Canalis Ernesto, Zanotti Stefano, Smerdel-Ramoya Anna
Department of Research, Saint Francis Hospital and Medical Center, Hartford, CT, 06105; The University of Connecticut School of Medicine, Farmington, CT, 06030.
Department of Research, Saint Francis Hospital and Medical Center, Hartford, CT, 06105.
Bone. 2014 Jul;64:273-80. doi: 10.1016/j.bone.2014.04.028. Epub 2014 May 2.
Connective tissue growth factor (Ctgf) or CCN2 is a protein synthesized by osteoblasts necessary for skeletal homeostasis, although its overexpression inhibits osteogenic signals and bone formation. Ctgf is induced by bone morphogenetic proteins, transforming growth factor β and Wnt; and in the present studies, we explored whether Notch regulated Ctgf expression in osteoblasts. We employed Rosa(Notch) mice, where the Notch intracellular domain (NICD) is expressed following the excision of a STOP cassette, placed between the Rosa26 promoter and NICD. Notch was activated by transduction of adenoviral vectors expressing Cre recombinase (Ad-CMV-Cre). Notch induced Ctgf mRNA levels in a time dependent manner and increased Ctgf heterogeneous nuclear RNA. Notch also destabilized Ctgf mRNA shortening its half-life from 13h to 3h. The effect of Notch on Ctgf expression was lost following Rbpjκ downregulation, demonstrating that it was mediated by Notch canonical signaling. However, downregulation of the classic Notch target genes Hes1, Hey1 and Hey2 did not modify the effect of Notch on Ctgf expression. Wild type osteoblasts exposed to immobilized Delta-like 1 displayed enhanced Notch signaling and increased Ctgf expression. In addition to the effects of Notch in vitro, Notch induced Ctgf in vivo, and calvariae and femurs from Rosa(Notch) mice mated with transgenics expressing the Cre recombinase in cells of the osteoblastic lineage exhibited increased expression of Ctgf. In conclusion, Ctgf is a target of Notch canonical signaling in osteoblasts, and may act in concert with Notch to regulate skeletal homeostasis.
结缔组织生长因子（Ctgf）或CCN2是成骨细胞合成的一种蛋白质，对骨骼稳态至关重要，尽管其过度表达会抑制成骨信号和骨形成。Ctgf由骨形态发生蛋白、转化生长因子β和Wnt诱导产生；在本研究中，我们探讨了Notch是否调节成骨细胞中Ctgf的表达。我们使用了Rosa(Notch)小鼠，在该小鼠中，Notch细胞内结构域（NICD）在位于Rosa26启动子和NICD之间的STOP盒切除后表达。通过转导表达Cre重组酶的腺病毒载体（Ad-CMV-Cre）激活Notch。Notch以时间依赖性方式诱导Ctgf mRNA水平，并增加Ctgf不均一核RNA。Notch还使Ctgf mRNA不稳定，将其半衰期从13小时缩短至3小时。Rbpjκ下调后，Notch对Ctgf表达的影响消失，表明其由Notch经典信号介导。然而，经典Notch靶基因Hes1、Hey1和Hey2的下调并未改变Notch对Ctgf表达的影响。暴露于固定化Delta样1的野生型成骨细胞显示出增强的Notch信号和增加的Ctgf表达。除了Notch在体外的作用外，Notch在体内也诱导Ctgf表达，与在成骨细胞谱系细胞中表达Cre重组酶的转基因小鼠交配的Rosa(Notch)小鼠的颅骨和股骨显示出Ctgf表达增加。总之，Ctgf是成骨细胞中Notch经典信号的靶标，可能与Notch协同作用以调节骨骼稳态。
