Liu Yingjie, Huang Lulu, Hao Baohui, Li Hao, Zhu Shuanglong, Wang Qiangsong, Li Ruixin, Xu Yunqiang, Zhang Xizheng
Tianjin Medical University General Hospital, Tianjin, China.
Institute of Medical Equipment, Academy of Military Medical Science, Tianjin, China.
Cell Physiol Biochem. 2017;41(4):1605-1615. doi: 10.1159/000470896. Epub 2017 Mar 28.
BACKGROUND/AIMS: Mechanical loading plays an important role in the regulation of bone mass. However, bone cells are not always under physiological stress. In some cases, bone tissue is subjected to an overloaded mechanical environment. For example, a person who is weight training and a stevedore often experience bone pain, inflammation and other bone fatigue damage symptoms. Icariin is the major ingredient of Herba epimedii, which has been widely used for the treatment of bone injury in traditional Chinese medicine, but its mechanism remains unknown. The aim of this study was to probe the effect of icariin on the proliferation and differentiation of osteoblasts exposed to overload and to determine whether the Wnt/β-catenin signalling pathway is involved in the drug response in osteoblasts.
Mouse MC3T3-E1 cells were exposed to mechanical tensile strain using a four- point bending device to create an overload damage model. An MTT assay was performed to determine the effects of icariin on MC3T3-E1 cell proliferation. The mRNA and protein levels of ALP, COL-I, OCN, RUNX2 and β-catenin were assessed using RT-PCR and immunoblotting. The protein levels of β-catenin in the MC3T3-E1 cells were also determined using fluorescence microscopy. The mineralization of osteoblasts was assessed using Alizarin Red S staining.
We found that icariin enhanced the proliferation of osteoblasts exposed to overload and promoted MC3T3-E1 cell differentiation and mineralization. Furthermore, the gene and protein expression levels of β-catenin and RUNX2 all increased with icariin treatment compared with those in the damage group.
Our study suggested that icariin promotes proliferation and differentiation in osteoblasts exposed to overload. The effect of icariin on osteoblastic differentiation acted by activating the RUNX2 promoter and the Wnt/β- catenin pathway.
背景/目的：机械负荷在骨量调节中起重要作用。然而，骨细胞并非总是处于生理应激状态。在某些情况下，骨组织会受到过载的机械环境影响。例如，进行重量训练的人和搬运工人经常会经历骨痛、炎症及其他骨疲劳损伤症状。淫羊藿苷是淫羊藿的主要成分，在传统中药中已被广泛用于治疗骨损伤，但其作用机制尚不清楚。本研究旨在探讨淫羊藿苷对暴露于过载环境下的成骨细胞增殖和分化的影响，并确定Wnt/β-连环蛋白信号通路是否参与成骨细胞对该药物的反应。
使用四点弯曲装置对小鼠MC3T3-E1细胞施加机械拉伸应变，以建立过载损伤模型。采用MTT法检测淫羊藿苷对MC3T3-E1细胞增殖的影响。通过RT-PCR和免疫印迹法评估碱性磷酸酶（ALP）、I型胶原蛋白（COL-I）、骨钙素（OCN）、Runx2和β-连环蛋白的mRNA和蛋白水平。还使用荧光显微镜测定MC3T3-E1细胞中β-连环蛋白的蛋白水平。通过茜素红S染色评估成骨细胞的矿化情况。
我们发现淫羊藿苷增强了暴露于过载环境下的成骨细胞的增殖，并促进了MC3T3-E1细胞的分化和矿化。此外，与损伤组相比，淫羊藿苷处理后β-连环蛋白和Runx2的基因和蛋白表达水平均升高。
我们的研究表明，淫羊藿苷促进暴露于过载环境下的成骨细胞的增殖和分化。淫羊藿苷对成骨细胞分化的作用是通过激活Runx2启动子和Wnt/β-连环蛋白通路实现的。
