化瘀补肾方对去卵巢小鼠骨折愈合及氧化应激的影响

程韶; 陶渝仁; 舒冰; 王晶; 赵永见; 赵世天; 张岩; 唐德志; 赵东峰; 孙悦礼; 施杞; 王拥军

doi:10.16306/j.1008-861x.2021.04.012

您当前的位置：

首页 >

文章列表页 >

化瘀补肾方对去卵巢小鼠骨折愈合及氧化应激的影响

实验研究 | 更新时间：2022-09-09

- 化瘀补肾方对去卵巢小鼠骨折愈合及氧化应激的影响
- Effects of Huayu Bushen Formula on fracture healing and oxidative stress in ovariectomized mice
- 上海中医药大学学报 2021年35卷第4期页码：79-88
- 作者机构：
  
  1.上海中医药大学附属龙华医院（上海　200032）
  2.上海市中医药研究院脊柱病研究所（上海　200032）
  3.教育部筋骨理论与治法重点实验室（上海　200032）
- 作者简介：
  
  程韶，男，在读博士生，主要从事中医药防治慢性筋骨疾病研究。陶渝仁，男，在读硕士生，主要从事中医药防治骨与关节疾病研究（本文贡献与第一作者等同）
  王拥军，教授、主任医师，博士生导师；E-mail：yjwang8888@126.com
- 基金信息：
  
  国家重点研发计划项目(2018YFC1704302);国家自然科学基金资助项目(81973876;81929004;81730107);教育部创新团队发展计划项目(IRT1270);科技部重点领域创新团队项目(2015RA4002);上海市进一步加快中医药事业发展三年行动计划项目(ZY（2018-2020）-CCCX-3003);上海市卫计委科研基金资助项目(20174Y0192);上海中医药大学研究生创新创业能力培养项目(Y2019010)
- DOI：10.16306/j.1008-861x.2021.04.012
  中图分类号：
扫描看全文
程韶, 陶渝仁, 舒冰, 等. 化瘀补肾方对去卵巢小鼠骨折愈合及氧化应激的影响[J]. 上海中医药大学学报, 2021,35(4):79-88.

CHENG Shao, TAO Yuren, SHU Bing, et al. Effects of Huayu Bushen Formula on fracture healing and oxidative stress in ovariectomized mice[J]. Academic Journal of Shanghai University of Traditional Chinese Medicine, 2021,35(4):79-88.
程韶, 陶渝仁, 舒冰, 等. 化瘀补肾方对去卵巢小鼠骨折愈合及氧化应激的影响[J]. 上海中医药大学学报, 2021,35(4):79-88. DOI： 10.16306/j.1008-861x.2021.04.012.

CHENG Shao, TAO Yuren, SHU Bing, et al. Effects of Huayu Bushen Formula on fracture healing and oxidative stress in ovariectomized mice[J]. Academic Journal of Shanghai University of Traditional Chinese Medicine, 2021,35(4):79-88. DOI： 10.16306/j.1008-861x.2021.04.012.

摘要

目的：,2,观察化瘀补肾方对去卵巢小鼠骨折愈合及氧化应激的影响。,方法：,2,3月龄雌性C57BL/6小鼠随机分为假手术组、模型组、化瘀补肾方组，每组10只。模型组和化瘀补肾方组小鼠行双侧卵巢切除术（OVX），假手术组仅切除卵巢周围部分脂肪组织。术后3个月，各组小鼠均建立左侧胫骨骨折复合髓内固定模型。造模后，化瘀补肾方组小鼠灌胃给予7.33 g/kg化瘀补肾方药液，另两组小鼠灌胃给予等体积0.9%NaCl溶液，连续14 d。末次给药后，取血，分离胫骨组织。X线及Micro-CT扫描观察骨折愈合情况；阿尔新蓝/橙黄G染色后镜下观察骨痂组织形态学变化。试剂盒测定血清超氧化物歧化酶（SOD）、丙二醛（MDA）水平；免疫组化检测骨痂组织Ⅰ型胶原蛋白（CollagenⅠ）表达；免疫荧光检测骨痂组织Runt相关转录因子2（Runx2）、SOD表达；试剂盒检测骨痂组织MDA含量。TUNEL法检测骨痂组织细胞凋亡。,结果：,2,①胫骨X线及Micro-CT检测显示，模型组小鼠骨痂稀疏，骨折线清晰；化瘀补肾方组小鼠骨痂较致密，骨折线略模糊。②阿尔新蓝/橙黄G染色显示，模型组小鼠骨痂组织中软骨基质较多，骨小梁稀疏；化瘀补肾方组骨痂组织中软骨基质较少，骨小梁致密。③模型组骨痂组织Runx2、CollagenⅠ的表达较假手术组显著降低（,P,<,0.01），化瘀补肾方组骨痂组织Runx2、CollagenⅠ的表达较模型组明显升高（,P,<,0.05）。④化瘀补肾方组骨痂组织细胞凋亡较模型组明显减少（,P,<,0.01）。⑤与假手术组相比，模型组小鼠血清及骨痂组织SOD水平显著降低（,P,<,0.01），MDA含量显著升高（,P,<,0.01）；与模型组相比，化瘀补肾方组小鼠血清及骨痂组织SOD水平明显升高（,P,<,0.05），MDA含量明显下降（,P,<,0.01）。,结论：,2,化瘀补肾方能够促进去卵巢小鼠的骨折愈合，其作用机制可能与提高血清及局部骨痂组织的抗氧化酶水平、减轻氧化应激状态、抑制细胞凋亡有关。

Abstract

Objective:,2,To observe the effects of Huayu Bushen Formula on fracture healing and oxidative stress in ovariectomized mice.,Methods:,2,Three months old female C57BL/6 mice were randomly divided into the sham operation group, model group and Huayu Bushen Formula group, 10 mice in each group.Bilateral ovariectomy(OVX) was performed in the model group and Huayu Bushen Formula group, while only part of the adipose tissue around the ovary was removed in the sham operation group.Three months after operation, the model of left tibial fracture combined with intramedullary fixation was established in all groups.After modeling, mice in the Huayu Bushen Formula group were treated with decoction of Huayu Bushen Formula at dose of 7.33 g/kg by intragastric administration for 14 days, while mice in the sham operation group and model group were treated with 0.9% NaCl solution at equal volume.After last administration, blood was taken and tibia tissue was separated.The fracture healing was observed by X-ray and Micro-CT scanning.After alcian blue/orange G staining, the histomorphological changes of callus were observed under microscope.The serum levels of superoxide dismutase(SOD) and malondialdehyde(MDA) were measured by kits.The expression of CollagenⅠ in callus was detected by immunohistochemistry.The expressions of Runt-related transcription factor 2(Runx2) and SOD in callus were detected by immunofluorescence.The content of MDA in callus was detected by kit.The cell apoptosis in callus was detected by TUNEL assay.,Results:,2,①X-ray and Micro-CT examination of tibia showed that, the calluses were few and scattered and the fracture line was clear in the model group, while the calluses were dense and the fracture line was slightly blurred in the Huayu Bushen Formula group.②Alcian blue/orange G staining showed that, there were more cartilage matrix and less bone trabecula in callus of the model group, but less cartilage matrix and dense bone trabecular were observed in callus of the Huayu Bushen Formula group.③The expressions of Runx2 and CollagenⅠ in callus of the model group were significantly lower than those of the sham operation group(,P,<,0.01), and the expressions of Runx2 and CollagenⅠ in callus of the Huayu Bushen Formula group were significantly higher than those of the model group(,P,<,0.05) .④The cell apoptosis of callus tissue in the Huayu Bushen Formula group was significantly decreased compared with that in the model group(,P,<,0.01) .⑤Compared with the sham operation group, the SOD level in serum and callus tissue of the model group was significantly decreased(,P,<,0.01), and the MDA content was significantly increased(,P,<,0.01) .Compared with the model group, the SOD level in serum and callus tissue of the Huayu Bushen Formula group was significantly increased(,P,<,0.05), and the MDA content was significantly decreased(,P,<,0.01) .,Conclusion:,2,Huayu Bushen Formula can promote the fracture healing in ovariectomized mice, and its mechanisms may be related to increasing the levels of antioxidant enzymes in serum and local callus, alleviating oxidative stress and inhibiting cell apoptosis.

关键词

化瘀补肾方骨质疏松性骨折氧化应激小鼠

Keywords

Huayu Bushen Formulaosteoportic fractureoxidative stressmouse

references

中华医学会骨质疏松和骨矿盐疾病分会.原发性骨质疏松症诊疗指南（2017）[J].中国骨质疏松杂志，2019，25(3)：281-309.

HU Z B，WEI B，WU S K，et al. Changes in bone mineral density and bone metabolic indexes in ankylosing spondylitis mouse model complicated with osteoporosis[J]. Exp Ther Med，2018，16(2)：811-815.

LI G，ZHANG L，WANG L，et al. Dual modulation of bone formation and resorption with zoledronic acid-loaded biodegradable magnesium alloy implants improves osteoporotic fracture healing：An in vitro and in vivo study[J]. Acta Biomater，2018，65：486-500.

RUSSOW G，JAHN D，APPELT J，et al. Anabolic Therapies in Osteoporosis and Bone Regeneration[J]. Int J Mol Sci，2018，20(1)：83.

YU F，XIA W. The epidemiology of osteoporosis，associated fragility fractures，and management gap in China[J]. Arch Osteoporos，2019，14(1)：32.

SEVAL Y，EMRE K，ERHAN Y，et al. Effect of acupuncture therapy on fracture healing in rats with femur fractures[J]. J Tradit Chin Med，2020，40(2)：275-283.

YELER H，TAHTABAS F，CANDAN F. Investigation of oxidative stress during fracture healing in the rats[J]. Cell Biochem Funct，2005，23(2)：137-139.

YALIN S，SAGÍR O，COMELEKOGLU U，et al. Strontium ranelate treatment improves oxidative damage in osteoporotic rat model[J]. Pharmacol Rep，2012，64(2)：396-402.

ZHANG Y B，ZHONG Z M，HOU G，et al. Involvement of oxidative stress in age-related bone loss[J]. J Surg Res，2011，169(1)：e37-e42.

KUBO Y，WRUCK C J，FRAGOULIS A，et al. Role of Nrf2 in Fracture Healing：Clinical Aspects of Oxidative Stress[J]. Calcif Tissue Int，2019，105(4)：341-352.

LIU M，XU H，ZHANG L，et al. Salvianolic acid B inhibits myofibroblast transdifferentiation in experimental pulmonary fibrosis via the up-regulation of Nrf2[J]. Biochem Biophys Res Commun，2018，495(1)：325-331.

ZHANG Z，XU P，YU H，et al. Luteolin protects PC-12 cells from H2O2-induced injury by up-regulation of microRNA-21[J]. Biomed Pharmacother，2019，112：108698. doi：10.1016/j.biopha.2019.108698http://doi.org/10.1016/j.biopha.2019.108698.

DU W，AN Y，HE X，et al. Protection of Kaempferol on Oxidative Stress-Induced Retinal Pigment Epithelial Cell Damage[J]. Oxid Med Cell Longev，2018，2018：1610751. doi：10.1155/2018/1610751http://doi.org/10.1155/2018/1610751.

AHMED O M，FAHIM H I，AHMED H Y，et al. The Preventive Effects and the Mechanisms of Action of Navel Orange Peel Hydroethanolic Extract，Naringin，and Naringenin in N-Acetyl-p-aminophenol-Induced Liver Injury in Wistar Rats[J]. Oxid Med Cell Longev，2019，2019：2745352. doi：10.1155/2019/2745352http://doi.org/10.1155/2019/2745352.

ILYAS A，ODATSU T，SHAH A，et al. Amorphous Silica：A New Antioxidant Role for Rapid Critical-Sized Bone Defect Healing[J]. Adv Healthc Mater，2016，5(17)：2199-2213.

CHEUNG W H，MICLAU T，CHOW S K，et al. Fracture healing in osteoporotic bone[J]. Injury，2016，47 Suppl 2：S21-S26.

BIGARELLA C L，LIANG R，GHAFFARI S. Stem cells and the impact of ROS signaling[J]. Development，2014，141(22)：4206-4218.

DENU R A，HEMATTI P. Effects of Oxidative Stress on Mesenchymal Stem Cell Biology[J]. Oxid Med Cell Longev，2016，2016：2989076. doi：10.1155/2016/2989076http://doi.org/10.1155/2016/2989076.

ATASHI F，MODARRESSI A，PEPPER M S. The role of reactive oxygen species in mesenchymal stem cell adipogenic and osteogenic differentiation：a review[J]. Stem Cells Dev，2015，24(10)：1150-1163.

庞坚，王翔，陈元川，等.中药治疗骨质疏松性骨折的组方用药研究[J].时珍国医国药，2015，26(1)：238-239.

刘丹.骨质疏松症患者骨折特点分析及其与中医证型的关系[D].北京：中国中医科学院，2016.

金镇雄，舒冰，王乾，等.中医辨证分型治疗骨质疏松性骨折的研究进展[J].中国骨质疏松杂志，2020，26(12)：1843-1846.

KOMORI T. Molecular Mechanism of Runx2-Dependent Bone Development[J]. Mol Cells，2020，43(2)：168-175.

ZHANG X，AUBIN J E，INMAN R D. Molecular and cellular biology of new bone formation：insights into the ankylosis of ankylosing spondylitis[J]. Curr Opin Rheumatol，2003，15(4)：387-393.

SHEN M J，WANG G G，WANG Y Z，et al. Nell-1 Enhances Osteogenic Differentiation of Pre-Osteoblasts on Titanium Surfaces via the MAPK-ERK Signaling Pathway[J]. Cell Physiol Biochem，2018，50(4)：1522-1534.

LI X F，XU H，ZHAO Y J，et al. Icariin Augments Bone Formation and Reverses the Phenotypes of Osteoprotegerin-Deficient Mice through the Activation of Wnt/β-Catenin-BMP Signaling[J]. Evid Based Complement Alternat Med，2013，2013：652317. doi：10.1155/2013/652317http://doi.org/10.1155/2013/652317.

SHU B，ZHAO Y，WANG Y，et al. Oleanolic Acid Enhances Mesenchymal Stromal Cell Osteogenic Potential by Inhibition of Notch Signaling[J]. Sci Rep，2017，7(1)：7002.

XIE B P，SHI L Y，LI J P，et al. Oleanolic acid inhibits RANKL-induced osteoclastogenesis via ERα/miR-503/RANK signaling pathway in RAW264.7 cells[J]. Biomed Pharmacother，2019，117：109045. doi：10.1016/j.biopha.2019.109045http://doi.org/10.1016/j.biopha.2019.109045.

CAO S，DONG X L，HO M X，et al. Oleanolic Acid Exerts Osteoprotective Effects and Modulates Vitamin D Metabolism[J]. Nutrients，2018，10(2)：247.

ZHANG Y，HAN B，WEI Y，et al. Icariin Promotes Fracture Healing in Ovariectomized Rats[J]. Med Sci Monit，2020，26：e924554. doi：10.12659/MSM.924554http://doi.org/10.12659/MSM.924554.

席庆菊.墨旱莲的化学成分、药理作用、加工炮制及临床应用研究进展[J].中国处方药，2018，16(8)：15-17.

洪雅丹，张凌风，潘娉娉，等.HPLC法同时测定淫羊藿-骨碎补药对中7种成分[J].中成药，2019，41(6)：1316-1321.

WANG X，LIU R，ZHANG W，et al. Oleanolic acid improves hepatic insulin resistance via antioxidant，hypolipidemic and anti-inflammatory effects[J]. Mol Cell Endocrinol，2013，376(1-2)：70-80.

JING X，DU T，CHEN K，et al. Icariin protects against iron overload-induced bone loss via suppressing oxidative stress[J]. J Cell Physiol，2019，234(7)：10123-10137.

浏览量

323

下载量

CSCD

CNKI被引量

文章被引用时，请邮件提醒。

提交

工具集

关联资源

当飞利肝宁胶囊调控肝脏氧化应激改善非酒精性脂肪性肝病小鼠的机制研究

Mst1减轻小鼠非酒精性脂肪性肝炎的机制研究

虎杖苷通过调控Keap1/Nrf2/HO⁃1抑制H₂O₂诱导的SH⁃SY5Y细胞氧化应激和线粒体损伤

中医药通过调控氧化应激通路治疗糖尿病肾病研究进展

芪龙祛瘀合剂对缺血性脑卒中的保护作用及其初步机制探究