Salubrinal通过调节内质网应激治疗骨质疏松的机制研究

As a metabolic bone disease, osteoporosis is a serious public health hazard. Thus, it is important to develop new drugs and to study new therapeutic mechanisms. Salubrinal is a selective inhibitor of a phosphatase specific to eukaryotic translation initiation factor 2α (eIF2α), and it protects cells from endoplasmic reticulum (ER) stress. Our previous studies showed that salubrinal inhibited dephosphorylation of eIF2α and induced activation of activating transcription factor 4 (ATF4), through which stimulated bone remodeling and accelerated bone wound healing. In addition, our studies further displayed that salubrinal reduced both the mRNA and protein levels of nuclear factor of activated T-cells, cytoplasmic 1 (NFATc1), and suppressed the development of osteoclast. However,the role of ER stress in the development of osteoporosis remains unknown. The underlying mechanisms of salubrinal regulating bone remodeling is also largely unexplored. The overall objective of this project aims to better understand the molecular mechanisms that salubrinal promotes bone remodeling for osteoporosis treatment. This project will adopt osteoporotic animal models and the techniques of histology, cytology, and molecular biology to test the hypothesis that ER stress plays an importment role in the development of osteoporosis, and salubrinal stimulates the differentiation of osteoblast and inhibits the development of osteoclast by elevating eIF2α phosphorylation which regulates ATF4，NFATc1, and RAC1 GTPase expression and activity. The effects of salubrinal on the ER stress associated signaling pathway such as NFκB and p38 MAPK, and its beneficial affects including the promotion of bone formation and inhibition of bone resorption will also be investigated. This study will provide new insights into physiological and pathological importance of the roles of ER stress signaling in the initiation and development of osteoporosis, and provide a novel target for treatment of osteoporosis.

骨质疏松严重危害公共健康，研究新药物及治疗新机制意义重大。Salubrinal是eIF2α去磷酸化的选择性抑制剂，保护细胞免于内质网应激。我们前期工作表明salubrinal通过抑制eIF2α去磷酸化、激活ATF4、促进骨重建而加速骨损伤愈合；salubrinal还降低NFATc1表达，抑制破骨细胞发育。然而，内质网应激在骨质疏松中的作用尚不明确，salubrinal调节骨重建的信号通路尚需阐明。为此，本项目将采用骨质疏松动物模型，组织学、细胞学和分子生物学手段验证以下假设：内质网应激在骨质疏松发生发展中起重要作用，通过上调应激中的eIF2α磷酸化水平，salubrinal不仅改变ATF4、NFATc1和RAC1 GTPase的表达，而且还调节NFκB和p38 MAPK信号通路，促进成骨细胞分化和抑制破骨细胞发育，通过促骨生成和抗骨吸收治疗骨质疏松。本项目的研究将为骨质疏松治疗提供新靶点。

骨质疏松是严重影响生存质量的全身性疾病，新的药物治疗方法及机制有待研究。Salubrinal是一种eIF2α去磷酸化的选择性抑制剂，保护细胞免于内质网应激，但是内质网应激在骨质疏松中的作用未明。本课题通过不同时间的鼠尾悬吊小鼠来探讨内质网应激在骨质疏松发生发展中的重要作用；体内皮下Salubrinal给药后使用骨密度测量、micro-CT和组织学染色等手段方法验证Salubrinal对鼠尾悬吊和卵巢去势导致的骨质疏松小鼠中均有显著疗效；利用骨髓细胞诱导的成骨细胞分化和MC3T3-E1成骨细胞系验证Salubrinal通过eIF2α和Atg7调节成骨细胞的内质网应激-自噬轴，从而促进成骨细胞骨形成。使用骨髓细胞诱导的破骨细胞形成和RAW264.7破骨细胞前体细胞系，阐明Salubrinal通过调节eIF2α、Rac1和NFATc1来改变破骨细胞的分化来减弱骨丢失。使用透射电镜、免疫荧光、细胞学和分子生物学等手段，阐述Salubrinal通过上调内质网应激中的eIF2α磷酸化水平，改变多靶点信号通路ATF4、NFATc1和Rac1 GTPase的表达和活性，调控内质网应激和自噬，从而促骨生成和抗骨吸收治疗骨质疏松。综上所述，Salubrinal是一种通过内质网应激和自噬，既可促进成骨细胞分化而增加骨生成，又可抑制破骨细胞发育而减少骨吸收，具有双重疗效的潜在骨质疏松治疗药物。