复方龙脉宁对急性心肌梗死模型大鼠TLR4/MyD88/NF-κB p65信号通路的影响
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摘要
目的:探讨复方龙脉宁汤对盐酸异丙肾上腺素诱导的心肌梗死大鼠模型的保护作用及其对Toll样受体4(TLR4)/髓样分化因子(MyD88)/核转录因子-κB(NF-κB)信号通路的影响。方法:取雄性SD大鼠48只,随机分成6组:正常组,模型组,复方丹参滴丸组(0.072 9 g·kg~(-1)),复方龙脉宁低、中、高剂量组(0.36,0.71,1.43 g·kg~(-1))。采用背部皮下多点注射盐酸异丙肾上腺素的方法建立急性心肌梗死动物模型。采用苏木素-伊红(HE)染色观察大鼠心肌组织病理学变化;酶联免疫吸附测定(ELISA)检测大鼠血清中白细胞介素-1β(IL-1β),白细胞介素-6(IL-6),肿瘤坏死因子-α(TNF-α),单核细胞趋化蛋白-1(MCP-1),一氧化氮(NO)的含量;免疫组化法测定大鼠心肌组织中NF-κB激酶β抑制蛋白(IKKβ),NF-κB抑制蛋白α(IκBα)的表达水平;蛋白免疫印迹法(Western blot)检测大鼠心肌组织中TLR4,MyD88,NF-κB p65蛋白的表达。结果:与正常组比较,模型组大鼠心肌损伤明显,血清中IL-1β,IL-6,TNF-α,MCP-1,NO含量明显升高(P<0.05),心肌组织中IκBα蛋白表达水平明显降低(P<0.05),心肌组织中IKKβ,TLR4,MyD88,NF-κB p65蛋白表达水平明显升高(P<0.05);与模型组比较,复方龙脉宁低、中、高剂量组能明显改善大鼠心肌损伤,明显升高IκBα蛋白的表达(P<0.05),明显降低血清中IL-1β,IL-6,TNF-α,MCP-1,NO的活性及心肌组织中IKKβ,TLR4,MyD88,NF-κB p65蛋白的表达(P<0.05)。结论:复方龙脉宁对心肌梗死的保护作用,可能与调控TLR4/MyD88/NF-κB p65信号通路相关因子的表达有关。
Objective: To investigate the protective effect of compound Longmaining isoprenaline hydrochloride-induced myocardial infarction model and its effect on Toll-like receptor 4( TLR4)/myeloid differentiation factor 88( MyD88)/nuclear transcription factor-kappa B( NF-κB) signaling pathway. Method:Forty-eight male SD rats were randomly divided into 6 groups: normal group,model group,compound salvia miltiorrhiza drop pill group( 0. 072 9 g·kg~(-1)),and low,medium and high-dose compound Longmaining decoction groups( 0. 36,0. 71,1. 43 g·kg~(-1)). The acute myocardial infarction model was induced through subcutaneous injection with isoproterenol. The pathological changes of myocardial tissue were examined by hematoxylin-eosin( HE) staining. The levels of interleukin-1β( IL-1β),interleukin-6( IL-6),tumor necrosis factor-α( TNF-α),monocyte chemotaxis protein-1( MCP-1) and nitrogen( NO) in serum were measured by enzyme linked immunosorbent assay( ELISA). The expression levels of inhibitors of NF-κB kinase subunit-β( IKKβ),NF-κB inhibitor α( IκBα),TLR4,MyD88 and NF-κB p65 were measured by immunohistochemical staining and Western blot. Result: Compared with normal group,the myocardial injury in model group was obvious. The levels of IL-1β,IL-6,TNF-α,MCP-1 and NO in serum increased significantly( P < 0. 05),the expression level of IκBαdecreased significantly( P < 0. 05),and the expression levels of IKKβ,TLR4,MyD88 and NF-κB p65 increased significantly in myocardial tissue( P < 0. 05). Compared with model group,low,medium and high-dose compound Longmaining groups could significantly alleviate the degree of myocardial injury in rats,significantly decrease IL-1β,IL-6,TNF-α,MCP-1 and NO levels in the serum( P < 0. 05),significantly increase the expression level of IκBα( P < 0. 05),and decreased the expression levels of IKKβ,TLR4,MyD88 and NF-κB p65( P < 0. 05).Conclusion: Compound Longmaining plays a protective effect on acute myocardial infarction by regulatingthe expressions of TLR4/MyD88/NF-κB p65 signaling pathway and relevant inflammatory factors.
Objective: To investigate the protective effect of compound Longmaining isoprenaline hydrochloride-induced myocardial infarction model and its effect on Toll-like receptor 4( TLR4)/myeloid differentiation factor 88( MyD88)/nuclear transcription factor-kappa B( NF-κB) signaling pathway. Method:Forty-eight male SD rats were randomly divided into 6 groups: normal group,model group,compound salvia miltiorrhiza drop pill group( 0. 072 9 g·kg~(-1)),and low,medium and high-dose compound Longmaining decoction groups( 0. 36,0. 71,1. 43 g·kg~(-1)). The acute myocardial infarction model was induced through subcutaneous injection with isoproterenol. The pathological changes of myocardial tissue were examined by hematoxylin-eosin( HE) staining. The levels of interleukin-1β( IL-1β),interleukin-6( IL-6),tumor necrosis factor-α( TNF-α),monocyte chemotaxis protein-1( MCP-1) and nitrogen( NO) in serum were measured by enzyme linked immunosorbent assay( ELISA). The expression levels of inhibitors of NF-κB kinase subunit-β( IKKβ),NF-κB inhibitor α( IκBα),TLR4,MyD88 and NF-κB p65 were measured by immunohistochemical staining and Western blot. Result: Compared with normal group,the myocardial injury in model group was obvious. The levels of IL-1β,IL-6,TNF-α,MCP-1 and NO in serum increased significantly( P < 0. 05),the expression level of IκBαdecreased significantly( P < 0. 05),and the expression levels of IKKβ,TLR4,MyD88 and NF-κB p65 increased significantly in myocardial tissue( P < 0. 05). Compared with model group,low,medium and high-dose compound Longmaining groups could significantly alleviate the degree of myocardial injury in rats,significantly decrease IL-1β,IL-6,TNF-α,MCP-1 and NO levels in the serum( P < 0. 05),significantly increase the expression level of IκBα( P < 0. 05),and decreased the expression levels of IKKβ,TLR4,MyD88 and NF-κB p65( P < 0. 05).Conclusion: Compound Longmaining plays a protective effect on acute myocardial infarction by regulatingthe expressions of TLR4/MyD88/NF-κB p65 signaling pathway and relevant inflammatory factors.
引文
[1]胡佳,杨杭燕,刘清海,等.中医药治疗急性心肌梗死的研究进展[J].湖南中医杂志,2018,34(3):171-173.
[2]CHEN G,LI J L,SONG M C,et al.A mixed component supramolecular hydrogel to improve mice cardiac function and alleviate ventricular remodeling after acute myocardial infarction[J].Adv Funct Mater,2017,27(34):1701798.
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[4]董林娟,张文娟,刘诗雨,等.龙脉宁方中葛根素在心肌缺血大鼠体内药动学-药效学相关性研究[J].中国中药杂志,2016,41(8):1535-1540.
[5]王昌利,王莎,史亚军,等.复方龙脉宁不同配比药效学实验研究[J].辽宁中医药大学学报,2014,16(7):7-11.
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[7]张文娟,董林娟,宋逍,等.复方龙脉宁提取液对小鼠急性心肌缺血缺氧损伤的保护作用[J].中药新药与临床药理,2015,26(5):618-622.
[8]孙旭晖,何正飞,余健.瑞舒伐他汀对急性心肌梗死患者MMP-9、炎症因子、血脂水平及心功能的影响[J].中国生化药物杂志,2016,36(5):66-68.
[9]方芳,甘受益,冯光瑞,等.长期服用复方丹参滴丸对老年急性心肌梗死患者PCI术后左心室重构及炎症因子水平的影响[J].现代生物医学进展,2017,17(3):544-546,566.
[10]潘洁丽,郏国炯,王萌.中药制剂复方丹参滴丸对急性心肌梗死患者体内核因子-κB p65介导的炎症反应的影响[J].药物生物技术,2017,24(2):120-123.
[11]范金斌,李军,朱丹.血清Mb,c Tn I及炎症因子IL-1β,TNF-α检测对急性心肌梗死的临床诊断意义[J].实验与检验医学,2017,35(5):724-727.
[12]徐玲,许朝祥,蔡银链.Toll样受体3信号通路介导自身免疫性心肌炎炎症反应及作用机制[J].免疫学杂志,2017,33(8):655-660.
[13]章元沛.药理学实验[M].北京:人民卫生出版社,2010:238.
[14]吴广均,张云.Langendorff法、冠脉结扎法及ISO诱导心肌缺血模型的比较[J].现代中西医结合杂志,2011,20(22):2849-2851.
[15]HUANG H,GENG Q,YAO H,et al.Protective effect of scutellarin on myocardial infarction induced by isoprenaline in rats[J].Iran J Basic Med Sci,2018,21(3):267-276.
[16]楚纪明,马树运,李海峰,等.葛根有效成分及其药理作用研究进展[J].食品与药品,2015,17(2):142-146.
[17]罗仁书,何治勇.川芎有效成分药理作用的研究进展[J].中国医院用药评价与分析,2018,18(9):1294-1296.
[18]姜宇懋,王丹巧.川芎嗪药理作用研究进展[J].中国现代中药,2016,18(10):1364-1370.
[19]张宁,于栋华,周琦,等.穿山龙药理作用的研究进展[J].中国药房,2015,26(4):547-550.
[20]金平,张宇,王会,等.蜂胶的化学成分及药理作用[J].吉林中医药,2018,38(4):432-434.
[21]Prabhu S D,Frangogiannis N G.The biological basis for cardiac repair after myocardial infarction:from inflammation to fibrosis[J].Circul Res,2016,119(1):91-112.
[22]胡珍,陈景瑞,魏静,等.冠状动脉结扎制备大鼠心肌梗死模型及评价实验研究[J].天津中医药,2016,33(2):90-95.
[23]齐翌婷,但家立,晁晶,等.超声心动图对慢性心功能衰竭患者的左心功能评价的价值[J].医学影像学杂志,2017,27(8):1592-1594.
[24]贺忠梅,张红霞,杨敏,等.小鼠心肌梗死后补体系统激活与炎症反应的相关性研究[J].心肺血管病杂志,2014,33(1):98-102.
[25]孙丹丹,吴玉鹏,杨军,等.Toll样受体-4基因启动子区多态性与急性心肌梗死发病风险的关系[J].山东医药,2017,57(17):17-20.
[26]刁爱芹,王卉,潘爱萍,等.大豆异黄酮下调TLR4/My D88介导的信号通路保护心肌肥厚的作用机制[J].重庆医学,2018,47(20):2659-2662.
[27]李年忠.LPS诱导下池蝶蚌免疫相关基因LBP、TLR4、My D88和AP-1的表达分析以及TLR4和My D88互作关系[D].南昌:南昌大学,2018.
[28]刘振华,王世军.调营养心汤治疗亚健康失眠及对血清核转录因子-κB,炎症因子的作用研究[J].中国实验方剂学杂志,2016,22(6):177-180.
[29]赵运旺.NF-κB信号通路研究进展[J].甘肃科技,2016,32(21):117-123,112.
[30]陈恒文,李军,林飞,等.宣痹安痛方对心肌梗死后心室重构大鼠心功能的改善及机制[J].中国实验方剂学杂志,2017,23(23):71-78.
[31]李叶静,谈弋.紫草素通过TLR4/NF-κB信号通路抑制由脂多糖诱导的巨噬细胞炎症研究[J].安徽医药,2017,21(8):1384-1387.
[32]陈涛,陈文江,陈灿.NF-κB信号通路与心肌梗死关系的研究进展[J].现代医药卫生,2017,33(5):707-709.
[33]崔艳,曹小平.核转录因子-κB的研究进展[J].川北医学院学报,2014,29(5):510-513.
[34]朱雪坤,杨日芳,孟艳秋.Toll样受体4配体的研究进展[J].中国药理学与毒理学杂志,2016,30(4):389-396.
[35]杨燕燕,王训立.TLR4信号通路在中药抗动脉粥样硬化中的研究进展[J].中国中医基础医学杂志,2017,23(1):144-147.
[2]CHEN G,LI J L,SONG M C,et al.A mixed component supramolecular hydrogel to improve mice cardiac function and alleviate ventricular remodeling after acute myocardial infarction[J].Adv Funct Mater,2017,27(34):1701798.
[3]贾杰,周瑛,韦中奎.中药治疗心血管疾病研究概况[J].北方药学,2014,11(2):63-64.
[4]董林娟,张文娟,刘诗雨,等.龙脉宁方中葛根素在心肌缺血大鼠体内药动学-药效学相关性研究[J].中国中药杂志,2016,41(8):1535-1540.
[5]王昌利,王莎,史亚军,等.复方龙脉宁不同配比药效学实验研究[J].辽宁中医药大学学报,2014,16(7):7-11.
[6]马海芳.复方龙脉宁标准汤剂的研究[D].咸阳:陕西中医药大学,2015.
[7]张文娟,董林娟,宋逍,等.复方龙脉宁提取液对小鼠急性心肌缺血缺氧损伤的保护作用[J].中药新药与临床药理,2015,26(5):618-622.
[8]孙旭晖,何正飞,余健.瑞舒伐他汀对急性心肌梗死患者MMP-9、炎症因子、血脂水平及心功能的影响[J].中国生化药物杂志,2016,36(5):66-68.
[9]方芳,甘受益,冯光瑞,等.长期服用复方丹参滴丸对老年急性心肌梗死患者PCI术后左心室重构及炎症因子水平的影响[J].现代生物医学进展,2017,17(3):544-546,566.
[10]潘洁丽,郏国炯,王萌.中药制剂复方丹参滴丸对急性心肌梗死患者体内核因子-κB p65介导的炎症反应的影响[J].药物生物技术,2017,24(2):120-123.
[11]范金斌,李军,朱丹.血清Mb,c Tn I及炎症因子IL-1β,TNF-α检测对急性心肌梗死的临床诊断意义[J].实验与检验医学,2017,35(5):724-727.
[12]徐玲,许朝祥,蔡银链.Toll样受体3信号通路介导自身免疫性心肌炎炎症反应及作用机制[J].免疫学杂志,2017,33(8):655-660.
[13]章元沛.药理学实验[M].北京:人民卫生出版社,2010:238.
[14]吴广均,张云.Langendorff法、冠脉结扎法及ISO诱导心肌缺血模型的比较[J].现代中西医结合杂志,2011,20(22):2849-2851.
[15]HUANG H,GENG Q,YAO H,et al.Protective effect of scutellarin on myocardial infarction induced by isoprenaline in rats[J].Iran J Basic Med Sci,2018,21(3):267-276.
[16]楚纪明,马树运,李海峰,等.葛根有效成分及其药理作用研究进展[J].食品与药品,2015,17(2):142-146.
[17]罗仁书,何治勇.川芎有效成分药理作用的研究进展[J].中国医院用药评价与分析,2018,18(9):1294-1296.
[18]姜宇懋,王丹巧.川芎嗪药理作用研究进展[J].中国现代中药,2016,18(10):1364-1370.
[19]张宁,于栋华,周琦,等.穿山龙药理作用的研究进展[J].中国药房,2015,26(4):547-550.
[20]金平,张宇,王会,等.蜂胶的化学成分及药理作用[J].吉林中医药,2018,38(4):432-434.
[21]Prabhu S D,Frangogiannis N G.The biological basis for cardiac repair after myocardial infarction:from inflammation to fibrosis[J].Circul Res,2016,119(1):91-112.
[22]胡珍,陈景瑞,魏静,等.冠状动脉结扎制备大鼠心肌梗死模型及评价实验研究[J].天津中医药,2016,33(2):90-95.
[23]齐翌婷,但家立,晁晶,等.超声心动图对慢性心功能衰竭患者的左心功能评价的价值[J].医学影像学杂志,2017,27(8):1592-1594.
[24]贺忠梅,张红霞,杨敏,等.小鼠心肌梗死后补体系统激活与炎症反应的相关性研究[J].心肺血管病杂志,2014,33(1):98-102.
[25]孙丹丹,吴玉鹏,杨军,等.Toll样受体-4基因启动子区多态性与急性心肌梗死发病风险的关系[J].山东医药,2017,57(17):17-20.
[26]刁爱芹,王卉,潘爱萍,等.大豆异黄酮下调TLR4/My D88介导的信号通路保护心肌肥厚的作用机制[J].重庆医学,2018,47(20):2659-2662.
[27]李年忠.LPS诱导下池蝶蚌免疫相关基因LBP、TLR4、My D88和AP-1的表达分析以及TLR4和My D88互作关系[D].南昌:南昌大学,2018.
[28]刘振华,王世军.调营养心汤治疗亚健康失眠及对血清核转录因子-κB,炎症因子的作用研究[J].中国实验方剂学杂志,2016,22(6):177-180.
[29]赵运旺.NF-κB信号通路研究进展[J].甘肃科技,2016,32(21):117-123,112.
[30]陈恒文,李军,林飞,等.宣痹安痛方对心肌梗死后心室重构大鼠心功能的改善及机制[J].中国实验方剂学杂志,2017,23(23):71-78.
[31]李叶静,谈弋.紫草素通过TLR4/NF-κB信号通路抑制由脂多糖诱导的巨噬细胞炎症研究[J].安徽医药,2017,21(8):1384-1387.
[32]陈涛,陈文江,陈灿.NF-κB信号通路与心肌梗死关系的研究进展[J].现代医药卫生,2017,33(5):707-709.
[33]崔艳,曹小平.核转录因子-κB的研究进展[J].川北医学院学报,2014,29(5):510-513.
[34]朱雪坤,杨日芳,孟艳秋.Toll样受体4配体的研究进展[J].中国药理学与毒理学杂志,2016,30(4):389-396.
[35]杨燕燕,王训立.TLR4信号通路在中药抗动脉粥样硬化中的研究进展[J].中国中医基础医学杂志,2017,23(1):144-147.