右归丸治疗骨质疏松性骨缺损的机制研究
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摘要
目的:探讨右归丸介导β-链蛋白(β-catenin)信号通路调控骨髓基质干细胞(BMSCs)治疗骨质疏松性骨缺损的机制。方法:进行小鼠骨髓间充质干细胞分离和体外扩增,鉴定其生物学活性。选择6月龄小鼠,建立骨质疏松性骨缺损模型,并给予右归丸[(18.9g/(kg·d)]进行治疗6周。采用酶联免疫吸附法(Elisa)检测小鼠血清中碱性磷酸酶(ALP)、抗酒石酸酸性磷酸酶(TRAP)、血清骨钙素(BGP)和雌二醇(E2);采用免疫组织化学法检测β-Catenin表达;采用实时PCR法检测骨钙素(Osteocalcin)、骨桥蛋白(Osteopontin)、成骨细胞特异性转录因子(Osterix)、骨细胞的特异转录因子(Runx2/Cbfa1)和β-Catenin mRNA表达。结果:与模型组相比,右归丸组和右归丸+BMSCs组小鼠β-Catenin含量显著升高;ALP和TRAP含量显著降低,BGP和E2含量显著升高;Osteopontin含量显著降低,Osterix,BMP-2,Runx2/Cbfa1和β-Catenin含量显著升高,差异有统计学意义(P<0.05)。与右归丸组相比,右归丸+BMSCs组β-Catenin含量显著升高,ALP和TRAP含量显著降低,BGP和E2含量显著升高;Osteopontin含量显著降低,Osterix,BMP-2,Runx2/Cbfa1和β-Catenin含量显著升高,差异有统计学意义(P<0.05)。结论:β-Catenin信号途径在骨质疏松性骨缺损骨愈合过程中具有重要的作用。右归丸是通过激活β-Catenin促进BMSCs成骨分化,从而促进骨质疏松性骨缺损骨愈合。
Objective:To investigate the mechanism of Yougui pill mediatedβ-catenin signaling pathway in the treatment of osteoporotic bone defect induced by BMSCs.Methods:Mouse bone marrow mesenchymal stem cells were isolated and expanded in vitro to identify their biological activity.Six-month-old mice were selected to establish an osteoporotic bone defect model and treated with Yougui pill[18.9 g/(kg·d)]for 6 weeks.Serum alkaline phosphatase(ALP),tartrate-resistant acid phosphatase(TRAP),serum osteocalcin(BGP)and estradiol(E2)were detected by enzyme-linked immunosorbent assay(Elisa).The expression of β-catenin was detected by immunohistochemistry.The expression of osteocalcin,osteopontin,osterix,osteoclast-specific transcription factor(Runx2/Cbfa1)and β-catenin mRNA were detected by real-time PCR.Results:Compared with the model group,the content of β-catenin in two group was significantly increased,and ALP and tartrate-resistant acid phosphatase(TRAP)were significantly decreased,while serum BGP and E2 levels were significantly increased.The contents of osteopontin,osterix,BMP-2,Runx2/Cbfa1 and β-catenin were significantly decreased,with statistical significance(P<0.05).Compared with Yougui pill group,the content ofβ-Catenin in Yougui pill and BMSCs group was significantly increased,the contents of ALP and TRAP were significantly decreased,and BGP and E2 contents were significantly increased.The contents of osteopontin,osterix,BMP-2,Runx2/Cbfa1 and β-catenin were significantly decreased,with statistical significance(P<0.05).Conclusion:The β-catenin signaling pathway plays an important role in the process of osteogenesis of osteoporotic bone defects.Yougui pill promotes osteogenic differentiation of BMSCs by activatingβ-catenin and promotes bone healing of osteoporotic bone defects.
Objective:To investigate the mechanism of Yougui pill mediatedβ-catenin signaling pathway in the treatment of osteoporotic bone defect induced by BMSCs.Methods:Mouse bone marrow mesenchymal stem cells were isolated and expanded in vitro to identify their biological activity.Six-month-old mice were selected to establish an osteoporotic bone defect model and treated with Yougui pill[18.9 g/(kg·d)]for 6 weeks.Serum alkaline phosphatase(ALP),tartrate-resistant acid phosphatase(TRAP),serum osteocalcin(BGP)and estradiol(E2)were detected by enzyme-linked immunosorbent assay(Elisa).The expression of β-catenin was detected by immunohistochemistry.The expression of osteocalcin,osteopontin,osterix,osteoclast-specific transcription factor(Runx2/Cbfa1)and β-catenin mRNA were detected by real-time PCR.Results:Compared with the model group,the content of β-catenin in two group was significantly increased,and ALP and tartrate-resistant acid phosphatase(TRAP)were significantly decreased,while serum BGP and E2 levels were significantly increased.The contents of osteopontin,osterix,BMP-2,Runx2/Cbfa1 and β-catenin were significantly decreased,with statistical significance(P<0.05).Compared with Yougui pill group,the content ofβ-Catenin in Yougui pill and BMSCs group was significantly increased,the contents of ALP and TRAP were significantly decreased,and BGP and E2 contents were significantly increased.The contents of osteopontin,osterix,BMP-2,Runx2/Cbfa1 and β-catenin were significantly decreased,with statistical significance(P<0.05).Conclusion:The β-catenin signaling pathway plays an important role in the process of osteogenesis of osteoporotic bone defects.Yougui pill promotes osteogenic differentiation of BMSCs by activatingβ-catenin and promotes bone healing of osteoporotic bone defects.
引文
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[15]张哲,徐秀娟,刘欣,等.老鹳草素通过Wnt/β-catenin信号通路影响小鼠骨髓基质干细胞的增殖和成骨分化[J].中华中医药学刊,2017,35(1):215-218.
[16]韦国雨,陈清雄,唐永亮,等.基于Wnt-β-catenin信号调控BMP-2表达探讨当归四逆汤防治类风湿性关节炎临床研究[J].中华中医药学刊,2017,35(1):243-246.
[17]刘兴振,范洁,李甲,等.骨关节炎关节软骨自然退变过程中Dvl2及β-catenin表达变化[J].中国现代应用药学,2016,33(11):1355-1358.
[2]卢建华,卢烨超,金红婷,等.红曲对去卵巢大鼠骨质疏松模型血清ALP,TRAP及骨组织TNF-α,RANK表达的影响[J].中国中医骨伤科杂志,2017,25(9):5-8.
[3]姚洪春,吴世军,刘正,等.骨代谢标志物的测定在老年骨质疏松脊柱脆性骨折中的意义[J].检验医学与临床,2017,14(20):3079-3081.
[4]刘新梅,常红.经皮椎体成形术治疗骨质疏松椎体压缩性骨折两种护理方法的随机对照研究[J].检验医学与临床,2016,13(16):2261-2263.
[5]黄成校,余化龙,高超,等.骨髓间充质干细胞复合Ⅰ型胶原修饰的聚乳酸聚乙醇酸对骨质疏松大鼠骨缺损的影响[J].安徽医药,2017,21(4):618-622.
[6]黄馗,罗道文,王雷,等.大鼠骨质疏松伴颅骨极量缺损模型的建立[J].中国组织工程研究,2017,21(8):1236-1240.
[7]颜学亮,曹奇,陈小明.小剂量骨水泥经皮椎体成形术治疗老年性骨质疏松症多节段跳跃性脊柱压缩性骨折的疗效分析[J].中南医学科学杂志,2016,44(4):368-370.
[8]孟玥,任艳玲,孙月娇,等.左归丸、右归丸及其拆方对去卵巢骨质疏松症模型大鼠肾脏碱性磷酸酶、骨钙素表达的影响[J].中医杂志,2016,57(5):423-427.
[9]阳松威,孙晓峰,贺又舜,等.左归丸对化疗致卵巢早衰小鼠卵巢功能的影响[J].中成药,2016,38(4):717-722.
[10]孙千惠,任艳玲,吴琼,等.左、右归丸对去卵巢大鼠BMSCs成骨、成脂分化后Caspase-3/Bcl-2的影响[J].中成药,2017,39(10):2004-2008.
[11]李凌汉,麦培根,陈宝红.右归丸对原发性骨质疏松症患者骨密度、骨代谢及脆性骨折发生率的影响[J].现代中西医结合杂志,2017,26(19):2088-2090.
[12]雷宁波,彭冉东,邓强,等.依降钙素联合损伤胶囊对骨质疏松性椎体压缩骨折PKP术后早期临床疗效观察[J].中国中医骨伤科杂志,2017,25(5):9-12.
[13]张添昊,任艳玲,艾思羽,等.左归丸和右归丸对去卵巢大鼠股骨骨髓PPARγ、C/EBPβ、C/EBPα蛋白表达的影响[J].中医杂志,2017,58(6):511-515.
[14]王君化.HER2和β-catenin在食管癌中表达的临床病理意义[J].中南医学科学杂志,2016,44(5):531-535.
[15]张哲,徐秀娟,刘欣,等.老鹳草素通过Wnt/β-catenin信号通路影响小鼠骨髓基质干细胞的增殖和成骨分化[J].中华中医药学刊,2017,35(1):215-218.
[16]韦国雨,陈清雄,唐永亮,等.基于Wnt-β-catenin信号调控BMP-2表达探讨当归四逆汤防治类风湿性关节炎临床研究[J].中华中医药学刊,2017,35(1):243-246.
[17]刘兴振,范洁,李甲,等.骨关节炎关节软骨自然退变过程中Dvl2及β-catenin表达变化[J].中国现代应用药学,2016,33(11):1355-1358.