Figure 5.

miR-142a-3p induced NET formation via direct targeting of WASL

(A) WASL mRNA expression was analyzed by qRT-PCR (n = 7–10 mice per group). (B) Wild-type and mutated m-WASL-3′ untranslated regions (UTRs) were cloned into psi-CHECK-2, and four predicted binding sites of miR-142a-3p were identified in the 3′ UTR of the WASL gene; dual-luciferase reporter assay performed on HEK293T cells transfected with WASL UTR reporter plasmid together with miR-142a-3p mimic or control mimic. (C and D) Expression of N-WASP (translated from WASL) in mice livers was analyzed by western blotting and immunohistochemistry (n = 6 mice per group). (E) Neutrophils of infected mice were isolated, and the expression of N-WASP was analyzed by immunohistochemistry. (F and G) Neutrophils were treated with PMA (500 nM, 4 h) or Wiskostatin (N-WASP inhibitor, 20 μM, 24 h) + PMA (500 nM, 4 h), and NET formation was evaluated by SEM and immunohistochemistry. (A, C, and D–G) One-way ANOVA with Dunnett’s multiple comparison test. (B) Unpaired two-sample t test.