伊班膦酸钠对卵巢切除所致骨质疏松模型大鼠的影响及机制

Effects and mechanisms of sodium ibandronate in ovariectomy-induced osteoporosis rats

背景:双膦酸盐类药物是一种新型骨吸收抑制剂,能有效抑制破骨细胞的活性和功能。目的:观察伊班膦酸钠对骨质疏松大鼠髁突软骨中牙本质基质蛋白1、Caspase3及Bcl-2、Bax表达的影响。方法:将36只雌性大鼠随机分组为假手术组、骨质疏松组及伊班膦酸钠组,每组各12只。假手术组术中不切除卵巢,骨质疏松组和伊班膦酸钠组切除双侧卵巢,术后7 d给予伊班膦酸钠组大鼠10μg/kg/次的伊班膦酸钠,每隔7 d腹腔注射1次,90 d后取材。microCT测量各组大鼠骨密度变化;采用甲苯胺蓝染色、TUNEL染色观察髁突软骨变化;免疫组织化学检测牙本质基质蛋白1蛋白表达;Western blot检测Caspase3、Bcl-2、Bax蛋白表达。实验方案经南华医院动物实验伦理委员会批准(批准号为SLXD_201804010)。结果与结论:①与假手术组或伊班膦酸钠组比较,骨质疏松组骨密度值显著降低(P<0.05);②甲苯胺蓝染色结果,伊班膦酸钠组髁突软骨肥大层明显厚于骨质疏松组;与假手术组或伊班膦酸钠组相比,骨质疏松组大鼠髁突软骨及软骨下骨凋亡细胞数均明显上升(P<0.05);③与假手术组比较,骨质疏松组TRAP阳性细胞数增多,而经伊班膦酸钠治疗后TRAP阳性细胞数下降(P<0.05);④骨质疏松组牙本质基质蛋白1蛋白表达较假手术组明显下降,而经伊班膦酸钠治疗后牙本质基质蛋白1蛋白表达上升(P<0.05);⑤与假手术组相比,骨质疏松组Caspase3、Bax表达明显上调,Bcl-2表达下降,而经伊班膦酸钠治疗后Caspase3、Bax表达水平明显下降,Bcl-2表达水平上调;⑥结果说明,伊班膦酸钠能够抑制骨质疏松状态下髁突软骨细胞凋亡及破骨细胞数量,可能与调控Caspase3、Bcl-2、Bax及牙本质基质蛋白1表达有关。

BACKGROUND:Bisphosphonates are a novel inhibitor of bone resorption that can inhibit the activity and function of osteoclasts.OBJECTIVE:To observe the effects of sodium ibandronate on the expression of dentin matrix protein 1,Caspace3,Bcl-2 and Bax in condylar cartilage in osteoporosis rats.METHODS:Thirty-six female rats were randomly divided into sham group,osteoporosis group and sodium ibandronate group,twelve in each group.The sham group did not excise ovaries during surgery.Bilateral ovaries of rats were removed in the osteoporosis and sodium ibandronate groups.On the 7th day after operation,rats in the sodium ibandronate group were intraperitoneally given sodium ibandronate 10μg/kg,once every 7 days.After 90 days,the rat ovaries were taken.Bone mineral density was measured in each group.The changes of condylar cartilage were observed by toluidine blue staining and TUNEL staining.The expression of dentin matrix protein 1 protein was detected by immunohistochemistry.The levels of Caspase3,Bcl-2 and Bax were detected by western blot assay.The study protocol was approved by the Animal Ethics Committee of Nanhua Hospital in China with the approval No.SLXD_201804010.RESULTS AND CONCLUSION:Compared with the sham group or sodium ibandronate group,the bone mineral density in the osteoporosis group was significantly decreased(P<0.05).The results of toluidine blue staining showed that the hypertrophic layer of condylar cartilage in the sodium ibandronate group was significantly thicker than that in the osteoporosis group.Compared with the sham group or sodium ibandronate group,the number of apoptotic cells in condylar cartilage and subchondral bone increased significantly in the osteoporosis group(P<0.05).The expression of dentin matrix protein 1 protein was significantly lower in the osteoporosis group than the sham group,but it increased after treatment with sodium ibandronate(P<0.05).Compared with the sham group,the expression of Caspase 3 and Bax in the osteoporosis group increased significantly,and the expression of Bcl-2 decreased.However,treatment with sodium ibandronate decreased the expression of Caspase 3 and Bax and increased the expression of Bcl-2 significantly.Overall,our findings reveal that sodium ibandronate can inhibit the apoptosis of condylar chondrocytes and the number of osteoclasts in osteoporotic state,which may be related to the regulation of Caspase 3,Bcl-2,Bax and dentin matrix protein 1 expression.