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Year : 2018  |  Volume : 14  |  Issue : 58  |  Page : 641-646

Osthole promote differentiation and inhibit proliferation of osteoblast by activating wnt signaling and endoplasmic reticulum stress

1 Laboratory of New Techniques of Restoration and Reconstruction of Orthopedics and Traumatology, Institute of Traumatology and Orthopedics, Nanjing University of Chinese Medicine, Nanjing 210029, China
2 Laboratory of New Techniques of Restoration and Reconstruction of Orthopedics and Traumatology, Institute of Traumatology and Orthopedics, Nanjing University of Chinese Medicine; Department of Traumatology and Orthopedics, Affiliated Hospital of Nanjing University of Chinese Medicine, Nanjing 210029, China

Correspondence Address:
Yang Guo
138 Xianlin Road, Nanjing 210023
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Source of Support: None, Conflict of Interest: None

DOI: 10.4103/pm.pm_591_17

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Background: Osthole is extracted from Fructus Cnidii and is proved to be effective in the treatment of osteoporosis in rats. However, data are still scarce and the mechanism remains elusive. Objective: To investigate the effect of Osthole on proliferation and differentiation of osteoblast. Materials and Methods: Cells were divided into five groups: control group, β-estradiol group (10−8 M), and Osthole groups (10−6 M, 10−5 M, and 10−4 M). Osteoblast proliferation was evaluated by Cell Counting Kit-8 (CCK-8) assay. Alkaline phosphatase (ALP) activity was detected by ALP staining and enzymatic measurement. Mineralization was detected by alizarin-red staining. The level of osteocalcin was measured by enzyme-linked immunosorbent assay (ELISA). The expression of key proteins of Wnt/β-catenin signaling pathway and endoplasmic reticulum stress (ERS) was analyzed by Western blot. Results: Cell proliferation was retarded in moderate-dose and high-dose Osthole group at 1d, 2d and 3d and in low-dose Osthole group at 1d and 2d (P < 0.05). ALP activity was enhanced in high-dose Osthole group from 1d to 3d and in moderate-dose Osthole group at 2d (P < 0.05). Mineralization of bone matrix was promoted in high-dose Osthole group at 21d (P < 0.05). The secretion of osteocalcin was promoted in Osthole groups at 21d (P < 0.05). Expression of CHOP, GRP78, PDI, Wnt1, and β-catenin was upregulated in high-dose Osthole group at 2d, indicating that both of ERS and Wnt/β-catenin signaling pathway were activated. Conclusion: It can be concluded that the effect of Osthole on inhibition of proliferation is relevant with activation of ERS, and activation of Wnt/β-catenin signaling pathway is one of the mechanisms how Osthole promotes osteoblast differentiation. In summary, this study provided more evidence for Osthole as a potential anti-osteoporosis medicine. Abbreviations used: ALP: Alkaline phosphatase; ELISA: Enzyme-linked immunosorbent assay; CCK-8: Cell Counting Kit-8; ERS: endoplasmic reticulum stress; DMEM: Dulbecco's Modified Eagle Medium; α-MEM: α-minimum essential medium; EDTA: trypsin/ethylenediaminetetraacetic acid; DMF: N,N-dimethylformamide; TBS-T: Tris-buffered saline-Tween 20; RIPA: Radio Immuno Precipitation Assay; HRP: Horseradish peroxidase; GAPDH: Glyceraldehyde 3-phosphate dehydrogenase; BCA: Bicinchoninic acid; ANOVA: Analysis of variance.

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