miR-664-3p靶向调节Smad4和Osterix促进骨质疏松发生的研究

Osteoporosis (OP) is becoming a serious health care issue in the countries worldwide including China now. MiR-664-3p is one of the differentially expressed miRNAs during the osteoblast differentiation screened by our group using high-throughput sequencing, whose expression was down-regulated with the maximal fold. We have demonstrated that both Smad4 and Osterix were target genes of miR-664-3p. Smad4 is a common Smad of BMP2 signal pathway. Osterix is a key regulator of osteoblast differentiation. Moreover, we found that miR-664-3p was down-regulated continuously in the process of osteoblast differentiation. However, the levels of miR-664-3p were markedly increased in the bone tissues of ovariectomy-induced osteoporotic mice. Furthermore, we found that the levels of miR-664-3p in the serum of osteoporotic patients were much higher than that of the matched control (13 patient vs 10 control). Based on our findings and information from the literatures, we postulate that miR-664-3p plays an inhibitory role in osteoblast differentiation and its overexpression promotes the occurrence of OP. We will test this hypothesis using osteoblast differentiation model, osteoblast-specific transgenic mice construction as well as clinical investigations. In addition, the underlying mechanisms by which miR-664-3p inhibits osteoblast differentiation and promotes OP genesis will be explored by gene overexpression and knockdown assays. Results of this project will not only shed new light on the molecular mechanisms underlying osteoblast differentiation and OP occurrence, but also provide a novel target for the prevention and treatment of OP. At the same time, results of this project may provide a novel biomarker for the diagnosis/auxiliary diagnosis of OP.

骨质疏松目前在我国乃至全球都是一个值得关注的严重的健康问题。miR-664-3p是本课题组筛选出的成骨细胞分化过程中下调倍数最大的miRNA。我们已经证明BMP信号通路的信号转导分子Smad4和成骨细胞分化关键调节因子Osterix是miR-664-3p的靶基因。我们还发现：成骨细胞分化过程中miR-664-3p的表达持续下调；骨质疏松小鼠骨组织以及骨质疏松病人血清中的miR-664-3p的水平显著高于对照组。由此我们推测：miR-664-3p具有抑制成骨细胞分化的功能，其表达上调促进骨质疏松的发生。本项目拟通过细胞实验、成骨细胞特异性转基因小鼠构建和临床标本检测对此假说进行验证；并对miR-664-3p作用的分子机制进行深入探讨。本项目的研究结果不仅可以有助于成骨细胞分化和骨质疏松发病的分子机制的阐明；而且可以为骨质疏松的预防和治疗提供新的靶点，为其诊断/辅助诊断提供新的生物标志物。

骨质疏松症（osteoporosis, OP）是一种以骨量减少、骨微结构破坏、骨脆性增加以及易发生骨折为特征的常见的代谢性骨病。激活成骨细胞功能从而促进骨形成是治疗骨质疏松症的策略之一。microRNAs（miRNAs）是在基因转录后水平发挥调控作用的小分子非编码RNA，几乎参与了所有的生理病理过程。研究发现在骨质疏松症中一些miRNAs确实发生了异常表达，但它们是否控制成骨分化和骨形成以及是否可以成为骨质疏松症的治疗靶点，仍需要深入研究。.在本研究中，我们鉴定出miR-664-3p是一个新的成骨细胞分化和骨形成中的关键抑制子。我们的结果表明，miR-664-3p表达在成骨细胞分化过程中显著下调，且过表达miR-664-3p显著下调成骨细胞活性和基质矿化。生物信息学分析表明，Smad4和Osterix是miR-664-3p的潜在靶基因，并且得到荧光素酶报告基因实验的验证。我们构建了成骨细胞特异性的miR-664-3p转基因小鼠，发现这些小鼠的骨形成和骨量明显减少；抑制miR-664-3p表达可以显著减轻miR-664-3p转基因小鼠因去卵巢而导致的骨质疏松。此外，我们还发现与正常对照相比，骨质疏松病人血清中的miR-664-3p水平显著升高。.总之，该研究首次证明miR-664-3p至少部分通过下调Smad4和Osterix的表达而抑制成骨发生。miR-664-3p有可能成为骨质疏松症诊断/辅助诊断的新型分子标志物以及临床上骨质疏松治疗和预后的新靶点。