Clines Gregory A, Mohammad Khalid S, Bao Yongde, Stephens Owen W, Suva Larry J, Shaughnessy John D, Fox Jay W, Chirgwin John M, Guise Theresa A
Department of Internal Medicine, Division of Endocrinology and Metabolism, Aurbach Laboratory, The University of Virginia, P.O. Box 801419, Charlottesville, Virginia 22908-1419, USA.
Mol Endocrinol. 2007 Feb;21(2):486-98. doi: 10.1210/me.2006-0346. Epub 2006 Oct 26.
Tumor-produced endothelin-1 (ET-1) stimulates osteoblasts to form new bone and is an important mediator of osteoblastic bone metastasis. The anabolic actions of ET-1 in osteoblasts were investigated by gene microarray analyses of murine neonatal calvarial organ cultures. Targets of ET-1 action were validated by real-time RT-PCR in murine primary osteoblast cultures. IL-6, IL-11, the CCN (CYR61, CTGF, NOV) family members cysteine-rich protein 61 and connective tissue growth factor, inhibin beta-A, serum/glucocorticoid regulated kinase, receptor activator of nuclear factor kappaB ligand, snail homolog 1, tissue inhibitor of metalloproteinase 3, and TG-interacting factor transcripts were increased by ET-1. ET-1 decreased the transcript for the Wnt signaling pathway inhibitor, dickkopf homolog 1 (Dkk1). Calvarial organ cultures treated with ET-1 had lower concentrations of DKK1 protein in conditioned media than control cultures. High DKK1 concentrations in bone marrow suppress bone formation in multiple myeloma. We hypothesized that the converse occurs in osteoblastic bone metastasis, where ET-1 stimulates osteoblast activity by reducing autocrine production of DKK1. Recombinant DKK1 blocked ET-1-mediated osteoblast proliferation and new bone formation in calvarial organ cultures, whereas a DKK1-neutralizing antibody increased osteoblast numbers and new bone formation. ET-1 directed nuclear translocation of beta-catenin in osteoblasts, indicating activation of the Wnt signaling pathway. The data suggest that ET-1 increases osteoblast proliferation and new bone formation by activating the Wnt signaling pathway through suppression of the Wnt pathway inhibitor DKK1.
肿瘤产生的内皮素-1（ET-1）刺激成骨细胞形成新骨，是成骨性骨转移的重要介质。通过对小鼠新生颅骨器官培养物进行基因微阵列分析，研究了ET-1在成骨细胞中的合成代谢作用。通过实时逆转录聚合酶链反应在小鼠原代成骨细胞培养物中验证了ET-1作用的靶点。ET-1可增加白细胞介素-6、白细胞介素-11、CCN（富半胱氨酸蛋白61、结缔组织生长因子、肾母细胞瘤过度表达基因）家族成员富半胱氨酸蛋白61和结缔组织生长因子、抑制素β-A、血清/糖皮质激素调节激酶、核因子κB受体活化因子配体、蜗牛同源物1、金属蛋白酶组织抑制剂3和TG相互作用因子的转录本。ET-1降低了Wnt信号通路抑制剂Dickkopf同源物1（Dkk1）的转录本。用ET-1处理的颅骨器官培养物的条件培养基中DKK1蛋白浓度低于对照培养物。骨髓中高浓度的DKK1可抑制多发性骨髓瘤中的骨形成。我们推测，在成骨性骨转移中情况相反，ET-1通过减少DKK1的自分泌产生来刺激成骨细胞活性。重组DKK1可阻断ET-1介导的颅骨器官培养物中成骨细胞的增殖和新骨形成，而DKK1中和抗体则增加了成骨细胞数量和新骨形成。ET-1促使β-连环蛋白在成骨细胞中发生核转位，表明Wnt信号通路被激活。数据表明，ET-1通过抑制Wnt通路抑制剂DKK1来激活Wnt信号通路，从而增加成骨细胞增殖和新骨形成。
