Biomolecular Analysis on Decorin associated with Ossification of the Longitudinal Ligament

在OPLL患者和脊髓型颈椎病患者的脊柱韧带中，我们检测了转化生长因子-β和核心蛋白聚糖的表达，但没有显著差异。然后，我们对OPLL患者的颈部皮肤进行了同样的对比研究，此前发现OPLL患者颈部皮肤硬度异常。结果发现，OPLL患者表皮中转化生长因子-β的免疫染色较对照组稍强。OPLL患者表皮核心蛋白聚糖染色明显增强。我们进一步通过原位杂交检测了核心蛋白聚糖的mRNA。OPLL患者Decorin基因的表达也明显增强。考虑到OPLL患者皮肤的硬度异常，这些结果强烈提示OPLL患者皮肤细胞外基质的异常。为了研究核心蛋白聚糖在骨形成中的作用，我们还用免疫组织化学方法检测了大鼠骨折模型中转化生长因子-β和核心蛋白聚糖的表达。在软骨细胞骨化部位，转化生长因子β和核心蛋白聚糖的定位相反，提示核心蛋白聚糖在软骨细胞成骨过程中对转化生长因子β起着某种形式的调节作用。Decorin在韧带骨化中的作用尚不清楚。然而，这些结果与先前报道的血清纤维连接蛋白升高是一致的，反映了OPLL患者全身骨化的易感性，并证实了细胞外基质异常的存在。细胞外基质与生长因子一起在细胞的增殖和分化过程中发挥着重要的局部作用。因此，推测细胞外基质的异常在异位骨化的发生和发展中起着额外的作用。

In spinal ligaments collected from patients with OPLL and those with cervical spondylotic myelopathy, we examined the expressions of TGF-beta and decorin, but no significant differences were observed. Then, we made the same comparative study in the nuchal skin of OPLL patients in which hardness abnormality had been pointed out since befor. As a result, it was found that the immunostaining for TGF-beta in epidermis of OPLL patinets tended to show somewhat stronger than those of control patients. The staining of decorin in the epidermis of OPLL patients increased significantly. We further examined mRNA of decorin by in situ hybridization. Then, the expression of decorin mRNA was also enhanced in OPLL patients. Considering the hardness abnormality in the skin of OPLL patients, these results strongly suggest an abnormality in extracellular matrix of the skin of OPLL patients. To study the roles of decorin in bone formation, we also examined expression of TGF-beta and decorin immunohistochemically in rat fracture models. At the site of enchondral ossification, localization of TGF-beta and decorin were opposite to each other, suggesting that decorin works as a regulatory factor in some form against TGF-beta in the process of enchondral ossification. The action of decorin on the ossification of ligament has been not clarified yet. However, these results are consistent with an increase in serum fibronectin as previously reported, reflect OPLL patients' predisposition to systemic ossification and substantiate the presence of an abnormality in extracellular matrix. Extracellular matrix plays an important role locally, together with the growth factor, in proliferation and differentiation of the cells. Therefore, it is presumed that the abnormality in extracellular matrix has playd some additional role in the initiation or progression of heterotopic ossification.