Fig 1: SPRY2 is a direct target of miR-21 in corneal epithelial cells. Corneal epithelial cells were treated with miR-21 inhibitor and exposed to TGF-β1 (T), 1% O2 (H), or TGF-β1 plus 1% O2 (T+H). The protein levels of SPRY1, SPRY2, SPRY4, and p-ERK, were determined by Western blotting (A) followed by densitometry analysis using ImageJ software (B). Tubulin was used as the loading control. (C) miR-21 and its putative binding sequences in the mouse 3′-UTR of Spry2. The binding sequences in the complementary sites were replaced by the indicated nucleotides to produce mutant Spry2 3′-UTR luciferase reporter constructs. Marked sequences represent the conserved complementary nucleotides of the miR-21 binding sequence in humans and various mammals. (D, E) Luciferase activity of the wild-type and mutated 3′-UTR of Spry2 treated with miR-21 mimic (D) or inhibitor (E). *P < 0.05, **P < 0.01, ***P < 0.001, versus the untreated group (B) or mimic NC (D) or inhibitor NC group (E). Experiments were performed in triplicate.