Fig. 3: miR-199a-5p overexpression repressed Th17 cell differentiation through targeting STAT3.

A The above: the potential binding sites between miR-199a-5p and the 3′UTR region of STAT3 mRNA. The below: relative luciferase reporter activities of STAT3 3′UTR wild type (WT) and mutant type (MUT) were detected by dual-luciferase reporter gene assay. B miR-199a-5p expression and C STAT3 expression were detected in CD4⁺T cells transfected with miR-199a-5p agomir/antagomir/their negative controls (agomir NC or antagomir NC). CD4⁺T cells were transfected with pMIG-STAT3 or its negative control (pMIG) or agomir NC + pMIG-STAT3 or miR-199a-5p agomir +pMIG-STAT3, 48 h later, CD4⁺T cells were underwent induction of Th17 differentiation. The protein levels of STAT3, pSTAT3, and RORγt in CD4⁺T cells were detected by western blot. The quantification of band densitometry was shown in (D) and the representative bands were shown in (E). GAPDH and β-actin were used as internal controls. F The IL-17 level was measured in the supernate of cells using ELISA. The population of Th17 cells was measured in CD4⁺T cells using flow cytometry. The quantitative result was shown in ((G); ***P < 0.001; ^##P < 0.01) and representative histograms were shown in (H). ***P < 0.001 vs. agomir NC or pMIG, ^#P < 0.05, ^##P < 0.01, ^###P < 0.001 vs. antagomir NC or agomir NC + STAT3.