Although the mineralization in tendon tissue has been reported in a series of aging and disease models, the underlying mechanisms remain unknown. This study aimed to describe the appearance of heterotopic ossification in rat Achilles tendon and further verify whether this tissue metaplasia is related to the enhanced osteogenic differentiation of tendon stem/progenitor cells (TSPCs) owing to the higher expression of bone morphogenetic proteins (BMP-2/4/7) with aging. The male SD rats, aged 4, 8, and 20 months (M), were used. The analyses of ossification and BMP expression in tendon were tested by radiological view (X-ray and CT), histological staining [hematoxylin and eosin (HE), Alcian blue, and Alizarin red], immunohistochemistry, and Western blot. The osteogenic differentiation potential and BMP expression of TSPCs were examined by Alizarin red S staining and real-time PCR. TSPCs were treated with BMP-2 or noggin, and the osteogenic differentiation potential was also examined. X-ray and CT showed the appearance of heterotopic ossification in tendon, and the volume and density of ossification was increased with aging. Histological staining showed the appearance of calcified region surrounded by chondrocyte-like cells and the increased osteogenesis-related gene and BMP expression in ossified tendon with aging. Moreover, the osteogenic differentiation potential and BMP expression in TSPCs isolated from ossified tendon were increased with aging. Additionally, BMP-2 increased the calcium nodule formation and osteogenesis-related gene expression in TSPCs. The addition of noggin inhibited BMP-induced enhancement of osteogenic differentiation. Thus, these findings suggested that the enhanced osteogenic differentiation of TSPCs contributes to the increased heterotopic ossification in aged tendon, which might be induced by the higher expression of BMPs with aging.

尽管已经在一系列衰老和疾病模型中报告了肌腱组织中的矿化现象，但其潜在机制仍然未知。这项研究旨在描述大鼠跟腱异位骨化的外观，并进一步验证这种组织化生是否与由于骨形态发生蛋白（BMP-2）的较高表达而增强的腱干/祖细胞（TSPC）的成骨分化有关/ 4/7）。使用雄性SD大鼠，分别为4、8和20个月（M）。通过放射学检查（X射线和CT），组织学染色（苏木精和曙红（HE），阿尔辛蓝和茜素红），免疫组化和Western blot检测肌腱中骨化和BMP表达的分析。通过茜素红S染色和实时荧光定量PCR检测TSPCs的成骨分化潜能和BMP表达。用BMP-2或头蛋白处理TSPC，并检查其成骨分化潜能。X射线和CT显示肌腱中出现异位骨化，并且骨化的体积和密度随着年龄的增长而增加。组织学染色显示，随着年龄的增长，骨化肌腱中钙化区域的外观被软骨细胞样细胞包围，成骨相关基因和BMP表达增加。而且，随着年龄的增长，从骨化肌腱分离的TSPC中成骨分化潜能和BMP表达增加。此外，BMP-2增加了TSPC中钙结节的形成和成骨相关基因的表达。头蛋白的添加抑制了BMP诱导的成骨分化增强。因此，这些发现表明，TSPCs的成骨分化增强促进了老年肌腱异位骨化的增加，这可能是由于年龄增长的BMPs的高表达所致。