Research Paper Volume 12, Issue 13 pp 13477—13487

Hydrostatic pressure induces osteogenic differentiation of adipose-derived mesenchymal stem cells through increasing lncRNA-PAGBC

Figure 4. MiR-133b inhibits RUNX2 in AMSCs. (A) Bioinformatic prediction of the binding site for miR-133b on the 3’-UTR of RUNX2, which was used for generating 3'-UTR of wildtype RUNX2 mRNA (RUNX2 wt) and 3'-UTR of RUNX2 mRNA with a mutant at miR-133b-binding site (RUNX2 mut). (B) Plasmids that overexpress miR-133b or deplete it were prepared. Plasmids carrying a null sequence were used as controls. These plasmids were used to transfect AMSCs and RT-qPCR for miR-133b was assessed in these cells. (C, D) We prepared luciferase reporter for wildtype (wt) RUNX2 and RUNX2 with a mutant at the miR-133b binding site (mut). The luciferase reporter assay was performed on AMSCs, using either miR-133b (C) or as-miR-133b (D). N=5. *p<0.05. NS: non-significant.