-138 and RUNX3. The RUNX3-mimic plasmid was transfected into CD4+ T cells, and the effects on the levels of IFN-γ, IL-2, IL-4, IL-5, and IL-13 and the percentages of Th1 and Th2 cells were measured. Compared with the control group, the CVA group showed significantly decreased levels of IFN-γ and IL-2 from CD4+ T cells, significantly increased levels of IL-4, IL-5, and IL-13 from CD4+ T cells, significantly decreased Th1 cell percentage and Th1/Th2 ratio, and a significantly increased Th2 cell percentage (P<0.05). The CVA group showed significantly lower relative expression levels of RUNX3 mRNA and protein in CD4+ T cells in peripheral blood than the control group (P<0.001). The relative expression level of miR-138 was significantly higher in the CVA group than in the control group (P<0.001). MiR-138 could target the expression of RUNX3. Upregulating the expression of RUNX3 in CD4+ T cells induced significantly increased levels of IFN-γ and IL-2, significantly decreased levels of IL-4, IL-5, and IL-13, significantly increased Th1 cell percentage and Th1/Th2 ratio, and a significantly decreased Th2 cell percentage (P<0.05). MiR-138 regulates Th1/Th2 balance by targeting RUNX3 in children with CVA, providing a new direction for the treatment of CVA.