红芪中5种黄酮类成分对大鼠骨髓间充质干细胞和成骨细胞成骨分化的影响
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摘要
目的研究红芪中毛蕊异黄酮、芒柄花素、芒柄花苷、异甘草素和美迪紫檀素5种黄酮类成分对大鼠骨髓间充质干细胞(rat bone marrow stromal cells,rBMSCs)和颅骨成骨细胞(rat calvarial osteoblasts,ROBs)成骨性分化的影响。方法贴壁分离法培养rBMSCs;酶消解法分离培养ROBs;MTT法检测rBMSCs和ROBs细胞的增殖情况;试剂盒检测rBMSCs和ROBs的碱性磷酸酶(ALP)活性和Ca~(2+)含量;茜素红染色法检测矿化结节形成。结果 5种成分均能不同程度促进rBMSCs和ROBs的增殖,提高ALP活性,增加Ca~(2+)含量,增加钙化结节面积和数量(P<0.05)。其中毛蕊异黄酮促进rBMSCs的成骨分化作用最佳,美迪紫檀素促进ROBs的成骨分化作用最佳,毛蕊异黄酮次之。结论这5种黄酮类成分对rBMSCs和ROBs的成骨功能均有一定程度地改善作用,而毛蕊异黄酮对2种细胞表现的成骨活性均较佳,可发展作为良好的骨诱导活性因子。
Objective To investigate the effects of five kinds of flavonoids(calycosin, formononetin, ononin, isoliquiritigenin, and medicarpin) from Hedysari Radix on promoting osteogenic differentiation of rat bone marrow stromal cells(rBMSCs) and rat calvarial osteoblasts(ROBs). Methods rBMSCs were isolated according to plastic adherence. ROBs were isolated by enzyme digestion method. The proliferation of rBMSCs and ROBs were detected by MTT assay. ALP activity and calcium content of rBMSCs and ROBs cells were detected by alkaline phosphatase kit and calcium kit. Mineralized nodule formation was detected by alizarin red staining. Results The five components could promote proliferation, increase ALP activity, increase calcium content, and increase the area and number of calcified nodules of rBMSCs and ROBs(P < 0.05). Among them, calycosin had the best effect on promoting the osteogenic differentiation of rBMSCs, and medicarpin promoted the osteogenic differentiation of ROBs with the best effect, followed by calycosin. Conclusion Five flavonoids promoted the improvement of osteogenic function, while calycosin has better osteogenic activity on rBMSCs and ROBs and can be used as an excellent osteoinductive factor.
Objective To investigate the effects of five kinds of flavonoids(calycosin, formononetin, ononin, isoliquiritigenin, and medicarpin) from Hedysari Radix on promoting osteogenic differentiation of rat bone marrow stromal cells(rBMSCs) and rat calvarial osteoblasts(ROBs). Methods rBMSCs were isolated according to plastic adherence. ROBs were isolated by enzyme digestion method. The proliferation of rBMSCs and ROBs were detected by MTT assay. ALP activity and calcium content of rBMSCs and ROBs cells were detected by alkaline phosphatase kit and calcium kit. Mineralized nodule formation was detected by alizarin red staining. Results The five components could promote proliferation, increase ALP activity, increase calcium content, and increase the area and number of calcified nodules of rBMSCs and ROBs(P < 0.05). Among them, calycosin had the best effect on promoting the osteogenic differentiation of rBMSCs, and medicarpin promoted the osteogenic differentiation of ROBs with the best effect, followed by calycosin. Conclusion Five flavonoids promoted the improvement of osteogenic function, while calycosin has better osteogenic activity on rBMSCs and ROBs and can be used as an excellent osteoinductive factor.
引文
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[14]郭晓宇,李唯,陈克明,等.淫羊藿苷通过PI3K/AKT-e NOS信号途径促进大鼠骨髓基质细胞的成骨性分化[J].中国药理学通报,2013,29(7):966-970.
[15]方清清,李志忠,周建,等.低频脉冲电磁场通过cAMP/PKA信号通路促进成骨细胞分化的研究[J].南方医科大学学报,2016,36(11):1508-1513.
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[17]车晓明,陈亮,顾勇,等.茶多酚治疗骨质疏松症的研究进展[J].中国骨质疏松杂志,2015,21(2):235-240.
[18]农梦妮,曾高峰,宗少晖,等.黄精多糖调控骨髓间充质干细胞向成骨细胞分化[J].中国组织工程研究,2016,20(15):2133-2139.
[19]Calliste C A,Le B J,Trouillas P,et al.Chalcones:structural requirements for antioxidant,estrogenic and antiproliferative activities[J].Anticancer Res,2001,21(6A):3949-3956.
[20]赵雪巍,刘培玉,刘丹,等.黄酮类化合物的构效关系研究进展[J].中草药,2015,46(21):3264-3271.
[21]Srivastava A,Singh H,Mishra R,et al.Structural insights,protein-ligand interactions and spectroscopic characterization of isoformononetin[J].J Mol Struct,2017,1133:479-491.
[22]陈宇,薛志远,师志强,等.红芪活性组分抗骨质疏松作用的谱效关系研究[J].中草药,2018,49(1):160-166.
[2]王迎彬,王金花.脂联素在骨质疏松性骨代谢疾病中的研究进展[J].解剖学杂志,2016,39(6):746-748.
[3]Liu Y,Liu J P,Xia Y.Chinese herbal medicines for treating osteoporosis[J].Cochrane Database Syst Rev,2014,3(3):CD005467.
[4]薛志远,赵良功,师志强,等.红芪和黄芪对去卵巢致大鼠骨质疏松防治作用的研究[J].现代中药研究与实践,2017,31(6):21-24.
[5]Zhao L,Zhao H,Sheng X,et al.Structural characterization and stimulating effect on osteoblast differentiation of a purified heteropolysaccharide isolated from Hedysarum polybotrys[J].Carbohydr Polym,2014,111(1):714-721.
[6]蒋俊,崔莉,孙娥,等.基于淫羊藿黄酮类化合物的体内代谢阐述其抗骨质疏松药效物质基础[J].中草药,2014,45(5):721-729.
[7]Goel A,Raghuvanshi A,Kumar A,et al.9-Demethoxymedicarpin promotes peak bone mass achievement and has bone conserving effect in ovariectomized mice:Positively regulates osteoblast functions and suppresses osteoclastogenesis[J].Mol Cell Endocrinol,2015,411:155-166.
[8]苏艳杰,陈亚辉,崔燎.植物黄酮抗骨质疏松作用研究进展[J].中国骨质疏松杂志,2014,20(5):562-568.
[9]丁小刚,覃勇,鄂建设,等.骨碎补总黄酮对老年性骨质疏松症患者血清骨钙素水平及骨密度影响[J].中国骨质疏松杂志,2013,19(5):519-521.
[10]王庆丰,王孝辉,张彩丽,等.骨碎补总黄酮促进大鼠诱导膜中VEGF、BMP-2表达的实验研究[J].药物评价研究,2018,41(7):1220-1223.
[11]宋双红,余倩,王喆之,等.骨碎补防治骨质疏松研究概况[J].生命的化学,2015,35(1):73-80.
[12]丁淑琴,杨风琴,张玮,等.山柰酚治疗去卵巢大鼠骨质疏松的实验研究[J].中国新药杂志,2014,23(24):2925-2929.
[13]柳小亚,李继平,陈心悦,等.HPLC同时测定红芪中8个活性成分的含量及聚类分析[J].药学学报,2016,51(5):786-791.
[14]郭晓宇,李唯,陈克明,等.淫羊藿苷通过PI3K/AKT-e NOS信号途径促进大鼠骨髓基质细胞的成骨性分化[J].中国药理学通报,2013,29(7):966-970.
[15]方清清,李志忠,周建,等.低频脉冲电磁场通过cAMP/PKA信号通路促进成骨细胞分化的研究[J].南方医科大学学报,2016,36(11):1508-1513.
[16]Xie Y F,Shi W G,Zhou J,et al.Pulsed electromagnetic fields stimulate osteogenic differentiation and maturation of osteoblasts by upregulating the expression of BMPRII localized at the base of primary cilium[J].Bone,2016,93:22-32.
[17]车晓明,陈亮,顾勇,等.茶多酚治疗骨质疏松症的研究进展[J].中国骨质疏松杂志,2015,21(2):235-240.
[18]农梦妮,曾高峰,宗少晖,等.黄精多糖调控骨髓间充质干细胞向成骨细胞分化[J].中国组织工程研究,2016,20(15):2133-2139.
[19]Calliste C A,Le B J,Trouillas P,et al.Chalcones:structural requirements for antioxidant,estrogenic and antiproliferative activities[J].Anticancer Res,2001,21(6A):3949-3956.
[20]赵雪巍,刘培玉,刘丹,等.黄酮类化合物的构效关系研究进展[J].中草药,2015,46(21):3264-3271.
[21]Srivastava A,Singh H,Mishra R,et al.Structural insights,protein-ligand interactions and spectroscopic characterization of isoformononetin[J].J Mol Struct,2017,1133:479-491.
[22]陈宇,薛志远,师志强,等.红芪活性组分抗骨质疏松作用的谱效关系研究[J].中草药,2018,49(1):160-166.