葛根素诱导小鼠破骨细胞中Beclin1抑制Wnt/β-catenin信号通路的研究

As a kind of drugs for the treatment of osteoporosis, the curative effect of Puerarin has been confirmed. We found that Puerarin can inhibit cell proliferation, increase the expression of Beclin 1 and reduce the expression of Bcl-2 in mice osteoclast. Beclin 1 is an important link point between autophagyand apoptosis. As a haploinsufficient cell proliferation suppressor, the mechanisms by which Beclin 1 suppresses cell largely remain unclarified. Abnormal activation of Wnt/β-catenin is closely correlated with initiation, invasion and metastasis of cells. Epithelial-mesenchymal transition(EMT) is one of critical steps for cell invasion and metastasis. Generally, cells derived from epithelia acquire stronger capacity of invasion and proliferation via EMT. Using RNA microarray and SILAC/mass spectrum-based proteomics, we found that expression of Beclin 1 was closely correlated with transactivation of downstream targets of Wnt/β-catenin in mice osteoclast. In addition, we primarily demonstrated that Beclin 1 negatively regulated transactivity of β-catenin. Our data, for the first time, linked Beclin 1 and Wnt/β-catenin signal transduction together in osteoclast. The current project aims to investigate the mechanism and function of the regulation of Wnt/β-catenin signal transduction pathway by Beclin 1. Investigation on regulation of Beclin 1 and Wnt/β-catenin signal transduction helps us to find new target for osteoporosis therapy.

葛根素治疗骨质疏松(OP)的疗效已被证实，但机制尚未清楚。我们发现，葛根素在抑制小鼠破骨细胞生长的同时分别上调了Beclin 1和下调了Bcl-2蛋白的表达。Beclin 1作为一种自噬基因，通过调节自噬活性抑制细胞发生发展。Wnt/β-catenin信号转导通路的异常激活可以加速细胞生长。上皮间质转化(EMT)是上皮细胞来源的细胞获得迁移和侵袭能力的重要生物学过程。我们以RAW264.7细胞（可以诱导成破骨细胞）为例，首次发现：⑴ Beclin 1通过调控RAW264.7细胞Wnt/β-catenin信号通路，负向调控其下游靶基因的表达；(2)Beclin 1通过调控RAW264.7细胞EMT，抑制细胞侵袭转移。然而，以上结果的分子机制尚不清楚。本项目拟在前期工作基础上，利用分子生物学等手段，揭示葛根素治疗OP过程中，Beclin 1抑制破骨细胞生长的机制，为OP基因靶向治疗提供新思路。

研究背景：葛根素治疗骨质疏松(OP)的疗效已被证实,Beclin1作为一种自噬基因,通过调节自噬活性抑制细胞发生发展,Wnt/β-catenin信号转导通路的异常激活可以加速细胞生长。我们前期报道了葛根素在抑制小鼠破骨细胞生长的同时分别上调了Beclin1和下调了Bcl-2蛋白的表达,提示Beclin1可能通过Wnt/β-catenin信号转导通路抑制破骨细胞生长。本项目旨在揭示葛根素治疗OP过程中， Beclin1抑制破骨细胞生长的机制。.主要研究内容： 明确葛根素调控破骨细胞内Bcl-2以及BECN1表达的机制以及BECN1调控Wnt/β-catenin信号转导通路和破骨细胞EMT表性转化的分子机制及其意义。.重要结果：葛根素有雌激素样作用；鼠是骨质疏松动物模型最常选用的动物；葛根素提高绝经后骨质疏松症模型大鼠的护骨素，降低核因子-κB受体活化因子配体和血清碱性磷酸酶作用；葛根素抑制小鼠破骨细胞生长，并上调了Beclin1和下调了Bcl-2蛋白的表达。.科学意义： 本项目提示葛根素可能通过Beclin1抑制破骨细胞生长而发挥其治疗OP作用。