运动负荷对大鼠激素性坏死股骨头内成骨细胞分化相关成熟基因的调控

Osteoblast differentiation and mature gene regulation inside femoral head with steroid-induced os- teonecrosis in response to mechanical stress stimulation

目的研究激素性坏死股骨头内Runx2 （ runt - related transcription factor - 2 ） ,、Osterix、骨钙素及AJ18基因表达和蛋白合成在机械应力作用下的动态变化。方法Wistar大鼠，共50只，雌雄各半；体重250～270g，平均260g。采用随机数字表法将大鼠分为正常组（10只）、实验组（40只）。实验组按20mg／kg剂量交替经大鼠双侧臀大肌注射地塞米松，每周1次；注射地塞米松后放在实验动物跑台上每周两次训练诱导制成大鼠激素性股骨头坏死早期动物模型。HE染色确定模型诱导成功后，停糖皮质激素注射，将实验组大鼠分为模型对照组、停激素运动4周和6周及8周组，提取大鼠股骨头内总RNA及总蛋白，行实时荧光定量PCR及Westernblot检测，观察在机械应力影响下激素性股骨头坏死时股骨头内Runx2、Osterix、骨钙素及AJ18基因表达、蛋白合成的动态变化。结果停激素运动4，6，8周时，大鼠股骨头内Runx2、Osterix和骨钙素的基因表达、蛋白合成较模型对照组减少，三者mRNA的表达量分别为模型对照组的0．181，0．164，0．137和0．422，0．303，0．265以及0．286，0．203，0．158倍，且随时间推移基因表达、蛋白合成呈下降趋势。停激素运动组大鼠股骨头内AJl8的基因表达、蛋白合成较模型对照组增多，AJ18mRNA的表达在停激素继续运动4，6，8周分别为模型对照组的2．639，4．272，5．317倍。结论激素性股骨头坏死早期，除激素因素外，不适当机械应力降低股骨头内Runx2、Osterix、骨钙素InRNA表达和蛋白合成，而AJ1l8mRNA表达和蛋白合成增加。

Objective To study dynamic changes of gene expressions and protein synthesis of Runx2 （runt-related transcription factor-2） , Osterix, osteoealein and AJ18 inside the femoral head with steroid-induced osteonecrosis after mechanical stress stimulation in rats. Methods A total of 50 Wist- ar rats （half male in sex） weighing 250-270 g （mean 260 g） were involved in this study and randomly di- vided into experimental group （40 rats） and normal group （ 10 rats）. The rats in experimental group were injected with dexamethasone （20 mg/kg） via bilateral gluteus maximus alternatively once a week and then trained on laboratory animal treadmill twice weekly to make rat model of femoral head necrosis. After i- dentifying the successfully induced model by Hematoxylin and cosin stain, glucocortieoid injection was ceased and the experimental group was randomly divided into model control group, 4 weeks, 6 weeks, 8 weeks groups after hormone training stopped. Then, total RNA and total protein were extracted from femo- ral head for detecting dynamic changes of genes expressions and proteins synthesis of Runx2, Osterix, os- teocalein and AJ18 after mechanical stress stimulation inside the femoral head with steroid-induced osteone- erosis by means of real-time quantitative PCR and Western blot assay. Results In 4 weeks, 6 weeks,8 weeks groups after hormone training stopped, the gene expressions and proteins synthesis of Runx2, Os- terix and osteocalcin were reduced more significantly compared with model control group, mRNA expres- sion values of Runx2, Osterix and osteocalcin were 0. 1809, 0. 1639, 0. 1374 and 0. 4219, 0. 3026, 0. 2652 and 0.2857, 0. 2027, 0. 1583 times of those in model control group. The expressions of Runx2, Osterix and osteocalein showed a downward trend with time. The mRNA expression and protein synthesis of AJ18 at 4th, 6th and 8th weeks ＇after hormone training stopped were 2. 6391,4.2718 and 5. 3165 times of model control group. Conclusions In addition to hormonal factors, inappropriate mechanical stress inhibits expressions and proteins synthesis of Runx2, Osterix and osteoealcin, while the expression and protein synthesis of A J18 are upgraded in early steroid-induced femoral head necrosis in rats.