Effects of icariin on proliferation, differentiation and mineralization of osteoblasts

Abstract

Objective To study the effect of icariin the proliferation, differentiation and mineralization of osteoblasts cultured in vitro. Methods The MC3T3-E1 cells were used in the experiment of mice as a source of osteoblast precursor cells, with a final concentration of 0, 10-4, 10-5, 10-6, 10-7, and 10-8 mol/L icariin. The ability of cell proliferation was measured by methyl thiazol tetrazolium (MTT) assay. The cell morphology and the cell growth density were observed by hematoxylin and eosin (HE) staining. Alkaline phosphatase reagent kit was used to test osteoblastic alkaline phosphatase activity for the evaluation of ability of early differentiation of bone cells. The alizarin red staining was used to evaluate the capability of mineralization of osteoblasts. Results The results of MTT showed that the absorbance (A) values in icariin groups (0, 10-4, 10-5, 10-6, 10-7, and 10-8 mol/L) were 0.54±0.04, 0.28±0.04, 0.40±0.02, 0.67±0.03, 0.58±0.01 and 0.55±0.03 respectively, and those in 10-6 mol/L icariin group were significantly higher than those in the rest groups with the difference being statistically significant (P=0.002). HE staining showed that the normal osteoblasts displayed typical spindle, triangular, polygonal and other irregular morphology. As compared with the blank control group, number of cells was greatest, and the cell distribution density was largest in the 10-6 mol/L icariin group. In 10-4 mol/L icarrin group, the expanding osteoblasts were rarely seen. In icariin groups (0, 10-4, 10-5, 10-6, 10-7, and 10-8 mol/L), alkaline phosphatase activity was (1.75±0.11), (0.85±0.35), (1.50±0.21), (2.80±0.12), (2.10±0.36) and (1.80±0.23) U/g prot respectively. Alkaline phosphatase activity was the highest in 10-6 mol/L icariin group (P=0.001), and that in 10-4 mol/L icariin group was the lowest with the difference being statistically significant (P=0.001). The alkaline phosphatase activity in 10-8 mol/L ocariin group was slightly higher than that in the blank group, but the difference was not statistically significant (P=0.292). Alizarin red staining showed that 10-6 mol/L icariin significantly promoted the mineralization of osteoblasts, and a large number of calcium nodules and clusters formed. In 10-4 mol/L icariin group, the epimedium calcium nodules were almost not found. Conclusion Icariin can promote the proliferation, differentiation and mineralization of osteoblasts. Key words: Osteoblast; Icariin; Osteoporosis