Verani R, Cappuccio I, Spinsanti P, Gradini R, Caruso A, Magnotti M C, Motolese M, Nicoletti F, Melchiorri D
Department of Human Physiology and Pharmacology, University of Rome La Sapienza, Rome, Italy.
J Neurochem. 2007 Jan;100(1):242-50. doi: 10.1111/j.1471-4159.2006.04207.x. Epub 2006 Oct 25.
Cultured mouse D3 embryonic stem (ES) cells differentiating into embryoid bodies (EBs) expressed several Wnt isoforms, nearly all isotypes of the Wnt receptor Frizzled and the Wnt/Dickkopf (Dkk) co-receptor low-density lipoprotein receptor-related protein (LRP) type 5. A 4-day treatment with retinoic acid (RA), which promoted neural differentiation of EBs, substantially increased the expression of the Wnt antagonist Dkk-1, and induced the synthesis of the Wnt/Dkk-1 co-receptor LRP6. Recombinant Dkk-1 applied to EBs behaved like RA in inducing the expression of the neural markers nestin and distal-less homeobox gene (Dlx-2). Recombinant Dkk-1 was able to inhibit the Wnt pathway, as shown by a reduction in nuclear beta-catenin levels. Remarkably, the antisense- or small interfering RNA-induced knockdown of Dkk-1 largely reduced the expression of Dlx-2, and the neuronal marker beta-III tubulin in EBs exposed to RA. These data suggest that induction of Dkk-1 and the ensuing inhibition of the canonical Wnt pathway is required for neural differentiation of ES cells.
培养的小鼠D3胚胎干细胞分化形成胚状体（EBs）时，表达了几种Wnt亚型、几乎所有Wnt受体卷曲蛋白（Frizzled）的同种型以及Wnt/ Dickkopf（Dkk）共受体低密度脂蛋白受体相关蛋白5型（LRP）。用视黄酸（RA）进行为期4天的处理，可促进EBs的神经分化，显著增加Wnt拮抗剂Dkk-1的表达，并诱导Wnt/ Dkk-1共受体LRP6的合成。应用于EBs的重组Dkk-1在诱导神经标志物巢蛋白和远端缺失同源框基因（Dlx-2）的表达方面表现得与RA相似。重组Dkk-1能够抑制Wnt信号通路，这表现为细胞核内β-连环蛋白水平降低。值得注意的是，在暴露于RA的EBs中，反义或小干扰RNA诱导的Dkk-1敲低大大降低了Dlx-2和神经元标志物β-III微管蛋白的表达。这些数据表明，ES细胞的神经分化需要诱导Dkk-1并随后抑制经典Wnt信号通路。
