锂剂通过β-catenin/Wnt调控内源性间充质干细胞预防/治疗激素型股骨头坏死的实验研究

项目名称： 锂剂通过β-catenin/Wnt调控内源性间充质干细胞预防/治疗激素型股骨头坏死的实验研究

项目编号： No.81472066

项目类型： 面上项目

立项/批准年度： 2015

项目学科： 医药、卫生

项目作者： 张长青

作者单位： 上海交通大学

项目金额： 90万元

中文摘要： 激素型股骨头坏死（ONFH）目前尚无法通过药物预防或早期治疗，局部间充质干细胞（MSCs）增殖能力下降及分化异常是重要的发病因素。我们发现锂剂可通过激活MSCs增殖及分化重要调控通路β-catenin/Wnt，促进体外MSCs增殖并拮抗激素的抑制作用。据此推测锂剂对激素型ONFH可能具有预防或治疗作用。本课题拟在体外继续探讨锂剂拮抗激素对MSCs分化的影响，通过蛋白过表达及SiRNA技术正负向调控β-catenin/Wnt通路活性，探究锂剂拮抗激素的分子机制；体内建立他莫昔芬可诱导性β-catenin-LoxP+/-转基因小鼠模型，特异性地在MSCs瞬时过表达或敲除β-catenin蛋白，探究锂剂调控内源性MSCs，拮抗激素预防ONFH的效果及分子学机制；最后在经典的兔激素型ONFH模型中验证锂剂疗效。从调控内源性MSCs增殖及分化入手，为临床预防/早期治疗激素型ONFH进行新的探索。

中文关键词： 股骨头坏死；骨髓间充质干细胞；骨坏死；干细胞；间充质干细胞

英文摘要： There are currently no effective way to prevent or treat steroid induced osteonecrosis of femoral head (ONFH) at the early stage. The suppressed proliferating ability and the decreased number and the inbalanced differentiation of mesenchymal stem cells (MSCs) caused by steroid is one of the cellular mechanisms underlie.In our previous study, we showed for the first time that lithium, well known as an anti-depression drug, promoted proliferation of human bone marrow derived MSCs in vitro. Mechanism study revealed that this effects was dependent on the glycogen synthase kinase 3 beta (GSK3β) mediated canonical Wnt pathway activation. Lithium abolished dexamethasone's effect on suppressing MSCs proliferate, but the mechanism is still unclear. We thus hypothesize that steroid suppresses β-catenin/Wnt signaling pathway of mesenchymal stem cells and lead to osteonecrosis of the femoral head. Lithium suppresses GSK3β, which causes the accumulation of β-catenin and activation of Wnt pathway in vivo. Therefore, lithium may be effective for the prevention/early treatment of steroid induced osteonecrosis of femoral head. We will use a series of assays to confirm our hypothesis and further explore the mechanism. Firstly, we plan to up-regulate and down-regulate the expression of GSK3β and β-catenin to further investigate of lithium's protective role in vitro. Secondly, we will use a series of transgenic mice to confirm our findings regarding lithium's protective role. Three strains of transgenic mice will be involved in this study. They are Prx1-Cre/ERT-GFP mice, β-catenin-flox mice and β-cateninfx(Ex3)/fx(Ex3) mice. We will specifically activated the β-catenin gene in mesenchymal stem cells in adult mice by generating tamoxifen inducible β-catenin conditional activation mice through breeding of β-catenin fx(Ex3)/fx(Ex3) mice with Prx1-cre/ERT-GFP transgenic mice. We will use these mice to test whether steroid caused osteonecrosis of femoral head by down-regulating β-catenin. Meanwhile, We will generating tamoxifen inducible β-catenin conditional knock out mice through breeding of β-catenin-flox mice with Prx1-cre/ERT-GFP transgenic mice. These mice will be used to test whether lithium exerts its protective effect through up-regulating β-catenin. Finally, we plan to confirm the above findings in a classical rabbit steroid induced ONFH model, aiming to develop a novel way to prevent or treat steroid induced ONFH at early stage.

英文关键词： Necrosis of femoral head;Bone marrow cells;osteonecrosis;stem cells;mesenchymal stem cells