中药调节差异microRNAs表达调控细胞凋亡防治激素性股骨头缺血坏死机理研究

Steroid-induced osteonecrosis of femoral head（SONFH）is a common orthopaedic disease, its exact pathophysiological mechanism is unclear. In recent years, studies have found that apoptosis played an important role in pathogenesis of SONFH, highly conserved miRNA had a role in the regulation of cell proliferation and apoptosis. In this study, miRNAs microarray will be used to detect miRNAs expression profiles of SONFH femoral head and serum, based on which differential miRNAs which have the same trend in the femoral head and the serum will be screened out and validated by Real-time PCR. It is expected to find miRNA related with SONFH as molecular markers for early diagnosis; and analyze signaling pathway of target genes regulated by these differential miRNAs, to find significantly pathway regulated by miRNAs and genes corresponding to significantly pathway, in order to explore the mechanism of apoptosis regulated by miRNA; Finally rats models with steroid-induced osteonecrosis are made and apoptosis experiment of osteoblast in vitro induced by steroid are done to observe the impact of Chinese medicine(Yuan Shi Sheng Mai Cheng Gu Pian) in target miRNA, to explore the impact of Chinese medicine(Yuan Shi Sheng Mai Cheng Gu Pian) in predicted target genes and related signaling pathways during the course of treatment of SONFH. All we do are expected to clear the mechanism that prevention and treatment of SONFH with Chinese medicine regulating miRNA exprssion.

激素性股骨头缺血性坏死（SONFH）是骨科常见病、多发病，其确切病理生理学机制并未明确。近来发现细胞凋亡在SONFH发病中起着重要作用，高度保守的miRNA具有调控细胞增殖和凋亡的作用。本研究应用miRNAs芯片检测SONFH患者股骨头及血清miRNAs表达谱，基于miRNAs表达谱筛选出股骨头与血清中具有相同变化趋势的差异miRNAs，并以Real-time PCR进行验证。期望发现SONFH相关miRNA，作为早期诊断的分子标志物；对这些miRNAs可能调控的靶基因进行信号通路分析，发现其调控的显著性通路及通路对应的基因，探索miRNA调节细胞凋亡的作用机理；最后以激素诱导大鼠股骨头缺血坏死模型及激素诱导体外培养成骨细胞凋亡实验，观察中药袁氏生脉成骨片对目标miRNA的影响，探索股骨头缺血坏死治疗过程中对预测的靶基因及相关信号通路的影响，期望明确中药防治SONFH miRNA调控机理。

本研究应用 miRNAs 芯片检测 SONFH 患者股骨头及血清 miRNAs表达谱，基于 miRNAs 表达谱筛选出股骨头与血清中具有相同变化趋势的差异 miRNAs，并以Real-time PCR 进行验证。期望发现 SONFH 相关 miRNA，作为早期诊断的分子标志物；对这些 miRNAs 可能调控的靶基因进行信号通路分析，发现其调控的显著性通路及通路对应的基因，探索 miRNA 调节细胞凋亡的作用机理；近来发现细胞凋亡在 SONFH 发病中起着重要作用，高度保守的miRNA 具有调控细胞增殖和凋亡的作用。本研究在 SONFH 患者血浆中发现差异表达的 hsa-miR-887、hsa-miR-576-5p，靶基因进行生物信息分析，结果发现hsa-miR-887 对应交集靶基因为 GSK3A和CASK，主要调控解剖结构形态; hsa-miR-96-5p 检出交集靶基因 114 个、hsa-miR-576-5p 检出 59 个，主要调控间充质细胞增殖、细胞代谢过程、细胞内信号转导等生物学过程，可作为早期诊断的分子标志物；通过含药物血清细胞实验证明袁氏生脉成骨片抑制此类miRNA的表达起治疗作用。我们的实验发现miR-672-5p的表达最为明显，并预测出Angptl4、Ccdc51、RGD1306991、Ssbp3 作为miR-672-5p的靶基因。通过对临床股骨头坏死标本的芯片测定，我们发现hsa-miR-195-5p在塌陷区有下调表达。本研究通过基因芯片技术，多层面研究中医药预防激素型股骨头坏死的理论基础和结果，取得了初步进展。