创伤性股骨头缺血坏死过程中凋亡调控基因bcl-2、bax的表达及意义

背景：由于骨细胞是股骨头的主要功能细胞，因此股骨头中骨细胞的凋亡及影响凋亡的因子已经成为目前探讨缺血性股骨头坏死发病机制的研究热点。 目的：观察兔股骨头缺血坏死过程中凋亡调控基因bcl-2、bax 的表达变化，探讨股骨头缺血坏死的发病机制。 设计、时间及地点：动物观察试验，于2007-12/2008-09在遵义医学院珠海校区中心实验室完成。 材料：35只新西兰白兔随机分为对照组和实验组(缺血后3，6，12，24，48，96 h)共7组，每组5只。 方法：实验组采用开放手术中断兔股骨头血供致股骨头缺血坏死；对照组动物仅进行皮肤切开和肌肉分离至关节囊的手术。在缺血的不同时间段对股骨头内骨细胞以及Bcl-2、Bax两种蛋白进行测试及比较。 主要观察指标：应用苏木精-伊红染色，免疫组织化学(ABC法)来观测股骨头缺血后3，6，12，24，48，96 h股骨头内骨细胞陷窝变化及Bcl-2、Bax蛋白表达的变化情况。 结果：缺血12 h内苏木精-伊红染色见骨细胞变化较少，48 h出现部分骨细胞和成骨细胞消失，96 h空骨陷窝出现百分比明显高于48 h和对照组的 (P < 0.01)。Bcl-2在缺血后3 h开始出现阳性表达，缺血后12 h出现高峰，然后缓慢下降；Bax在缺血后3 h开始出现阳性表达，高峰出现在缺血后24 h；bcl-2/bax在24 h最小，然后逐渐升高。 结论：创伤性股骨头缺血坏死过程中凋亡基因bcl-2、bax表达变化说明其参与了股骨头细胞凋亡及股骨头缺血坏死过程，结合bcl-2/bax比率变化显示bcl-2、bax在缺血性股骨头坏死中通过调控骨细胞凋亡起作用。

Expression and significance of bcl-2 and bax genes during the development of traumatic avascular necrosis of the femoral head

BACKGROUND: Osteocytes are the main function cells in the femoral head. Therefore, the study of the apoptosis of osteocytes and the correlative stress genes has become the research focus for exploring the pathomechanism underlying avascular necrosis of femoral head. OBJECTIVE: To observe the expression changes of bcl-2 and bax genes during the development of traumatic avascular necrosis of femoral head in rabbits, and explore the pathomechanism underlying this disease. DESIGN, TIME AND SETTING: An animal observational experiment was performed at the Central Laboratory of Zhuhai Campus of Zunyi Medical College between December 2007 and September 2008. MATERIALS: A total of 35 New Zealand rabbits were randomly divided into 7 groups, namely, the control group and the experiment group which are subdivided into six ones at the time points of 3, 6, 12, 24, 48 and 96 hours postischemia respectively , with 5 animals in each group. METHODS: Rabbits in the experiment group were given an open surgery of interrupting the blood supply of their femoral heads to lead to avascular necrosis; rabbits in the control group only received skin incision and muscle-sparing ended in their joint capsules. Osteocytes, Bcl-2 and Bax proteins in femoral heads were determined and compared at different time points postischemia. MAIN OUTCOME MEASURES: Lacunae changes of osteocytes in femoral heads were detected with hematoxylin-eosin (HE) staining method, and the expression of Bcl-2 and Bax proteins with immunohistochemistry staining methods (ABC methods), after 3, 6, 12, 24, 48 and 96 hours of ischemia respectively. RESULTS: According to HE staining, ischemia of 12 hours or less resulted in no osteocyte lacunae change; after 48 hours of ischemia, parts of osteocytes and osteoblasts disappeared; at hour 96 postischemia, the percentage of empty osteocyte lacunae was higher obviously than that at hour 48 or that in the control group (P < 0.01). Bcl-2 expression began to increase at hour 3 postischemia, and reached a peak at hour 12, after which it decreased slowly; Bax expression begin to increase at hour 3 after ischeamia too, but with a peak at hour 24. bcl-2/bax kept a minimal at hour 24 and then gradually increased. CONCLUSION: The changed expression of bcl-2 and bax genes during the traumatic avascular necrosis of femoral head shows that bcl-2 and bax are involved in the apoptosis of osteocytes and the avascular necrosis of femoral head. Moreover, their ratio changes indicated that they play their role in avascular necrosis of femoral head in the way of influencing the apoptosis of osteocytes.