老鹳草素干预BMSCs成骨分化及其对Wnt/β-catenin信号通路调控作用研究

Development of new action mechanism of drug will be of great importance in preventing and curing osteoprosis(OP). The weakened osteogenetic differentiation function of bone marrow stroma stem cells (BMSCs) is possibly one of the major reasons of OP. Recent researches proved that Wnt/β-catenin signaling pathway plays an important regulating role in the process of osteoblastic proliferation. Our previous study showed that gerannin, isolated from Phyllanthus amarus L. native to Yunnan Province, had anti-osteoporotic effects. Our preliminary investigation suggested that geraniin can promote the proliferation of osteoblasts and the differentiation of rat BMSCs, and up-regulate the protein and its mRNA expression of Wnt3a in Wnt/β-catenin signaling pathway. Based on our these results, the model of OVX-induced osteoprosis is duplicated in this present subject, in order to intensively study the effects of geraniin on osteogenic differentiation of BMSCs and the expression and transposition of the key target β-catenin of Wnt/β-catenin signaling pathway and the intervening links of its upstream-downstream signaling molecules. The relationship is also analyzed between regulation of Wnt/β-catenin signaling pathway, intervening osteogenic differentiation of BMSCs and curing osteoprosis. This project will reveal the molecular mechanism of geraniin as a therapeutic agent for osteoprosis and can provide valuable theoretical and experimental evidences for geraniin as a self-innovated and plant-originated anti-osteoporotic drug.

开展防治骨质疏松（OP）药物的新作用机制研究具有重要意义。骨髓基质干细胞(BMSC)成骨分化减弱是OP的主要原因之一，研究表明经典Wnt/β-catenin信号对BMSC的成骨分化有重要调节作用。课题组研究发现，老鹳草素具有抗OP作用，促进成骨细胞增殖和正常大鼠BMSCs的成骨分化，增加Wnt3a蛋白及mRNA的表达。在此基础上，本课题拟复制大鼠OP模型，研究老鹳草素对源于OP大鼠BMSCs增殖、成骨分化能力的影响和与成骨密切相关的Wnt通路中关键分子β-catenin表达、转位及其对上下游信号分子的干预环节，探索其成骨分化作用与Wnt/β-catenin信号通路中上述关键靶标的关系，揭示老鹳草素诱导成骨分化和抗OP的新作用机制，为将老鹳草素研究成为自主创新和云南特色药用植物来源的OP防治药物提供进一步的理论和实验依据。

骨质疏松（osteoporosis, OP）及其并发症是老年患者致死致残的重要原因。随着人口老龄化的加剧，已成为全球范围内的重大公共健康问题。老鹳草素（Geraniin）是云南特色药用植物大戟科叶下珠（Phyllanthus amarus L.）的主要活性成分，动物实验发现体内具有抗去卵巢（Ovariectomy, OVX）或糖皮质激素诱发OP的作用，但其潜在的细胞分子生物学机制尚不明确。在前期研究基础上，本项目复制大鼠骨质疏松症（OP）模型，研究老鹳草素对OP大鼠骨髓间充质干细胞（BMSC）增殖和成骨分化的影响，探讨其量效关系，明确老鹳草素抗OP作用与Wnt/β-catenin信号通路上、下游增殖、分化关键因子的关系，进一步揭示老鹳草素促进BMSC增殖和成骨分化、从而治疗OP的潜在机制。研究结果提示OVX手术可抑制Wnt/β-catenin信号通路，导致OP大鼠BMSC/OB的增殖和成骨分化能力较正常大鼠显著下降。老鹳草素可显著增加OP大鼠BMSC的早期和晚期成骨分化，并促进BMSC/OB增殖。其分子机制为老鹳草素激活OP大鼠BMSC中Wnt/β-catenin信号通路，使下游增殖相关因子C-myc、Cyclin D1和分化相关因子Runx2、Osterix表达上调。老鹳草素激活Wnt/β-catenin信号通路及其下游骨形成相关因子的作用具有浓度依赖性，因而其促进BMSC/OB增殖和成骨分化的作用也具有浓度依赖性。老鹳草素促进β-catenin核转位和核内因子表达的作用具有饱和性。本研究初步探索了老鹳草素促进BMSC成骨分化的量效关系，为其成药性研究提供了参考，揭示了老鹳草素抗OP作用的分子靶点，并初步探讨了其基因调控网络及信号通路交互作用。本研究为研发云南特色药用植物来源的新型抗OP药物提供理论和实验依据。