The Role of MicroRNA in Osteoarthritis

摘要 近年来，骨关节炎(OA)的研究取得了重大进展，包括产生了 小鼠模型，包括遗传和手术诱导的表现为类骨关节炎的骨性关节炎小鼠模型。 这些小鼠模型的一个共同特征是上调了Runx2的表达。因为Runx2是一个 直接调节编码基质降解酶基因的表达的关键转录因子， 如Mmp9、Mmp13和Adamts5，这种上调导致这些基因的表达增加。 关键问题已经提出来了。1)Runx2是否是调节关节组织中骨性关节炎发生的中心分子？ 2)操控Runx2的表达能否用于治疗骨性关节炎？MiRNAs是内源性的非 编码RNA，在调节RNA稳定性和蛋白质表达方面发挥重要作用。以前的研究 我们实验室的研究表明，miR-204/-211这两个同源miRNAs结合Runx2并调节Runx2 在间充质干细胞(MSCs)中表达。要确定这些miRNAs在Runx2中的功能 在OA的发育过程中，我们最近创造了miR-204和miR-211等位基因，并进一步 通过培育miR-204/FLOX和miR-204/FLOX获得了miR-204/-211双KO小鼠(dmiRPrx1和dmiRAgc1er)。 211FLOX/FLOX小鼠，携带PRX1-CRE(靶向肢体MSCs)或Agc1-Creer(靶向成人关节软骨细胞) 转基因小鼠删除这些miRNAs。在dmiRPrx1和dmiRPrx1中观察到严重的OA样表型 MiR-204/-211缺失影响DmiRAgc1ER KO小鼠和整个膝关节组织，包括严重丢失 关节软骨、软骨下硬化症、软骨赘/骨赘形成和滑膜增生症。 我们的发现表明miR-204和miR-211是维持关节组织动态平衡和 可作为治疗骨性关节炎的关键靶点。这个项目的基本假设是miR- 204和miR-211是Runx2表达的关键调节因子，在维持关节组织中起着关键作用 动态平衡。已经提出了两个具体的目标来检验这一假设。在目标1中，我们将充分 研究miR-204/-211缺失对dmiRAgc1er KO小鼠和 确定软骨细胞中Runx2的缺失是否会逆转在dmiRAgc1er KO小鼠中观察到的OA表型。在……里面 目的2，我们将研究miR-204/-211在骨性关节炎标本中的表达变化，并确定在体内 应用miR-204将阻止手术诱导的骨性关节炎的发展或减缓骨性关节炎的进展 老鼠模型。这些拟议的研究将为OA的发展和机制提供新的见解 将有助于开发治疗骨性关节炎的新策略。

Abstract Significant progress has been made in recent years in osteoarthritis (OA) research, including the generation of mouse models, including genetic and surgically-induced OA mouse models that show an OA-like phenotype. One feature that is common to these mouse models of OA is the up-regulation of Runx2. Because Runx2 is a key transcription factor directly regulating the expression of genes encoding for matrix degradation enzymes, such as Mmp9, Mmp13 and Adamts5, this up-regulation leads to the increase in expression of these genes. The key questions have been raised. 1) Is Runx2 a central molecule mediating OA development in joint tissue? 2) Could manipulation of Runx2 expression be used to treat OA disease? miRNAs are endogenous non- coding RNAs and play important roles in regulation of RNA stability and protein expression. Previous studies by our lab have shown that miR-204/-211, two homologous miRNAs, bind Runx2 and regulate Runx2 expression in mesenchymal stem cells (MSCs). To determine the function of these miRNAs in Runx2 regulation and in OA development, we have recently created miR-204 and miR-211 floxed alleles and further generated miR-204/-211 double KO mice (dmiRPrx1 and dmiRAgc1ER) by breeding miR-204flox/flox and miR- 211flox/flox mice with Prx1-Cre (targeting limb MSCs) or Agc1-CreER (targeting adult articular chondrocytes) transgenic mice to delete these miRNAs. A severe OA-like phenotype was observed in dmiRPrx1 and dmiRAgc1ER KO mice and entire knee joint tissues were affected by miR-204/-211 deletion, including severe loss of articular cartilage, subchondral sclerosis, chondrophyte/osteophyte formation, and synovial hyperplasia. Our findings suggest that miR-204 and miR-211 are key regulators in maintaining joint tissue homeostasis and could be served as critical targets for OA treatment. The underlying hypothesis of this project is that miR- 204 and miR-211 are key regulators of Runx2 expression and play critical roles in maintaining joint tissue homeostasis. Two specific aims have been proposed to test this hypothesis. In Aim 1, we will fully characterize the longitudinal effects of miR-204/-211 deletion on OA development in dmiRAgc1ER KO mice and determine if deletion of Runx2 in chondrocytes will reverse OA phenotype observed in dmiRAgc1ER KO mice. In Aim 2, we will investigate changes in miR-204/-211 expression in OA samples and determine if in vivo administration of miR-204 will prevent OA development or decelerate OA progression in surgically-induced OA mouse model. These proposed studies will provide novel insights into mechanisms of OA development and will help the development of new strategies for OA treatment.