Molecular analysis for the mechanisms of intervertebral disc degeneration using experimental animal model

我们建立了连续负重的小鼠尾巴退变模型。在该模型中，IVD在术后两周内降解。利用这个模型，我们测试了体重负荷是否会诱导Runx2的表达，从而可能导致软骨细胞肥大。事实上，压迫一天的IVD的重量负荷显著增加了Runx2的表达。因此，我们接下来检测了Runx2和其他椎间盘降解标记物在犬IVD中的表达，并用半定量RT-PCR方法检测了RUNX2、X型胶原和基质金属蛋白酶-13在幼年完整和退变IVD中的表达。用免疫组织化学方法检测Runx2和X型胶原蛋白在正常和退行性IVD中的定位。RUNX2转录本和蛋白的表达与X型胶原和基质金属蛋白酶-13结合，在犬退行性室间隔发育中的表达增强。在纤维化骨关节炎软骨中，Runx2与基质金属蛋白酶-13共定位于软骨细胞簇中，在骨关节炎软骨细胞中，基质金属蛋白酶-13的表达受Runx2的调控。X型胶原和MMP13是Runx2在生长板软骨细胞中的下游靶点。考虑到IVD细胞表达软骨特异性基质蛋白的数量不同，根据其解剖情况，IVD细胞、关节软骨细胞和生长板软骨细胞可能存在类似的分子调控。

We have developed continuous weight bearing disc degeneration model for mouse tail. In this model, degradation of IVD was observed within two weeks after operation. Using this model, we tested if weight loading, a most common cause of intervertebral disc (IVD) degeneration, induces Runx2 expression, which may cause chondrocyte hypertrophy. Indeed, weight loading of the IVD by compression for a day significantly increased Runx2 expression. Thus, we next examined the expression of Runx2 with other disc degradation markers in canine IVD, The expression of RUNX2, type-X collagen and MMP-13 mRNA in young intact and degenerated IVD were examined by semi-quantitative RT-PCR analysis. The localization of Runx2 and type-X collagen protein in control and degenerated IVD were examined by imunohistochemistly. The expression of RUNX2 transcript and protein, in combination with type-X collagen and MMP-13, was enhanced in degenerated IVDs of the dog. RUNX2 co-localizes with MMP-13 in clusters of chondrocytes in fibrillated OA cartilage, and MMP-13 expression is regulated by Runx2 in osteoarthritis chondrocytes. Type-X collagen and MMP13 are downstream targets of Runx2 in the growth plate chondrocytes. Considering IVD cells all express cartilage-specific matrix proteins with quantitative differences, depending on their anatomic situation, the comparable molecular regulation may exist in IVD cells, articular chondrocytes and growth plate chondrocytes.