低密度脂蛋白受体相关蛋白(LRP5/6)对锶促进骨形成的作用机制

Osteoporosis and the related fragility fractures are a major public health concerns worldwide, especially with the aging population. It is more serious in China due to the large population. Extensive attention has been devoted to develop drugs for the treatment of osteoporosis. Anti-resorption drug and anabolic drug are the two major branches of treatment. In particular, Strontium (Sr) is the only drug that offers dual effects of both anti-bone resorption and bone formation..Strontium (Sr) has been verified clinically by reducing the risks of the vertebral and non-vertebral fracture in postmenopausal women. This effect has also been demonstrated in animal models of rat, mice and goat by our group. These findings motivated its application in early prevention of osteoporosis and incorporation in biomaterial. In past decade, we have developed strontium-containing hydroxyapatite and strontium-calcium compound for orthopedic applications. However, the exact working mechanism underlies this dual effects of Sr is still not clear..The dual effects of Sr are recognized by promoting osteogenesis and inhibiting the osteoclast activities. Our previous studies also show that Sr promotes the proliferation, differentiation of osteoblast, and osteogenesis of mesenchymal stem cell; the indirect down-regulation of osteoclast activities is regulated via osteoblasts generated osteoprotegrin. Currently, the exact signaling pathway working through for Sr to realize its effect still requires further exploration. .The similarity of Sr and Ca suggested the potential role of calcium-sensing receptor (CaSR) in promoting the anabolic effect of Sr in bone. However, unlike Ca and CaSR, Sr could not be homeostatically controlled, which indicates the possible involvement of other mechanism Recently, more studies have shown Sr activates Wnt signaling pathway in mesenchymal stem cells (MSCs) and osteoblasts. Low densitylipoprotein receptor-related protein 5/6 (LRP5/6) is a well-recognized co-receptors for Wnt signaling activation, which is crucial for osteoblast commitment and differentiation. In our pilot study, we found that Sr increases the phosphorylation of LRP6 and activate of Wnt signaling pathway in pre-osteoblasts. In addition, the genetic deletion of Lrp6 could block this effect in-vitro. .In conclusion, we hypothesize that the transmembrane receptor LRP5/6 is required for the anabolic effect of Sr in bone to promote osteoblast commitment, MSCs differentiation, and also enhance bone formation. Therefore, we aim to examine 1) whether Sr binds to LRP5 and/or LRP6 macromolecule and then stabilize β-catenin in MSCs in vitro; and 2) whether the increment of bone mass and the enhancement of osteogenesis of MSCs after Sr treatment is diminished in Lrp5 or Lrp6-KO mice in-vivo. This study will offer a new insight into the working mechanism of Sr in bone cells at ligand-receptor level, in particular the commitment and differentiation of MSCs towards osteogenesis.

骨质疏松是21世纪面临的最严重的老龄问题之一，在我国由于人口基数大而更为严峻。目前的治疗药物中，锶是唯一具有抗骨吸收和促进骨形成双重作用的口服药物，并已在临床上证明能够有效降低骨质疏松骨折的几率，但作用机制尚不明确。已有证据表明锶能够通过促进成骨细胞和干细胞的成骨分化，抑制破骨细胞活性而达到其治疗效果，但通过何种分子通路激活细胞相应的机能还有待进一步明确。越来越多的研究指出锶可能通过Wnt通路促进成骨，在我们的前期实验中也发现锶能够结合成骨细胞中Wnt的共同受体LRP5/6而激活其下游通路。基于此，本研究将进一步探讨Sr与成骨细胞和干细胞中的LRP5/6的结合情况及其对成骨分化的影响，并通过建立Lrp5/6条件基因敲除老鼠模型，明确LRP5/6在锶促进骨形成中的作用。本研究将会进一步从配体-受体水平上明确锶促进干细胞和成骨细胞的成骨分化作用机制，为其临床应用提供理论依据。

项目背景.骨质疏松是21 世纪面临的最严重的老龄问题之一，在我国由于人口基数大而更为严峻。目前的治疗药物中，锶是唯一具有抗骨吸收和促进骨形成双重作用的口服药物，并已在临床上证明能够有效降低骨质疏松骨折的几率，但作用机制尚不明确。.主要研究内容.1. 锶降低miR-9-5p、miR-675-5p和miR-138-5p表达水平(实验结果1部分)。2. miR-9-5p、miR-675-5p和miR-138-5p负调节成骨 (实验结果2-5部分)。因此，锶通过降低负调节因子（miR-9-5p、miR-675-5p和miR-138-5p）的表达而促进成骨。.重要结果.1. 锶盐在体内和体外改变miRNAs的表达。2. miR-9-5p、miR-675-5p和miR-138-5p抑制骨细胞增殖。3. miR-9-5p、miR-675-5p和miR-138-5p抑制骨细胞分化和黏附。4. miR-9-5p和miR-675-5p比miR-138-5p更易诱导细胞凋亡。5. miR-9-5p、miR-675-5p和miR-138-5p分别靶向于GSK3β、ATP8A2和 EIF4EBP1。6. LRP5在骨形成中起重要作用。.科学意义.本研究将会进一步从配体-受体水平上明确锶促进成骨细胞的成骨分化作用机制，为其临床应用提供理论依据。