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. 2019 Jun 12;17:223–234. doi: 10.1016/j.omtn.2019.06.001

Figure 4.

Figure 4

miRNA-26a-5p Targets PTEN to Functionally Inhibit Osteoblast Activity In Vitro

(A) Luciferase reporter constructs containing either a WT PTEN 3′ UTR (WT 3′ UTR) or this same region after site-directed mutagenesis (3′ UTR-Mut). Luc, luciferase. (B) Endogenous miRNA-26a-5p effects in MC3T3-E1 cells on WT PTEN 3′ UTR (luc-UTR), the PTEN 3′ UTR mutant (luc-UTR-Mut), as assessed after antagomiR-NC or antagomiR-26a-5p treatment. (C and D) qPCR (C) and western blot analysis (D) were used after control, agomiR-26a-5p, agomiR-NC, and antagomiR-26a-5p, and antagomiR-NC transfection to assess the expression of PTEN. (E) PTEN expression in calluses of control, fracture, and fracture+TBI patient groups. (F) PTEN expression in calluses of fracture and fracture+TBI mouse groups. (G and H) qPCR (G) and western blot analysis (H) were used after control, siRNA-NC, and siRNA-PTEN transfection to assess Col1a1, ALP, OCN, and Runx2 expression, with a scrambled siRNA used as the negative control (siRNA-NC). Data are means ± SD of triplicate experiments. *p < 0.05, **p < 0.01, ***p < 0.001.