Fig. 9

miRNA-22 inhibited the TGF-β signaling by targeting KDM3A. A The target sequence for miRNA-22-3p in the 3′-UTR of KDM3A and the mutated target sequence. B KDM3A-wt or KDM3A-mut was transfected into NSCs together with miRNA-22-3p mimics or mimics-NC. Dual-luciferase reporter analysis confirmed the direct recognition of the KDM3A 3′-UTR by miRNA-22-3p (n = 3; data are the mean ± S.D.; ∗p < 0.05, ns p > 0.05). C The transfection of miRNA-22-3p mimics and the coculture of LPS-As-NSCs-EVs into NSCs downregulated the expression of KDM3A and TGF-β after transfection or culture for 24 h. In contrast, the transfection of miRNA-22-3p inhibitors into NSCs upregulated the expression of these 2 genes in the presence of LPS-As-NSCs-EVs (n = 3; data are the mean ± S.D.; ∗p < 0.05). D Western blot analysis revealed that the expression of TGF-β and p-Smad 2 was downregulated by the transfection with miRNA-22-3p mimics or the coculture with LPS-As-NSCs-EVs for 24 h. Moreover, the LPS-As-NSCs-EVs-induced downregulation of TGF-β and p-Smad 2 expression was countered by the transfection with miRNA-22-3p inhibitors (n = 3; data are the mean ± S.D.; ∗p < 0.05). E Consistent with the in vitro data, the injection of miRNA-22-3p agomir or LPS-As-NSCs-EVs reduced the expression of TGF-β and p-Smad 2 at different time points following SCI. Moreover, this LPS-As-NSCs-EVs-induced mediation of the expression of TGF-β and p-Smad 2 was partly abolished by injection of the miRNA-22-3p antagomir (n = 3; data are the mean ± S.D.; ∗p < 0.05)