早期激素性股骨头缺血性坏死模型的建立及半胱天冬酶3活性测定

Establishment of an early steroid-induced femoral head necrosis model and determination of Caspase-3 activity

背景:有研究表明骨细胞凋亡与激素性股骨头缺血性坏死的发病机制密切相关,而活化的半胱天冬酶3可作用于其他半胱天冬酶家族成员,使蛋白酶级联反应放大,最终导致细胞凋亡。但半胱天冬酶3参与的骨细胞凋亡与早期激素性股骨头缺血性坏死的相关研究则罕见报道。目的:采用大剂量激素联合强迫兔直立负重的方法建立早期激素性股骨头缺血性坏死模型并测定骨细胞半胱天冬酶3活性。方法:将4月龄新西兰大白兔随机分成模型组和对照组。模型组每周3次臀肌注射地塞米松,每日强迫兔直立1h,对照组注射等量生理盐水。分别于3,6,9和12周处死动物,取其双侧股骨头,光镜及电镜下观察组织病理学改变,应用对-硝基酰苯胺比色法测定骨细胞半胱天冬酶3活性。结果与结论:给药3周后,模型组光镜下可见脂肪细胞肥大和增多,骨髓间充质干细胞及造血干细胞数量相对减少;给药6周后,模型组空骨陷凹率均大于对照组(P<0.05);给药9周后,模型组出现骨小梁变细、断裂,电镜下见模型组部分骨细胞体积缩小,染色质浓集,核固缩和碎裂等凋亡特征。给药6周后,模型组骨细胞半胱天冬酶3活性高于对照组(P<0.05)。实验以此推论采用大剂量激素联合强迫兔直立负重可以诱导建立早期激素性股骨头缺血性坏死模型,半胱天冬酶3参与的骨细胞凋亡可能与早期激素性股骨头缺血性坏死发病过程有关。

BACKGROUND： Previous studies showed that apoptosis in osteocyte had close relationship to the pathogenesis of steroid-induced femoral head necrosis, while the activation of Caspase-3 can act on the other members of Caspase family, lead to protease cascade amplification, and eventually result in apoptosis. However, a report about Caspase-3 participates in apoptosis in osteocyte and early steroid-induced femoral head necrosis remains rare. OBJECTIVE： To establish an early steroid-induced femoral head necrosis model and determine Caspase-3 activity in osteocyte. METHODS： The rabbits were randomly divided into the model and control groups. Rabbits in the model group were injected dexamethasone 3 times a week and forcing the rabbits held upright position 1 hour everyday, while rabbits in the control group were injected the same amount of saline. Animals were sacrificed at 3, 6, 9 and 12 weeks after operation, and the optical microscope and transmission electron microscopy were used to observe the histopathological changes, and p-nitroanilide colorimetry was performed to measure the Caspase-3 activity in osteocyte. RESULTS AND CONCLUSION： Histopathological changes could be observed under a light microscope at 3 weeks after administration, including increasing of empty osteocyte lacuna, adipocyte hypertrophy and relatively decreasing of mesenchymal stem cells. The percentage of empty osteocyte lacunae in the model group was significantly higher than in the control group at 6 weeks after administration （P 0.05）. Trabecular bone becomes thinner even collapse in model group at 9 weeks after administration. With the electron microscope, morphological changes in bone cells were observed. At 6 weeks after administration, osteocyte Caspase-3 activity in the model group was higher than the control group （P 0.05）. Accordingly, the use of high dose injection of dexamethasone and forcing the rabbits holding upright position can establish an early steroid-induced femoral head necrosis model. The apoptosis in osteocyte depending Caspase-3 may have some relationship to the pathogenesis.