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Updated: Jun 21, 2026
Quantitative PCR-based Assay to Measure Sonic Hedgehog Signaling in Cellular Model of Ciliogenesis
Published on: January 31, 2025
[Hedgehog pathway is associated with the proliferation and differentiation of osteoblasts]
Lei Han1, Xiao-ling Zhang, Xuan Li
1Department of Orthodontics, College of Stomatology, Ninth People's Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai Key Laboratory of Stomatology, Shanghai 200011, China. pinedream@163.com
Purpose:
The purpose of this study was to evaluate the effects of Hedgehog signaling pathway on osteoblast proliferation and differentiation.
Methods:
Primary osteoblasts harvested from newborn rat calvarial bone were cultured for 21 days. The temporary expression of Shh and Ihh was detected by RT-PCR. The culture medium was then treated with either a recombinant Shh N-terminals protein (N-Shh) or cyclopamine (cy), a Hedgehog inhibitor. The proliferation of osteoblasts was quantified by MTT and FCM. The osteoblast differentiation and matrix mineralization were evaluated by ALP and Alizarin-Red staining. The expression of Ptch and Smo was quantified by real-time PCR.SAS 8.0 software package was used for statistical analysis.
Results:
Osteoblast expressed both Shh and Ihh. The expression level of Shh decreased with culture time while the amount of Ihh expression increased. N-Shh treatment increased the cell population and the number of cells in S phase (P<0.05. N-Shh also promoted ALP activities (P<0.05) and bone matrix calcification. Moreover, N-Shh application led to an increase in Ptch and Smo mRNA level (P<0.05). Treatment with cy, on the other side, inhibited osteoblast proliferation and differentiation(P<0.05).
Conclusion:
Heghehog signaling pathway is actively involved in regulating osteoblast proliferation and differentiation.
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