激素性股骨头坏死早期细胞凋亡相关基因表达及阿仑磷酸钠的干预

背景：近年研究发现激素性股骨头坏死病例中凋亡的骨细胞增多，因此认为激素性股骨头坏死是骨细胞凋亡所致，而并非缺血性坏死。目的：观察大鼠激素性股骨头坏死早期bcl-2、半胱氨酸天冬氨酸蛋白酶3(cysteinyl aspartate specific protease 3，caspase-3)的表达，以及阿仑磷酸钠的干预效果。设计、时间及地点：随机对照动物实验，于2006-03/2007-02在吉林大学基础医学院病理实验室完成。材料：健康Wister大鼠100只，随机分为正常对照组、激素模型组、藻酸双酯钠组、阿仑磷酸钠组、联合用药组，20只/组。藻酸双酯钠片为吉林东丰制药厂产品，阿仑磷酸钠为杭州默沙东制药有限公司产品。方法：除正常对照组外，其余4组大鼠均给予醋酸泼尼松龙臀肌注射建立激素性股骨头坏死模型。藻酸双酯钠组、阿仑磷酸钠组分别灌服对应药物，联合用药组灌服藻酸双酯钠+阿仑磷酸钠。各组肌注用药1次/周，灌服药1次/d，实验持续6周。所有大鼠均强迫间断站立，4 h/d。主要观察指标：光镜观察股骨头病理变化。S-P免疫组织化学染色法检测股骨头组织bcl-2、caspase-3的表达。结果：给药第4周与正常对照组比较，激素模型组股骨头坏死现象明显，骨小梁变细甚至断裂，空骨陷窝数量明显增多(t=2.629，P < 0.01)；藻酸双酯钠组骨小梁和空骨陷窝数量介于前2组之间(t=1.713，P < 0.01)；阿仑磷酸钠组、联合用药组骨小梁和空骨陷窝数量接近正常对照组(t=1.462，t=1.657，P > 0.05)。与激素模型组比较，藻酸双酯钠组、阿仑磷酸钠组、联合用药组股骨头组织bcl-2阳性细胞率均明显升高(t=2.146，P < 0.05；t=2.815，P < 0.01；t=2.947，P < 0.01)，caspase-3阳性细胞率均明显降低(t=2.97，P < 0.01；t=2.462，P < 0.01；t=1.854，P < 0.05)，其中阿仑磷酸钠组及联合用药组升高尤为明显(t=2.503，t=1.959，P < 0.05)。结论：大剂量激素可能通过抑制bcl-2表达和促进caspase-3表达引起股骨头坏死。阿仑磷酸钠可能通过与其相反的途径来防止骨细胞凋亡，从而预防早期激素性股骨头坏死。

Changes in apoptosis related gene expression following alendronate intervention in the early srage of steroid-induced necrosis of the femoral head

BACKGROUND: The research demonstrates that steroid-induced necrosis of femoral head may caused by osteocytes apoptosis rather than ischemic necrosis.OBJECTIVE: To investigate the expression of bcl-2 and cysteinyl aspartate specific protease 3 (caspase-3) and intervention effect of alendronate sodium in femoral head of steroid-induced necrosis in rats.DESIGN, TIME AND SETTING: A randomized controlled animal study was performed in the Pathological Laboratory, Basic Medical School of Jilin University between March 2006 and February2007.MATERIALS: 100 healthy Wister rats were randomly divided into the normal control, glucocorticoid model, propylene glycol alginate sodium sulfate (PSS), alendronate sodium and PSS combined with alendronate groups, with 20 rats in each. The alginic sodium diester tablets were provided by the Dongfeng Pharmaceutical Factory, and alendronate sodium was supplied by the Hangzhou MSD Pharmaceutical Co., Ltd.METHODS: Hydroprednisone acetate was injected into all rats using intragluteal injection to produce necrosis models of bone cells of femoral head with the exception of the normal control group. The rats were drench with PSS, alendronate sodium, and PSS combined with alendronate in related groups, respectively. The experiment was performed 6 consecutive weeks with intramuscular administration once per week and drench administration once time per day. All rats were forced to stand at intervals four hours per day.MAIN OUTCOME MEASURES: Pathology changes of femoral head were observed under light microscope. The expression of bcl-2 and caspase-3 were detected by immunohistochemical method.RESULTS: Compared with the normal control group at 4 weeks after administration, phenomenon of femoral head necrosis was clear in the glucocorticoid model group, the spongy bone trabecula turned thinner or broke, the number of empty lacuna was obvious increased (t=2.629, P < 0.01). The number of bone trabecula and empty lacuna in the PSS group lies between the normal control and the glucocorticoid model groups (t=1.713, P < 0.01). However, the number of bone trabecula and empty lacuna in the alendronate sodium and PSS combined with alendronate group was near to that of the control group (t=1.462, t=1.657, P > 0.05). Compared with the glucocorticoid model, bcl-2 positive cell rate was obvious increased in the PSS, alendronate sodium and PSS combined with alendronate groups (t=2.146, P < 0.05; t=2.815, P < 0.01; t=2.947, P < 0.01), the caspase-3 positive cell rate was significantly decreased (t=2.97, P < 0.01; t=2.462, P < 0.01; t=1.854, P < 0.05), especially in the alendronate sodium and PSS combined with alendronate groups.CONCLUSION: A large dose of hormone may cause avascular necrosis of femoral head by decreasing expression of bcl-2 and increasing caspase-3. While alendronate sodium can prevent femoral head steroid-induced necrosis from osteocytes apoptosis by regulating expressions of bcl-2 and caspase-3.