载骨碎补总黄酮磷酸钙骨水泥对骨缺损模型大鼠诱导膜中成骨细胞分化的影响及其机制研究
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篇名: 载骨碎补总黄酮磷酸钙骨水泥对骨缺损模型大鼠诱导膜中成骨细胞分化的影响及其机制研究
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摘要: 目的:探究载骨碎补总黄酮磷酸钙骨水泥对骨缺损模型大鼠诱导膜成骨分化的作用及其机制。方法:分别以磷酸钙骨水泥(CPC)、聚甲基丙烯酸甲酯(PMMA)骨水泥为载体,以强骨胶囊内容物(有效成分为骨碎补总黄酮)制备载药CPC和载药PMMA骨水泥。选取64只雄性SD大鼠,随机分为载药CPC组、载药PMMA骨水泥组、未载药CPC组、未载药PMMA骨水泥组,每组16只。分离大鼠股骨并行截骨,以制备骨缺损模型,然后分别植入相应骨水泥。造模4周后,各组大鼠切开并保护好诱导膜,取出骨水泥,植入自体松质骨。于造模后第4周对大鼠后肢骨进行X射线拍片;于造模后第4周和植骨后第6周时分别取大鼠骨缺损区诱导膜和新生骨,采用苏木精-伊红染色法观察诱导膜组织形态学,并测定新生骨组织的骨小梁宽度及成骨细胞数;采用免疫组化法检测诱导膜中人骨成型蛋白2(BMP-2)、血管内皮生长因子(VEGF)的蛋白表达水平;采用Western blotting法检测新生骨组织中Smad1、Smad4、Smad7的蛋白表达水平。结果:与其余3组比较,载药CPC组大鼠骨缺损区的骨水泥降解更明显,且可见诱导膜组织形成,骨缺损区域更小;诱导膜中毛细血管内皮细胞丰富、排列有序;新生骨组织中骨小梁宽度及成骨细胞数均显著增加(P<0.05);诱导膜中BMP-2、VEGF的蛋白表达水平和新生骨组织中Smad1、Smad4、Smad7的蛋白表达水平均显著升高(P<0.05)。结论:载骨碎补总黄酮CPC具有促进骨缺损模型大鼠诱导膜成骨的作用。这可能是通过激活BMP-2/Smad通路,从而调节成骨细胞分化;同时通过提高VEGF表达,促进血管内皮细胞分化,加快形成毛细血管网,从而促进骨愈合。
ABSTRACT: OBJECTIVE: To investigate the effects and its mechanism of calcium phosphate bone cement (CPC) loading total flavonoids of Davallia mariesii on osteogenic differentiation of induced membrane in rats. METHODS: Drug-loading CPC and drug-loading polymethyl methacrylate (PMMA) cement were prepared with the contents of Qianggu capsules (total flavonoids of D. mariesii as active ingredient) using CPC and PMMA cement as carrier. Totally 64 male SD rats were randomly divided into drug-loading CPC group, drug-loading PMMA cement group, no-drug CPC group, no-drug PMMA cement group, with 16 rats in each group. The femur of rats was separated and osteotomized to prepare bone defect model, and then the corresponding bone cement was implanted. Four weeks after modeling, the induced membranes of rats were cut and protected. Bone cement was taken out and autogenous cancellous bone was implanted. At the 4th week after modeling, X-ray photographs were taken on the hind limb bones of rats. At the 4th week after modeling and 6th week after bone grafting, induced membranes and new bone were taken from the bone defect area of rats respectively. HE staining was used to observe the morphology of induced membrane, and the width of bone rabecular and the number of osteoblasts of new bone tissue were measured. Immunohistochemistry was used to detect the protein expression of BMP-2 and VEGF in induced membrane. Western blotting assay was used to detect the protein expression of Smad1, Smad4 and Smad7 in new bone. RESULTS: Compared with other 3 groups, the degradation of bone cement in drug-loading CPC group was more obvious in the bone defect areas, which showed that the formation of induced membrane was observed and the bone defect areas were smaller; capillary endothelial cells were abundant and orderly arranged in the induced membranes, and the width of bone trabeculae and the number of osteoblasts in the new bone tissue increased significantly (P<0.05); the protein expression of BMP-2 and VEGF in the induced membrane, the protein expression of Smad1, Smad4 and Smad7 in new bone were increased significantly (P<0.05). CONCLUSIONS: CPC loading total flavonoids of D. mariesii promotes the formation of induced membrane osteoblast in bone defect model rats, which may be associated with regulating osteoblast differentiation by activating BMP-2/Smad pathway; at the same time, it can promote bone healing by promoting the differentiation of vascular endothelial cells, accelerating the formation of capillary network and increasing the expression of vascular endothelial cells.
期刊: 2019年第30卷第10期
作者: 董航,黄嘉华,麦喆钘,陈柏行,黄培镇,蔡群斌,陈超,纪树亮,孙伟鹏,黄尹滢,周琦石
AUTHORS: DONG Hang,HUANG Jiahua,MAI Zhexing,CHEN Boxing,HUANG Peizhen,CAI Qunbin,CHEN Chao,JI Shuliang,SUN Weipeng,HUANG Yinying,ZHOU Qishi
关键字: 骨碎补;总黄酮;磷酸钙骨水泥;诱导膜;骨缺损;成骨分化;机制;大鼠
KEYWORDS: Davallia mariesii;Total flavonoids; Calcium phosphate bone cement;Induced membrane;Bone defects;Osteogenic differentiation; Mechanism; Rat
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