Figure 4.

Icariin-induced osteogenesis was inhibited after cAMP/PKA/CREB signaling was blocked. A, the protein expression levels of phosphorylated PKA (p-PKA), phosphorylated CREB (p-CREB), COL-1, and RUNX2 in the cells pretreated by 2′,3′-dideoxyadenosine (DDA) (an adenylyl cyclase inhibitor) and treated with/without icariin. B, ALP activities after 3 and 6 days. C, the relative mRNA expression levels of ALP, COL1α2, and RUNX2. D, representative images of calcified nodules formed after 12 days. E and F, the numbers and areas of calcified nodules. Each experiment was conducted at least three times independently. Data are represented as mean ± S.D. (n = 3); *, p < 0.05; **, p < 0.01; ***, p < 0.001, when compared with control (Ctrl) or icariin group.