中国组织工程研究 ›› 2022, Vol. 26 ›› Issue (29): 4638-4642.doi: 10.12307/2022.840

• 骨组织构建 bone tissue construction • 上一篇    下一篇

骨碎补总黄酮对骨组织重建及骨形态发生蛋白2、血管内皮生长因子、CD31的影响

赵  盾1,方  斌2,易春智2,何敏聪2,郑嘉乾1,李  悦2   

  1. 1广州中医药大学第一临床医学院,广东省广州市  510000;2广州中医药大学第一附属医院骨肿瘤科,广东省广州市  510000
  • 收稿日期:2021-06-07 接受日期:2021-11-23 出版日期:2022-10-18 发布日期:2022-03-26
  • 通讯作者: 李悦,医师,博士,广州中医药大学第一附属医院骨肿瘤科,广东省广州市 510000
  • 作者简介:赵盾,男,1997年生,河南省新乡市人,汉族,硕士,主要从事骨与软组织肿瘤研究。
  • 基金资助:
    国家自然科学基金青年科学基金项目(8160150894),项目负责人:李悦;广东省教育厅高校科研项目青年创新人才项目(2019KQNCX017),项目负责人:何敏聪

Effects of total flavonoids of Rhizoma drynariae on bone remodeling and expression of bone morphogenetic protein-2, vascular endothelial growth factor and CD31

Zhao Dun1, Fang Bin2, Yi Chunzhi2, He Mincong2, Zheng Jiaqian1, Li Yue2   

  1. 1The First Clinical Medical College, Guangzhou University of Chinese Medicine, Guangzhou 510000, Guangdong Province, China; 2Department of Orthopedic Oncology, The First Affiliated Hospital of Guangzhou University of Chinese Medicine, Guangzhou 510000, Guangdong Province, China
  • Received:2021-06-07 Accepted:2021-11-23 Online:2022-10-18 Published:2022-03-26
  • Contact: Li Yue, Physician, MD, Department of Orthopedic Oncology, The First Affiliated Hospital of Guangzhou University of Chinese Medicine, Guangzhou 510000, Guangdong Province, China
  • About author:Zhao Dun, Master, The First Clinical Medical College, Guangzhou University of Chinese Medicine, Guangzhou 510000, Guangdong Province, China
  • Supported by:
    the National Natural Science Foundation of China (Youth Program), No. 8160150894 (to LY); Guangdong Provincial Education Department Scientific Innovative Project for Young Talents, No. 2019KQNCX017 (to HMC)

摘要:

文题释义:
骨碎补总黄酮:骨碎补是补肾法思想指导下的代表药物,中医学认为其性温、味苦,归肾、肝经,具有补肾强骨、续伤止痛之功效。现代药理学研究证实,骨碎补具有促骨折愈合、抗骨质疏松、 固牙健齿、活血化瘀等多种药理活性。骨碎补总黄酮是干燥根茎中提取的有效成分,在实验及临床研究中使用非常广泛。
Masquelet技术:为法国医生所创,其过程分为2步:①诱导膜形成期:首先对骨缺损区进行彻底清创,然后用聚甲基丙烯酸甲酯骨水泥填充塑形为棒状并连接骨断端,用软组织包裹骨水泥,并固定骨缺损两端;②骨组织重建期:在一期术后6-8周,切开诱导膜,取出骨水泥,打通纤维膜两端,使骨髓腔与诱导膜囊腔相通,同时在膜腔内填满切碎的自体骨或骨替代材料,借助诱导膜丰富的血供实现成骨。

背景:多项研究表明骨碎补总黄酮具有促进骨愈合的作用,然而其发挥作用的具体分子机制尚不清楚。
目的:探讨骨碎补总黄酮对骨缺损区域骨组织重建的影响。
方法:将36只雄性SD大鼠随机分为空白组、诱导膜组、诱导膜+中药组,每组12只,均于右侧股骨中段截取4 mm骨组织后采用钢板固定,诱导膜组、诱导膜+中药组置入聚甲基丙烯酸甲酯骨水泥旷置;4周后取出之前旷置的骨水泥,诱导膜组及诱导膜+中药组植入自体尾骨,术后诱导膜+中药组灌胃给予骨碎补总黄酮 0.22 g/(kg•d),其余两组灌胃给予等量生理盐水,直至取材。自体骨植入6周后,通过X射线片、组织形态学观察缺损区域新生骨组织情况,免疫组化染色检测骨形态发生蛋白2、血管内皮生长因子、CD31(血小板内皮黏附分子)的表达。
结果与结论:①X射线片:诱导膜+中药组骨缺损区已形成连续骨质,髓腔结构形成,骨皮质重建良好;诱导膜组未形成完整的骨皮质,髓腔未完全形成,骨重建不连续;空白组仍可见清晰骨缺损区域;②组织形态学观察:空白组缺损区域无连续的骨质形成,其他两组均有不同程度的骨质形成,其中诱导膜+中药组处于骨质塑形阶段,诱导膜组虽有骨质生成,但成骨不均匀,局部仍有少量炎症细胞浸润和结缔组织增生;③免疫组化染色:诱导膜+中药组骨形态发生蛋白2、血管内皮生长因子蛋白表达高于空白组、诱导膜组(P < 0.05),CD31蛋白表达高于空白组(P < 0.05);④结表表明:骨碎补总黄酮可以调控骨形态发生蛋白2、血管内皮生长因子蛋白的表达,促进骨缺损区的骨质矿化和重建。 

https://orcid.org/0000-0002-7636-4092 (赵盾) 

中国组织工程研究杂志出版内容重点:组织构建;骨细胞;软骨细胞;细胞培养;成纤维细胞;血管内皮细胞;骨质疏松;组织工程

关键词: 诱导膜, Masquelet技术, 骨碎补总黄酮, 骨缺损, 骨组织重建, 骨形态发生蛋白2, 血管内皮生长因子, CD31

Abstract: BACKGROUND: Numerous studies have shown that the total flavonoids of Rhizoma drynariae can promote bone healing, but the specific molecular mechanism is still unclear.
OBJECTIVE: To investigate the effect of total flavones of Rhizoma drynariae on bone remodeling after bone defects.
METHODS: Thirty-six male Sprague-Dawley rats were randomly divided into three groups (n=12 per group): blank group, induced membrane group, induced membrane+traditional Chinese medicine group). After a 4-mm bone tissue sample was taken from the middle part of the right femur of each rat, the femur was fixed with a steel plate and inserted with polymethylmethacrylatebone cement. Four weeks after fixation, the second operation was performed to remove the previously detached bone cement. The blank group did not receive bone graft, and the induced membrane group and the induced membrane+Chinese medicine group received autologous coccyx bone graft. Afterwards, the induction membrane+traditional Chinese medicine group was given the total flavonoids of Rhizoma drynariae 0.22 g/(kg•d) by gavage, and the other two groups were given the same amount of normal saline by gavage. Six weeks after bone grafting, X-ray and histomorphological examinations were performed to observe the formation of new bone tissue in the defect area. Immunohistochemistry staining was used to detect the protein expressions of bone morphogenetic protein 2, vascular endothelial growth factor, and CD31 (platelet-endothelial cell adhesion molecule) in bone tissue.       
RESULTS AND CONCLUSION: X-ray films indicated that there was continuous bone tissue in the bone defect area, the medullary cavity was formed, and the bone cortex was reconstructed well in the induced membrane+traditional Chinese medicine group; the induced membrane group did not form a complete cortical bone, in which the medullary cavity was not completely formed and no continuous bone tissue formed; the bone defect area in the blank group was still visible. Histomorphological observations indicated that there was no continuous bone formation in the defect area of the blank group, while new bone tissue was formed to different extent in the defect area of the other two groups. The induced membrane+traditional Chinese medicine group was in the stage of bone shaping. In the induced membrane group, new bone tissue formed unevenly and there was still inflammatory cell infiltration and connective tissue hyperplasia in the defect area. Immunohistochemical findings revealed that the protein expression of bone morphogenetic protein-2 and vascular endothelial growth factor was significantly higher in the induced membrane+traditional Chinese medicine group than the other two groups (P < 0.05), while the protein expression of CD31 was significantly higher in the induced membrane+traditional Chinese medicine group than the blank group (P < 0.05). To conclude, total flavones of Rhizoma drynariae can regulate the expression of bone morphogenetic protein-2 and vascular endothelial growth factor proteins and promote bone mineralization and remodleing in the bone defect area. 

Key words: induced membrane, Masquelet technique, total flavones of Rhizoma drynariae, bone defect, bone remodeling, bone morphogenetic protein-2, vascular endothelial growth factor, CD31

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