Cyclic Tensile Stress to Rat Thoracolumbar Ligamentum Flavum Inducing the Ossification of Ligamentum Flavum

Study Design. Western blot, reverse transcription-polymerase chain reaction (RT-PCR), radiological, and histological analyses of the rat ossification of ligamentum flavum (OLF) induced by cyclic tensile stress. Objective. The aim of this study was to induce the OLF using cyclic tensile stress to rat thoracolumbar ligamentum flavum, and to investigate the possible molecular mechanism of tension-induced OLF. Summary of Background Data. Tensile stress has been considered as an important factor leading to the OLF. So far, however, no OLF induced by tension has been reported. Methods. Forty rats were randomly divided into five equal groups. For control groups, the blank and anesthesia groups were not subjected to tension. For experimental groups, the 4-, 8-, and 12-week groups were subjected to cyclic tensile stress of ligamentum flavum after abdominal anesthesia for 4weeks, 8 weeks, and 12 weeks, respectively, using an original stress apparatus for rats. The radiological and morphological changes of rat spine, as well as the protein and mRNA expressions of CD44, bone morphogenetic protein-2 (BMP-2), integrin beta 3, collagen protein type I (COL1), osteopontin (OPN), runt-related transcription factor 2 (RUNX-2), and vascular endothelial growth factor (VEGF), were concerned. Results. The micro-CT showed OLF in the 4-, 8-, and 12-week group. The axial maximum occupied area of ossifications was 1.42 mm(2), 3.35 mm(2), and 7.28 mm(2), respectively. In histopathology, chondrocytes proliferated in the experimental model; woven bone arose in the 8- and 12-week groups, and was more noticeable in the 12-week group. According to western blot and RT-PCR, the expressions of seven osteogenesis-related molecules were all increased in three experimental groups. Conclusion. Cyclic tensile stress to the ligamentum flavum in rats can induce the OLF, and the longer the duration, the more visible the osteogenesis. The upregulation and synergism of osteogenesis-related molecules may contribute to the OLF induced by tensile stress.