黄芪总黄酮对RAW264.7细胞细胞因子和介质分泌水平的影响
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摘要
试验采用传统中药黄芪的主要成分黄芪总黄酮为研究对象,探讨了黄芪总黄酮的体外免疫作用。以RAW264.7细胞为模型,通过细胞毒性试验拟定黄芪总黄酮的试验剂量,分别设立药物低、中、高剂量组以及空白组,通过ELISA法测定细胞因子IL-6、IL-1β、IFN-γ、TNF-α和介质PGE2的含量,Griess法测定介质NO分泌水平,Western blot法分析iNOS和COX-2蛋白含量变化。结果表明:黄芪总黄酮剂量依赖性的促进了正常条件下RAW264.7细胞上清液中细胞因子IL-6、IL-1β、IFN-γ、TNF-α和细胞介质NO、PGE2的分泌,上调了iNOS和COX-2蛋白的表达,表明黄芪总黄酮具有体外免疫增强作用。
In this study,we investigated the immunological effects of total flavonoids of astragalus(TFA)in vitro which is the main components of traditional Chinese medicine Astragalus.The experimental dose of TFA was established through cytotoxicity test by using the RAW264.7 cells.TFA groups(low,medium and high doses)and blank group were set up,respectively.The levels of cytokines IL-6,IL-1β,IFN-γ,TNF-α,mediator PGE2 were detected by ELISA kits,NO secretion level was detected by Griess reagent and COX-2,iNOS protein expression levels were determined by Western blot.The results showed that the levels of IL-6,IL-1β,IFN-γ,TNF-α,NO and PGE2 were significantly increased,and the expression levels of iNOS and COX-2 proteins were up-regulated in TFA treated groups by dose dependent.It is shown that TFA could enhance immunity in vitro.
In this study,we investigated the immunological effects of total flavonoids of astragalus(TFA)in vitro which is the main components of traditional Chinese medicine Astragalus.The experimental dose of TFA was established through cytotoxicity test by using the RAW264.7 cells.TFA groups(low,medium and high doses)and blank group were set up,respectively.The levels of cytokines IL-6,IL-1β,IFN-γ,TNF-α,mediator PGE2 were detected by ELISA kits,NO secretion level was detected by Griess reagent and COX-2,iNOS protein expression levels were determined by Western blot.The results showed that the levels of IL-6,IL-1β,IFN-γ,TNF-α,NO and PGE2 were significantly increased,and the expression levels of iNOS and COX-2 proteins were up-regulated in TFA treated groups by dose dependent.It is shown that TFA could enhance immunity in vitro.
引文
[1]王健楠,阮洪生,崔玉东,等.身痛逐瘀汤通过调P38MAPK信号通路抑制LPS诱导的RAW264.7巨噬细胞一氧化氮的分泌[J].黑龙江八一农垦大学学报,2015,27(03):75-78,87.
[2]丛媛媛,热米拉·米吉提,帕丽达·阿不力孜,等.胀果甘草多糖对RAW264.7巨噬细胞免疫功能的影响[J].中华中医药学刊,2018,36(05):1043-1047.
[3]汪小莉,刘晓,夏春燕,等.防己黄芪汤药理作用及各单味药化学成分研究进展[J].中草药,2016,47(19):3527-3534.
[4]胡明.黄芪在处方中配伍临床应用的分析[J].世界最新医学信息文摘,2016,16(87):213.
[5]王静.加减黄芪桂枝五物汤对口腔黏膜白斑免疫应答的影响[J].新中医,2018,50(06):160-162.
[6]田永利,许志宇,葛林,等.总黄酮类化合物含量测定方法和药理作用研究进展[J].河北医药,2010,32(15):2094-2096.
[7]Guo Z,Xu H Y,Xu L,et al.In vivo and in vitro immunomodulatory and anti-inflammatory effects of total flavonoids of astragalus[J].African Journal of Traditional Complementary and Alternative Medicines,2016,13(4):60-73.
[8]刘馨宇,徐璐,王娜,等.黄芪总黄酮对大鼠佐剂性关节炎病理损伤的影响[J].动物医学进展,2017,38(10):73-77.
[9]袁红,张淑芳,贾绍辉,等.黄芪生物活性及其在保健食品中的应用研究进展[J].食品科学,2014,35(15):330-334.
[10]金鹰,秀荣,呼格吉乐图.炎症的发展规律及鉴别诊断[J].动物保健,2006(11):54-55.
[11]付丽佳,王玉华,徐莲琴.儿科药学服务[J].中国医药指南,2013,11(24):383-385.
[12]李冀,于雪,马育轩,等.中药及其有效成分抗炎机制的研究进展[J].中医药学报,2010,38(02):134-137.
[13]姜宁,宋新波.蒲公英的药理研究进展[J].中国中医药杂志,2008,6(12):19-23.
[14]张会宗,李成森,贾天柱.中药蒲公英临床应用[J].时珍国医国药,2001,12(01):84-85.
[15]刘轩,魏育林,李波,等.人参皂甙对脂多糖诱导小鼠肺泡巨噬细胞分泌肿瘤坏死因子的作用及机制探讨[J].中日友好医院学报,2001,15(02):83-86.
[16]贺拥军,康龙丽,王英,等.类风湿性关节炎病人血清及关节滑液中一氧化氮、超氧化物歧化酶、丙二醛含量变化观察[J].西藏医药杂志,2005,26(03):43-45.
[17]李艳云,史秋梅,王晓珊,等.紫锥菊多糖对内毒素血症小鼠炎症的保护作用[J].河北科技师范学院学报,2015,27(03):8-11.
[18]宗炎,阚童童,潘求真,等.绵羊IL-6启动子的克隆及pIL6-TLR4载体构建[J].黑龙江八一农垦大学学报,2017,29(1):45-48.
[19]李晓婷,宋佰芬,崔玉东.鸡白介素18与鸡γ干扰素基因融合表达载体构建[J].黑龙江八一农垦大学学报,2017,29(1):49-53.
[20]田中秋,邓立普.TNF-a,IL-6在全身炎症反应综合征表达的研究进展[J].蛇志,2008,20(04):275-278.
[21]潘灵辉.细胞因子平衡在炎症反应中作用的研究进展[J].医学综述,2005,11(09):775-777.
[22]江泽波,胡金萍,温晓文,等.猪苓多糖对巨噬细胞RAW264.7的双向免疫调节作用[J].免疫学杂志,2014,30(12):1033-1038.
[23]苏娣,张芸,戴竹青,等.毛蕊花糖对巨噬细胞RAW264.7的免疫调节作用[J].食品科学,2013,34(7):250-253.
[24]林红,胡格,伊鹏霏,等.黄芩苷,连翘酯苷对肺微血管内皮细胞分泌IFN-α,IFN-γ的影响[J].中国畜牧兽医,2009,36(06):30-34.
[25]闫姝含.多拉菌素的抗炎作用及其对相关信号转导通路的调控[D].长春:吉林大学,2013.
[26]陈榕芳,吴小南,陈洁.浒苔多糖粗提物调节巨噬细胞RAW264.7免疫功能研究[J].海峡预防医学杂志,2012,18(2):4-7.
[27]司天雷.DHA与槲皮素的抗炎作用及其对NFκB、MAPK信号通路影响的研究[D].武汉:华中农业大学,2015.
[28]刘光耀,王海燕,白彩虹,等.威灵仙不同提取部位对脂多糖诱导的RAW264.7细胞NO生成及iNOS和TNF-α表达的影响[J].三峡大学学报(自然科学版),2014,36(3):102-107.
[29]武小兰,佟强,胡玉荣.蒙药查格德日丸对偏头痛大鼠脑和血中NO和NOS的影响[J].内蒙古民族大学学报(自然科学版),2015,30(06):515-517.
[30]张彦燕,李佳川,张小金,等.鱼腥草素钠对人脐静脉内皮细胞NO合成及NOS活力的影响[J].西南民族大学学报(自然科学版),2014,40(5):711-714.
[31]赵恒光,罗福玲.粉防己碱对脂多糖诱导下RAW264.7细胞COX-2/PGE2、iNOS/NO表达的影响[J].中国药科大学学报,2011,42(02):141-144.
[32]杨光锐,管又飞.前列腺素E2合酶的研究进展[J].生理科学进展,2006,37(02):182-186.
[33]Murakami M,Naraba H,Tanioka T,et al.Regulation of prostaglandin E2biosynthesis by inducible membrane-associated prostaglandin E2synthase that acts in concert with cyclooxygenase-2[J].Journal of Biological Chemistry,2000,275(42):32783-32792.
[34]Jakobsson P J,Thorén S,Morgenstern R,et al.Identification of human prostaglandin E synthase:a microsomal,glutathione-dependent,inducible enzyme,constituting apotential novel drug target[J].Proceedings of the National Academy of Sciences,1999,96(13):7220-7225.
[35]程瑶,郭泽,王平,等.蒲公英甾醇对LPS诱导的RAW264.7细胞iNOS和COX-2表达的影响及其机制研究[J].免疫学杂志,2015,31(10):860-863.
[36]徐永明,高智,肖良,等.HIF-1α和COX-2在力学诱导大鼠终板软骨细胞退变模型中的表达及意义[J].赤峰学院学报(自然科学版),2017,33(13):23-25.
[2]丛媛媛,热米拉·米吉提,帕丽达·阿不力孜,等.胀果甘草多糖对RAW264.7巨噬细胞免疫功能的影响[J].中华中医药学刊,2018,36(05):1043-1047.
[3]汪小莉,刘晓,夏春燕,等.防己黄芪汤药理作用及各单味药化学成分研究进展[J].中草药,2016,47(19):3527-3534.
[4]胡明.黄芪在处方中配伍临床应用的分析[J].世界最新医学信息文摘,2016,16(87):213.
[5]王静.加减黄芪桂枝五物汤对口腔黏膜白斑免疫应答的影响[J].新中医,2018,50(06):160-162.
[6]田永利,许志宇,葛林,等.总黄酮类化合物含量测定方法和药理作用研究进展[J].河北医药,2010,32(15):2094-2096.
[7]Guo Z,Xu H Y,Xu L,et al.In vivo and in vitro immunomodulatory and anti-inflammatory effects of total flavonoids of astragalus[J].African Journal of Traditional Complementary and Alternative Medicines,2016,13(4):60-73.
[8]刘馨宇,徐璐,王娜,等.黄芪总黄酮对大鼠佐剂性关节炎病理损伤的影响[J].动物医学进展,2017,38(10):73-77.
[9]袁红,张淑芳,贾绍辉,等.黄芪生物活性及其在保健食品中的应用研究进展[J].食品科学,2014,35(15):330-334.
[10]金鹰,秀荣,呼格吉乐图.炎症的发展规律及鉴别诊断[J].动物保健,2006(11):54-55.
[11]付丽佳,王玉华,徐莲琴.儿科药学服务[J].中国医药指南,2013,11(24):383-385.
[12]李冀,于雪,马育轩,等.中药及其有效成分抗炎机制的研究进展[J].中医药学报,2010,38(02):134-137.
[13]姜宁,宋新波.蒲公英的药理研究进展[J].中国中医药杂志,2008,6(12):19-23.
[14]张会宗,李成森,贾天柱.中药蒲公英临床应用[J].时珍国医国药,2001,12(01):84-85.
[15]刘轩,魏育林,李波,等.人参皂甙对脂多糖诱导小鼠肺泡巨噬细胞分泌肿瘤坏死因子的作用及机制探讨[J].中日友好医院学报,2001,15(02):83-86.
[16]贺拥军,康龙丽,王英,等.类风湿性关节炎病人血清及关节滑液中一氧化氮、超氧化物歧化酶、丙二醛含量变化观察[J].西藏医药杂志,2005,26(03):43-45.
[17]李艳云,史秋梅,王晓珊,等.紫锥菊多糖对内毒素血症小鼠炎症的保护作用[J].河北科技师范学院学报,2015,27(03):8-11.
[18]宗炎,阚童童,潘求真,等.绵羊IL-6启动子的克隆及pIL6-TLR4载体构建[J].黑龙江八一农垦大学学报,2017,29(1):45-48.
[19]李晓婷,宋佰芬,崔玉东.鸡白介素18与鸡γ干扰素基因融合表达载体构建[J].黑龙江八一农垦大学学报,2017,29(1):49-53.
[20]田中秋,邓立普.TNF-a,IL-6在全身炎症反应综合征表达的研究进展[J].蛇志,2008,20(04):275-278.
[21]潘灵辉.细胞因子平衡在炎症反应中作用的研究进展[J].医学综述,2005,11(09):775-777.
[22]江泽波,胡金萍,温晓文,等.猪苓多糖对巨噬细胞RAW264.7的双向免疫调节作用[J].免疫学杂志,2014,30(12):1033-1038.
[23]苏娣,张芸,戴竹青,等.毛蕊花糖对巨噬细胞RAW264.7的免疫调节作用[J].食品科学,2013,34(7):250-253.
[24]林红,胡格,伊鹏霏,等.黄芩苷,连翘酯苷对肺微血管内皮细胞分泌IFN-α,IFN-γ的影响[J].中国畜牧兽医,2009,36(06):30-34.
[25]闫姝含.多拉菌素的抗炎作用及其对相关信号转导通路的调控[D].长春:吉林大学,2013.
[26]陈榕芳,吴小南,陈洁.浒苔多糖粗提物调节巨噬细胞RAW264.7免疫功能研究[J].海峡预防医学杂志,2012,18(2):4-7.
[27]司天雷.DHA与槲皮素的抗炎作用及其对NFκB、MAPK信号通路影响的研究[D].武汉:华中农业大学,2015.
[28]刘光耀,王海燕,白彩虹,等.威灵仙不同提取部位对脂多糖诱导的RAW264.7细胞NO生成及iNOS和TNF-α表达的影响[J].三峡大学学报(自然科学版),2014,36(3):102-107.
[29]武小兰,佟强,胡玉荣.蒙药查格德日丸对偏头痛大鼠脑和血中NO和NOS的影响[J].内蒙古民族大学学报(自然科学版),2015,30(06):515-517.
[30]张彦燕,李佳川,张小金,等.鱼腥草素钠对人脐静脉内皮细胞NO合成及NOS活力的影响[J].西南民族大学学报(自然科学版),2014,40(5):711-714.
[31]赵恒光,罗福玲.粉防己碱对脂多糖诱导下RAW264.7细胞COX-2/PGE2、iNOS/NO表达的影响[J].中国药科大学学报,2011,42(02):141-144.
[32]杨光锐,管又飞.前列腺素E2合酶的研究进展[J].生理科学进展,2006,37(02):182-186.
[33]Murakami M,Naraba H,Tanioka T,et al.Regulation of prostaglandin E2biosynthesis by inducible membrane-associated prostaglandin E2synthase that acts in concert with cyclooxygenase-2[J].Journal of Biological Chemistry,2000,275(42):32783-32792.
[34]Jakobsson P J,Thorén S,Morgenstern R,et al.Identification of human prostaglandin E synthase:a microsomal,glutathione-dependent,inducible enzyme,constituting apotential novel drug target[J].Proceedings of the National Academy of Sciences,1999,96(13):7220-7225.
[35]程瑶,郭泽,王平,等.蒲公英甾醇对LPS诱导的RAW264.7细胞iNOS和COX-2表达的影响及其机制研究[J].免疫学杂志,2015,31(10):860-863.
[36]徐永明,高智,肖良,等.HIF-1α和COX-2在力学诱导大鼠终板软骨细胞退变模型中的表达及意义[J].赤峰学院学报(自然科学版),2017,33(13):23-25.