中国组织工程研究 ›› 2025, Vol. 29 ›› Issue (6): 1159-1167.doi: 10.12307/2025.304

• 口腔组织构建 oral tissue construction • 上一篇    下一篇

杜仲促进去势大鼠牙槽骨成骨的作用

郑  琳1,靳文君1,罗珊珊1,黄  芮1,王  杰1,程余婷1,安哲庆1,熊  玥1,巩仔鹏2,廖  健1   

  1. 1贵州医科大学口腔医学院/附属口腔医院,贵州省贵阳市  550004;2贵州医科大学民族药与中药开发应用教育部工程研究中心,贵州省贵阳市550004

  • 收稿日期:2024-01-23 接受日期:2024-03-22 出版日期:2025-02-28 发布日期:2024-06-20
  • 通讯作者: 廖健,主任医师,教授,贵州医科大学口腔医学院/附属口腔医院,贵州省贵阳市 550004
  • 作者简介:郑琳,女,1998年生,贵州省贵阳市人,汉族,在读专业型硕士,主要从事口腔种植修复的基础与临床研究。
  • 基金资助:
    贵州省基础研究计划重点项目(黔科合基础-ZK[2023]重点037),项目负责人:廖健;贵州省卫生健康委科学技术基金项目(gzwkj2023-439),项目负责人:熊玥

Eucommia ulmoides promotes alveolar bone formation in ovariectomized rats 

Zheng Lin1, Jin Wenjun1, Luo Shanshan1, Huang Rui1, Wang Jie1, Cheng Yuting1, An Zheqing1, Xiong Yue1, Gong Zipeng2, Liao Jian1   

  1. 1School/Hospital of Stomatology, Guizhou Medical University, Guiyang 550004, Guizhou Province, China; 2Guizhou Provincial Engineering Research Center for Development and Application of Ethnic Medicines and Traditional Chinese Medicines, Guizhou Medical University, Guiyang 550004, Guizhou Province, China
  • Received:2024-01-23 Accepted:2024-03-22 Online:2025-02-28 Published:2024-06-20
  • Contact: Liao Jian, Chief physician, Professor, School/Hospital of Stomatology, Guizhou Medical University, Guiyang 550004, Guizhou Province, China
  • About author:Zheng Lin, Master candidate, School/Hospital of Stomatology, Guizhou Medical University, Guiyang 550004, Guizhou Province, China
  • Supported by:
    the Key Project of Guizhou Basic Research Program, No. ZK[2023]037 (to LJ); Science and Technology Fund Project of Guizhou Provincial Health Commission, No. gzwkj2023-439 (to XY)

摘要:




文题释义:
杜仲:是中国的一种传统中药材,其皮易折断,断面有细密、银白色、富弹性的橡胶丝相连;性甘温,具有补肝肾、强筋骨的功效,近年来已用于临床治疗骨质疏松等疾病。
Wnt/β-连环蛋白信号通路:是Wnt信号通路的经典通路,β-连环蛋白是该信号通路的关键分子,该通路的激活使得β-连环蛋白在细胞质内得以稳定,积聚一定量后进入细胞核,参与下游靶基因的转录。

背景:杜仲具有一定的成骨作用,能够促进成骨细胞的增殖及分化,但其对牙槽骨骨形成及Wnt/β-连环蛋白信号通路的影响尚不明确。
目的:基于Wnt/β-连环蛋白信号通路探讨杜仲促进去势大鼠牙槽骨成骨的作用机制。
方法:采用随机数字表法将60只雌性SD大鼠分为空白对照组、假手术组、模型组、杜仲低剂量组、杜仲高剂量组,每组12只。模型组、杜仲低剂量组、杜仲高剂量组通过双侧卵巢切除术建立骨质疏松大鼠模型,假手术组切除双侧卵巢周围同等质量的脂肪组织。造模3个月后,杜仲低、高剂量组大鼠分别灌胃给予2.1,4.2 g/(kg•d)的杜仲,假手术组和模型组大鼠灌胃给予生理盐水。药物干预12周后取材,通过Micro-CT观察牙槽骨骨量变化,苏木精-伊红染色观察牙槽骨病理结构变化,ELISA法检测血清中碱性磷酸酶、骨钙素水平,Western blot检测牙槽骨组织中β-连环蛋白、卷曲蛋白9的蛋白表达,RT-qPCR检测牙槽骨组织中骨钙素、Runx2、碱性磷酸酶、β-连环蛋白、卷曲蛋白9 mRNA的表达。
结果与结论:①与空白对照组比较,模型组骨体积分数、骨小梁数目、骨小梁厚度、骨密度均降低(P < 0.05),骨小梁分离度升高(P < 0.05);病理观察可见骨小梁排列紊乱、不规则,骨小梁变细或断裂,骨髓腔变大,骨量明显减少;血清中碱性磷酸酶水平上升(P < 0.05),骨钙素水平下降(P < 0.05);碱性磷酸酶、骨钙素、Runx2、β-连环蛋白、卷曲蛋白9的mRNA表达均降低(P < 0.05);β-连环蛋白、卷曲蛋白9的蛋白表达降低(P < 0.05)。②与模型组比较,杜仲低、高剂量组大鼠牙槽骨骨体积分数、骨小梁数目、骨小梁厚度、骨密度均升高(P < 0.05),骨小梁分离度降低(P < 0.05);杜仲低、高剂量组骨小梁排列稍整齐,骨小梁较粗大,骨量增加;杜仲低、高剂量组血清中碱性磷酸酶水平下降(P < 0.05),骨钙素水平上升(P < 0.05);杜仲低剂量组碱性磷酸酶、骨钙素、β-连环蛋白、卷曲蛋白9的 mRNA表达升高(P < 0.05),杜仲高剂量组碱性磷酸酶、骨钙素、Runx2、β-连环蛋白、卷曲蛋白9的mRNA表达升高(P < 0.05);杜仲低剂量组卷曲蛋白9蛋白表达升高(P < 0.05),杜仲高剂量组β-连环蛋白、卷曲蛋白9的蛋白表达升高(P < 0.05);③与杜仲低剂量组比较,杜仲高剂量组对上述指标的改善作用更显著;④提示:杜仲能有效促进牙槽骨骨形成,其作用机制可能与激活Wnt/β-连环蛋白信号通路有关。
https://orcid.org/0009-0006-6112-0702(郑琳)

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: 杜仲, 牙槽骨, 骨形成, 去势大鼠, Wnt/β-连环蛋白信号通路, 骨质疏松

Abstract:
BACKGROUND: 
Eucommia ulmoides has a certain osteogenic effect, which can promote the proliferation and differentiation of osteoblasts. However, it is unclear whether Eucommia ulmoides has effects on alveolar bone formation and Wnt/β-Catenin signaling pathway.
OBJECTIVE: To investigate the mechanism by which Eucommia ulmoides promotes alveolar bone formation in ovariectomized rats based on the Wnt/β-Catenin signaling pathway.
METHODS: Sixty female Sprague-Dawley rats were selected and randomly divided into five groups: blank control group, sham-operation group, model group, low-dose group Eucommia ulmoides group, and high-dose Eucommia ulmoides group, with twelve rats in each group. Osteoporosis animal models were constructed by bilateral oophorectomy in the model group and the low-dose and high-dose Eucommia ulmoides groups. The sham-operation group underwent the same method to remove adipose tissue of equal mass around the bilateral ovaries. Three months after surgery, the low- and high-dose Eucommia ulmoides groups were given 2.1 g/kg/d and 4.2 g/kg/d Eucommia ulmoides by gavage, respectively. The sham-operation group and model group were given the same amount of physiological saline by gavage. After 12 weeks of drug intervention, the changes in alveolar bone mass of rats in each group were observed through Micro-CT; hematoxylin-eosin staining was used to observe the pathological structural changes of alveolar bone in rats; enzyme linked immunosorbent assay was used to detect the expression levels of alkaline phosphatase and osteocalcin in the serum of rats; western blot was used to detect the expression levels of β-Catenin and Frizzled9 receptor proteins in the alveolar bone of rats; and real-time fluorescence quantitative PCR was used to detect the expression of osteocalcin, Runt-related transcription factor 2 (Runx2), alkaline phosphatase, β-catenin, and frizzled9 mRNAs in alveolar bone tissues of rats.
RESULTS AND CONCLUSION: Compared with the blank control group, bone volume fraction, trabecular number, trabecular thickness, and bone mineral density were reduced in the model group (P < 0.05), and trabecular separation was elevated (P < 0.05). Pathological observation showed that the arrangement of trabeculae was disordered and irregular, the trabeculae were thinned or broken, and the marrow cavity was enlarged in the model group, with a significant reduction in bone volume; the level of alkaline phosphatase in the serum was increased (P < 0.05), and the level of osteocalcin was decreased (P < 0.05); mRNA expression of alkaline phosphatase, osteocalcin, Runx2, β-catenin, and frizzled9 were decreased (P < 0.05); protein expression of β-Catenin and Frizzled9 was decreased (P < 0.05). Compared with the model group, the low- and high-dose Eucommia ulmoides groups showed an increase in bone volume fraction, trabecular number, trabecular thickness, and bone mineral density (P < 0.05) and a decrease in trabecular separation (P < 0.05). In the low- and high-dose Eucommia ulmoides groups, bone trabeculae were slightly aligned and thickened, with a significant increase in bone mass. Compared with the model group, the serum level of alkaline phosphatase was reduced (P < 0.05) and the serum level of osteocalcin was elevated (P < 0.05) in the low- and high-dose Eucommia ulmoides groups. Compared with the model group, the mRNA expression of alkaline phosphatase, osteocalcin, Runx2, β-catenin, and frizzled9 were increased in the low- and high-dose Eucommia ulmoides groups (P < 0.05). Compared with the model group, the protein expression of Frizzled9 was increased in the low-dose Eucommia ulmoides group (P < 0.05), while the protein expression of β-Catenin and Frizzled9 was increased in the high-dose Eucommia ulmoides group (P < 0.05). Compared with the low-dose Eucommia ulmoides group, the high-dose Eucommia ulmoides group had a more significant improvement in the above indexes. To conclude, Eucommia ulmoides can effectively promote the alveolar bone formation, and its mechanism of action might be related to the activation of the Wnt/β-catenin signaling pathway.

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