苗药九仙罗汉接骨汤干预胫骨骨折模型大鼠的成骨分化及骨折愈合.

Effect of Miao medicine Jiuxian Luohan Jiegu Decoction on osteogenic differentiation and fracture healing in tibial fracture rats.

BACKGROUND: Jiuxian Luohan Jiegu Decoction can promote blood circulation, remove blood stasis, reduce swelling, and relieve pain. However, its specific mechanism underlying tibial fractures has not yet been fully elucidated. OBJECTIVE: To explore the effect of Miao medicine Jiuxian Luohan Jiegu Decoction on the osteogenic differentiation and fracture healing ability of rats with tibial fractures mediated by bone morphogenetic proteins (BMPs)/Smad signaling pathway. METHODS: Sixty Sprague-Dawley rats were randomly divided into a fracture group, a Miao medicine group, and a Panax notoginseng group. The Miao medicine group was given Jiuxian Luohan Jiegu Decoction (2 mL/kg) by gavage; the Panax notoginseng group was given the same volume of panax notoginseng powder (1 mL/kg) by gavage; and the model group was given the same amount of distilled water by gavage, once a day, for 28 days in total. Alizarin red staining was used to measure osteogenic differentiation of bone marrow mesenchymal stem cells. RT-PCR and western blot were used to measure BMP-2, transforming growth factor-β, Smad1, and Smad2 at mRNA and protein levels. Three-point bending test was used to measure biomechanical indexes. Hematoxylin-eosin staining was used to detect bone tissue morphology. ELISA was used to measure serum vascular endothelial growth factor and insulin growth factor-1. RESULTS AND CONCLUSION: (1) Compared with the model group, the differentiated cells of the Miao medicine group and the Panax notoginseng group grew in multiple layers and formed focal shapes, and a large number of calcium nodules were formed. The Miao medicine group showed better effects than the Panax notoginseng group. (2) Compared with the model group, the mRNA levels of BMP-2, transforming growth factor-β, Smad1, and Smad2 were increased in the Panax notoginseng group and the Miao medicine group increased (P < 0.05). (3) The maximum load and stiffness of the panax notoginseng group and the Miao medicine group were higher than those of the model group (P < 0.05). (4) The bone trabeculae of the model group were thin, arranged disorderly, and relatively loosely, the space between the medullary cavities was large, and a large number of uncalcified chondrocytes appeared. Similar bone tissue morphology was found in the Panax notoginseng group and the Miao medicine group is similar, in which the bone trabecula was thicker, the medullary cavity space was reduced, the distribution was more orderly, and the chondrocytes were more ossified compared with those in the model group. (5) Compared with the model group, the expression levels of vascular endothelial growth factor and insulin growth factor-1 in the Panax notoginseng group and the Miao medicine group increased to a certain extent (P < 0.05). (6) Radiological observations showed that there were obvious fracture lines in the model group, while the fractured ends of the samples in the Panax notoginseng group and the Miao medicine group reached complete healing. However, the scab density at fractured ends of the Miao medicine group was slightly higher than that of the Panax notoginseng group. (7) To conclude, Miao medicine (Jiuxian Luohan Jiegu Decoction) can promote osteoblast differentiation and accelerate fracture healing. Its mechanism of action may be achieved by activating the BMPs/Smad signaling pathway. [ABSTRACT FROM AUTHOR]

背景：九仙罗汉接骨汤具有活血散瘀、接骨消肿、止痛的作用，但目前其对胫骨骨折的具体作用机制尚未完全掌握。 目的：探究苗药九仙罗汉接骨汤介导BMPs/Smad信号通路对胫骨骨折大鼠成骨分化及骨折愈合能力的影响。 方法：60只SD大鼠随机分为模型组、苗药组、三七组，采用改良钻孔法建立不完全性胫骨骨折模型。造模成功当天，苗药组给予苗药九 仙罗汉接骨汤煎剂(2 mL/kg)灌胃；三七组给予同体积三七粉液(1 mL/kg)灌胃；模型组给予同等计量蒸馏水灌胃，1次/d，共28 d。28 d后 麻醉下取大鼠胫骨组织，茜素红染色测骨髓间充质干细胞成骨分化；三点弯曲试验测胫骨生物力学指标；苏木精-伊红染色测骨组织形 态；ELISA检测血清中血管内皮生长因子、胰岛素样生长因子1表达水平；RT-PCR和Western blot检测骨形态发生蛋白 2、转化生长因子β、 Smad1、Smad2的mRNA和蛋白水平；实验已经过贵州中医药大学第二附属医院动物伦理委员会批准(2018-072)。 结果与结论：①与模型组比较，苗药组与三七组骨髓间充质干细胞成骨分化后细胞多层重叠生长且形成灶状，有大量的钙结节形成，且 苗药组较三七组效果更佳；②与模型组相比，三七组与苗药组骨形态发生蛋白 2、转化生长因子β 、Smad1、Smad2的mRNA和蛋白水平均 有所上升(P < 0.05)；③三七组与苗药组最大载荷、刚度水平较模型组均有所升高(P < 0.05)；④模型组骨小梁较细，髓腔间空隙大，杂乱 无章，较为松散，有大量的未钙化软骨细胞；三七组与苗药组大鼠骨组织形态较为接近，均表现为骨小梁较粗，髓腔空隙减小，分布较 为有序，软骨细胞骨化较多；⑤与模型组相比，三七组与苗药组血管内皮生长因子、胰岛素样生长因子1表达水平有一定程度的提升(P < 0.05)；⑥模型组大鼠存在明显骨折线，三七组与苗药组标本骨折断端均达到完全愈合，但与三七组相比，苗药组标本骨折断端骨痂密度 稍高；⑦结果说明，苗药九仙罗汉接骨汤可促进成骨细胞分化，加速骨折愈合，其作用机制可能是通过激活BMPs/Smad信号通路来实现。 [ABSTRACT FROM AUTHOR]

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