INSERM U606, University Paris Diderot, and Hospital Lariboisière, Paris, France.
J Cell Biochem. 2012 Jun;113(6):2047-56. doi: 10.1002/jcb.24076.
Mesenchymal stem cells (MSC) can differentiate into osteoblasts upon activation of Wnt signaling. Identifying targets of Wnt signaling in MSC may help promote MSC osteoblast differentiation for bone regeneration. In this study, using microarray analysis we found that Wnt3a upregulates neuregulin 1 (NRG-1) during Wnt3a-induced osteoblast differentiation in primary human MSC and murine C3H10T1/2 mesenchymal cells. Western blot and qPCR analyses confirmed that NRG-1 is upregulated by Wnt3a, and that this effect was counterbalanced by decreased expression of the NRG-1 receptor ErbB3. Consistently, exogenous NRG-1 had no effect on alkaline phosphatase (ALP) activity, an early marker of osteoblast differentiation. In contrast, small interfering RNA-mediated silencing of endogenous NRG-1 increased basal and Wnt3a-induced ALP activity in MSC. We showed that short hairpin (sh) ErbB3 and Wnt3a additively increased β-catenin transcriptional activity and ALP activity in MSC. These effects were abrogated by DKK1, indicating that cross-talk between Wnt3a and ErbB3 control MSC osteoblast differentiation via Wnt/β-catenin signaling. Furthermore, ErbB3 silencing decreased Src expression. Pharmacological inhibition of Src signaling promoted ErbB3- and Wnt-induced ALP activity, suggestive of a role of Src signaling in the modulation of osteoblast differentiation by ErbB3 and Wnt3a. The results indicate that downregulation of ErbB3 induced by Wnt3a contributes to Wnt3a-induced early osteoblast differentiation of MSCs through increased canonical Wnt/β-catenin signaling and decreased Src signaling.
间充质干细胞 (MSC) 在 Wnt 信号激活时可分化为成骨细胞。鉴定 MSC 中 Wnt 信号的靶点可能有助于促进 MSC 成骨细胞分化以实现骨再生。在这项研究中，我们通过微阵列分析发现 Wnt3a 在人原代 MSC 和鼠 C3H10T1/2 间充质细胞的 Wnt3a 诱导的成骨细胞分化过程中上调神经调节蛋白 1 (NRG-1)。Western blot 和 qPCR 分析证实，Wnt3a 上调了 NRG-1 的表达，而这种作用被 NRG-1 受体 ErbB3 表达的下调所抵消。一致地，外源性 NRG-1 对碱性磷酸酶 (ALP) 活性（成骨细胞分化的早期标志物）没有影响。相反，内源性 NRG-1 的小干扰 RNA 介导的沉默增加了 MSC 的基础和成骨细胞诱导的 ALP 活性。我们表明，短发夹 (sh) ErbB3 和 Wnt3a 可协同增加 MSC 中的β-连环蛋白转录活性和 ALP 活性。这些作用被 DKK1 阻断，表明 Wnt3a 和 ErbB3 之间的串扰通过 Wnt/β-连环蛋白信号控制 MSC 成骨细胞分化。此外，ErbB3 沉默降低了Src 的表达。Src 信号的药理学抑制促进了 ErbB3 和 Wnt 诱导的 ALP 活性，提示 Src 信号在 ErbB3 和 Wnt3a 调节成骨细胞分化中的作用。结果表明，Wnt3a 下调 ErbB3 有助于通过增加经典 Wnt/β-连环蛋白信号和降低 Src 信号来诱导 MSC 的早期成骨细胞分化。
