Elucidation of molecular mechanism involved in mechanical stress signaling in bone - functional analysis, contribution to pathology, and cloning of related molecule of a novel gene Znt5 -

我们最近分离了一个新的基因Znt5，其表达随着兔主动脉扩张负荷的增加而增加。Znt5作为锌转运体，与野生型（WT）小鼠相比，Znt5^<-/->小鼠表现出轻微的生长迟缓和骨质疏松症。骨质流失似乎是由于成骨细胞功能障碍所致。为了了解该分子在骨机械应力信号中的作用，我们首先使用几个模型检查了骨对机械应力的表达。在Flexor cell系统拉伸刺激下培养MC3T3-E1、MLO-Y4和MLO-A5三种成骨细胞时，Znt5表达仅在MLO-Y4细胞中增加约2倍，MLO-Y4细胞是已知分化程度最高的成骨细胞。在Flexor cell系统拉伸刺激下培养MC3T3-E1、MLO-Y4和MLO-A5 3种成骨细胞时，Znt5的表达仅在MLO-Y4细胞中增加了约2倍，MLO-Y4细胞是分化程度最高的成骨细胞。在小鼠新生颅骨培养中，通过插入弹簧连续拉应力诱导Znt5表达9 h。在小鼠尾悬模型中，卸压3周后，小鼠胫骨和股骨中Znt5 mRNA水平下降至对照组的三分之一左右，重新加载后恢复到正常水平。当Znt5^<-/->小鼠尾悬处理3周时，骨密度下降（-5%）远小于WT组（-15%）。在胫骨骨折模型和实验性膝关节OA模型中，WT和Znt5^<-/->小鼠的骨折愈合和软骨破坏情况相似。为了探究Znt5的下游信号通路，我们比较了Znt5^<-/->和WT骨骼的基因表达谱，发现Zn5^<-/->中约有20个基因表达减少。然而，对这些mRNA表达的实时PCR分析显示，Znt5^<-/->与WT之间没有显著差异。我们认为，Znt5的表达在成熟成骨细胞或骨细胞中受到机械应力的正向调节，在机械应力维持骨体积中起着至关重要的作用。少

We recently isolated a novel gene Znt5 whose expression increased in response to expansion load of rabbit aorta. Znt5 functions as a zinc transporter and Znt5^<-/-> mice exhibited slight growth retardation and osteoporosis as compared to wild-type (WT) littermates. The bone loss seemed to be due to a dysfunction of osteoblasts. To learn the involvement of this molecule in the mechanical stress signaling in bone, we first examined the expression in bone responding to mechanical stress using several models. When three kinds of osteoblastic cell lines, MC3T3-E1,MLO-Y4 and MLO-A5,were cultured with stretching stimulation by the Flexor Cell system, about 2-fold increase of Znt5 expression was seen only in MLO-Y4 cells that are known to be the most differentiated osteoblasts. When three kinds of osteoblastic cell lines, MC3T3-E1,MLO-Y4 and MLO-A5,were cultured with stretching stimulation by the Flexor Cell system, about 2-fold increase of Znt5 expression was seen only in MLO-Y4 cells that ar … More e known to be the most differentiated osteoblasts. In the culture of mouse newborn calvariae, Znt5 expression was induced by continuous tensile stress by an inserted spring for 9 h. In the mouse tail suspension model, Znt5 mRNA level in tibiae and femora was decreased to about one third that of control after 3 weeks of unloading, and was restored to the normal level by reloading. When Znt5^<-/-> mice were treated with tail suspension for 3 weeks, the decrease in bone density (-5%) was much less than that in WT (-15%). In a tibial fracture model and an experimental knee OA model, fracture healing and cartilage destruction, respectively, was similarly seen in WT and Znt5^<-/-> mice. To explore the downstream signaling of Znt5,we compared the gene expression profile between Znt5^<-/-> and WT bones, and found that about 20 genes were decreased in Zn5^<-/->. However, real-time PCR analyses of those mRNA expressions revealed no significant differences between Znt5^<-/-> and WT. We conclude that Znt5, whose expression is positively regulated by mechanical stress in mature osteoblasts or osteocytes, plays an essential role in the maintenance of bone volume by mechanical stress. Less