2022, Vol. 26 ›› Issue (1): 64-69
Icariin promotes differentiation of mouse preosteoblasts via activating autophagy
Studies have found that icariin can promote the proliferation and differentiation of mouse preosteoblasts. Nuclear factor-κB signaling pathways and autophagy play an important role in the regulation of osteoblast differentiation. Whether they are involved in icariin promoting the proliferation and differentiation of mouse preosteoblasts remains unclear.
To investigate the effect of icariin on the proliferation and differentiation of mouse preosteoblasts and the role of nuclear factor-κB signaling and autophagy.
(1) Screening out the best intervention concentration of icariin: Mouse preosteoblasts MC3T3-E1 cells were divided into control group and various icariin groups (0.01, 0.1, 1, 10, 100 μmol/L). MTS was used to detect the proliferation activity of mouse preosteoblasts from different groups after icariin intervention. Osteogenic differentiation was assessed by Alizarin red staining. The osteoblastic differentiation ability and autophagy activity in mouse preosteoblasts from different groups were evaluated by western blot assay after icariin intervention. (2) 3-Methyladenine was added to inhibit autophagy. Four groups were set up: control group, 3-methyladenine group, icariin group, and 3-methyladenine + icariin group. Alizarin red staining was used to observe the mineralized nodules. Western blot assay was utilized to determine the expression levels of alkaline phosphatase, Runx2, p62, LC3, NF-κB p65 and p-NF-κB p65 protein.
(1) Icariin could promote the proliferation of mouse preosteoblasts and the effect was the most obvious at 10 μmol/L (P < 0.05). The formation of mineralized nodules in the 10 μmol/L icariin group was significantly more than that in the control group and the expression levels of alkaline phosphatase and Runx2 protein were significantly increased (P < 0.05). Compared with the control group, the expression levels of Beclin1, LC3 II/LC3 I increased significantly and the expression of p62 decreased in the 10 μmol/L icariin group (P < 0.05). (2) After 3-methyladenine inhibited autophagy, compared with the icariin group, the expression of alkaline phosphatase, Runx2, LC3 II/LC3 I decreased and the expression of p62 increased in the 3-methyladenine + icariin group (P < 0.05). (3) Compared with the control group, the expression of p-p65/p65 decreased in the 10 μmol/L icariin group (P < 0.05). (4) These results indicate that icariin may promote osteogenic differentiation of MC3T3-E1 cells by down-regulation of the nuclear factor-κB signaling and enhancing autophagy.
icariin, autophagy, osteoblasts, cell differentiation, nuclear factor-κB signaling pathway