The potential function of miR-135b-mediated JAK2/STAT3 signaling pathway during osteoblast differentiation.

Abstract

MC3T3-E1 cells were divided into Blank, miR-135b mimics, miR-135b inhibitors, AG490, and miR-135b inhibitors + AG490 groups. Cell viability was determined by MTT, alkaline phosphatase (ALP) activity by the corresponding kit, and mineralization by alizarin red staining. Furthermore, miR-135b, osteoblast-specific genes, and JAK2/STAT3 were detected through quantitative real-time polymerase chain reaction and Western blotting. MiR-135b downregulation was identified with increased JAK2 during osteoblast differentiation. JAK2 was confirmed as a target gene of miR-135b by dual-luciferase reporter assay. MC3T3-E1 cells in both miR-135b mimics and AG490 groups manifested decrease in cell viability, ALP activity, and mineralized nodes, as well as reductions in osteoblast-specific genes and proteins of JAK2, p-JAK2, and p-STAT3, but increase in cell apoptosis. However, opposite changes of the above factors were shown in cells from miR-135b inhibitors group. Notably, AG490 could reverse promotion effects of miR-135b inhibitors on osteoblast differentiation. Inhibiting miR-135b could activate the JAK2/STAT3 signaling pathway, thereby improving the cell viability and promoting the osteoblast differentiation.