喘可治注射液对甲型H1N1流感病毒FM1株致小鼠肺炎的影响
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摘要
目的:评价喘可治注射液对甲型H1N1流感病毒FM1株致小鼠肺炎模型的药效作用。方法:将ICR小鼠随机分为正常组、模型组、达菲组(27.5 mg·kg~(-1)·d~(-1))、喘可治注射液组(1.5 m L·kg~(-1)·d~(-1))。除正常组外,其余各组采用甲型H1N1流感病毒FM1株滴鼻感染ICR小鼠造成病毒性肺炎模型,分别进行死亡保护实验、治疗性给药实验和预防性给药实验。死亡保护实验中每组20只小鼠,以2倍半数致死量(LD_(50))流感病毒液滴鼻感染,感染后当天开始给药,每天1次,连续4 d,观察感染后14 d内动物的死亡情况,并计算死亡率、死亡保护率;治疗性给药实验中每组10只小鼠,以0.8倍LD_(50)流感病毒液滴鼻感染,感染后当天开始给药,每天1次,连续4 d,于实验第5天处死小鼠,采用酶联免疫吸附测定(ELISA)检测肺组织中白细胞介素-8(IL-8)含量及脑组织内发热介质前列腺素E_2(PGE_2),精氨酸加压素(AVP)的含量,实时荧光定量聚合酶链式反应(Real-time PCR)检测肺组织内相对病毒载量;预防性给药实验中每组10只小鼠,分组当天开始给药,每天1次,连续5 d,末次给药1 h后,除正常组外,其余各组以0.8倍LD_(50)流感病毒滴鼻感染ICR小鼠,感染后第4天称重后处死小鼠,计算各组肺指数、脾指数、胸腺指数及肺指数抑制率,Real-time PCR检测肺组织内相对病毒载量。结果:与正常组比较,模型组小鼠IL-8,PGE2含量,肺指数及肺组织内流感病毒株病毒载量均显著升高(P<0.01);与模型组比较,治疗性给予喘可治注射液可明显降低感染后小鼠的死亡率、肺组织内IL-8含量、脑组织内PGE_2含量及肺组织中流感病毒病毒载量(P<0.05,P<0.01);预防性给予喘可治注射液可显著提高感染小鼠的胸腺指数(P<0.01),明显降低感染小鼠肺指数及肺组织中流感病毒病毒载量(P<0.05);两种给药方式均可明显减轻支气管、血管周围及肺泡间质等组织炎性细胞浸润,减少管腔内红细胞渗出。结论:喘可治注射液对甲型H1N1流感病毒感染所致病毒性肺炎小鼠模型有较好的治疗和预防作用,其作用机制可能与减轻炎症反应和抑制病毒复制有关。
Objective: To evaluate the effect of Chuankezhi injection on mouse model of pneumonia induced by influenza A( H1N1) FM1 strain. Method: ICR mice were randomly divided into normal group,model group,tamiflu control group( 27. 5 mg·kg~(-1)·d~(-1)) and Chuankezhi injection group( 1. 5 m L·kg~(-1)·d~(-1)). In the death protection experiment,mice were infected with 2 × half lethal dose( LD_(50)) of influenza virus FM1. The Chuankezhi injection was given once a day for 4 days. The number of death animal within 14 days was counted.The mortality and the death protection rate were calculated. In the treatment experiment,mice were infected with0. 8 × LD_(50) of influenza virus,and the Chuankezhi injection was given once a day for 4 days. On the 5~(th)day after the infection,the levels of interleukin-8( IL-8) in lung,prostaglandin E_2( PGE_2) and vasopressin( AVP) in brain were tested by enzyme-linked immunosorbent assay( ELISA). The viral load of influenza virus in lung was tested by Real-time PCR. In the pre-treatment experiment,mice were given Chuankezhi injection once a day for 5 days. 1 hour after the last treatment,mice were infected with 0. 8 × LD_(50) influenza virus. 4 days after the infection,the lung index,spleen index,thymus index,and viral load in lung tissue were calculated. Result: Compared with normal group,the IL-8,PGE_2 content,lung index and viral load in the lung tissue of model group were significantly increased( P < 0. 01). Compared with model group,Treatment of Chuankezhi injection could reduce the rate of deaths. Significantly inhibit the level of IL-8,PGE_2,and the viral load of influenza( P < 0. 05,P <0. 01). Pre-treatment of Chuankezhi injection could significantly increase the thymus index of infected mice( P <0. 01),reduce lung index and the viral load of influenza( P < 0. 05). Both administration methods can significantly reduce the infiltration of inflammatory cells in the bronchial,perivascular and alveolar interstitium and reduce the exudation of red blood cells in the lumen. Conclusion: Chuankezhi injection could effectively prevent the mouse model of pneumonia induced by influenza A( H1N1) virus. The mechanism might be related to the reduction of inflammation and inhibiting viral replication.
Objective: To evaluate the effect of Chuankezhi injection on mouse model of pneumonia induced by influenza A( H1N1) FM1 strain. Method: ICR mice were randomly divided into normal group,model group,tamiflu control group( 27. 5 mg·kg~(-1)·d~(-1)) and Chuankezhi injection group( 1. 5 m L·kg~(-1)·d~(-1)). In the death protection experiment,mice were infected with 2 × half lethal dose( LD_(50)) of influenza virus FM1. The Chuankezhi injection was given once a day for 4 days. The number of death animal within 14 days was counted.The mortality and the death protection rate were calculated. In the treatment experiment,mice were infected with0. 8 × LD_(50) of influenza virus,and the Chuankezhi injection was given once a day for 4 days. On the 5~(th)day after the infection,the levels of interleukin-8( IL-8) in lung,prostaglandin E_2( PGE_2) and vasopressin( AVP) in brain were tested by enzyme-linked immunosorbent assay( ELISA). The viral load of influenza virus in lung was tested by Real-time PCR. In the pre-treatment experiment,mice were given Chuankezhi injection once a day for 5 days. 1 hour after the last treatment,mice were infected with 0. 8 × LD_(50) influenza virus. 4 days after the infection,the lung index,spleen index,thymus index,and viral load in lung tissue were calculated. Result: Compared with normal group,the IL-8,PGE_2 content,lung index and viral load in the lung tissue of model group were significantly increased( P < 0. 01). Compared with model group,Treatment of Chuankezhi injection could reduce the rate of deaths. Significantly inhibit the level of IL-8,PGE_2,and the viral load of influenza( P < 0. 05,P <0. 01). Pre-treatment of Chuankezhi injection could significantly increase the thymus index of infected mice( P <0. 01),reduce lung index and the viral load of influenza( P < 0. 05). Both administration methods can significantly reduce the infiltration of inflammatory cells in the bronchial,perivascular and alveolar interstitium and reduce the exudation of red blood cells in the lumen. Conclusion: Chuankezhi injection could effectively prevent the mouse model of pneumonia induced by influenza A( H1N1) virus. The mechanism might be related to the reduction of inflammation and inhibiting viral replication.
引文
[1]陈闻天,黄鹤,于汉杰,等.甲型流感病毒基因组的进化与变异[J].中国病毒病杂志,2018,8(5):411-420.
[2]LU C J,JIANG Z T,FAN X M,et al. Ametabonomic approach to the effect evaluation of treatment in patients infected with influenza A(H1N1)[J]. Talanta,2012,100(20):51-56.
[3]李立,杜雅薇,寇爽,等.金银花、连翘对甲型H1N1流感免疫调节通路影响的生物信息学分析[J].中国实验方剂学杂志,2017,23(10):201-204.
[4]王燕. H1N1流感病毒性肺炎临床诊治启示[J].临床肺科杂志,2018,23(11):2128-2129.
[5]陈琪,吴莹,齐晓宇,等.中医成药治疗甲型H1N1流感研究进展[J].现代生物医学进展,2016,16(19):3793-3796.
[6]王振中,鮑琳琳,孙兰,等.热毒宁注射液抗甲型H1N1流感病毒作用机制研究[J].中草药,2014,45(1):90-93.
[7]朱泽豪,廖柳,严夏.基于贝叶斯框架的中药注射液联合西药治疗小儿支气管哮喘的Meta分析[J].中成药,2018,40(8):1889-1894.
[8]吴文先,高雅,刘霞.喘可治注射液不同给药途径治疗婴幼儿毛细支气管炎临床疗效比较[J].中医临床研究,2016,8(25):113-114.
[9]郑红英,杨春莲,何丽林.喘可治注射液联合多索茶碱治疗慢性阻塞性肺疾病急性发作期的疗效观察[J].现代药物与临床,2018,33(4):809-812.
[10]肇静娴,曾耀英,王青,等.喘可治注射液对人外周血单个核细胞Th1/Th2细胞因子谱的影响[J].中国免疫学杂志,2006,22(4):356-359.
[11]李承刚.纳米材料PAMAM G3及甲型H1N1流感病毒导致小鼠急性肺损伤的致病机理研究[D].北京:北京协和医学院,2010.
[12]高晶晶,李鑫,乔瑞洁,等.百日咳小鼠呼吸道感染模型的建立与应用[J].微生物学免疫学进展,2018,46(5):28-35.
[13]郭姗姗,刘颖,高英杰,等.小儿肺热咳喘口服液对甲型H1N1流感病毒感染小鼠的影响[J].中国实验方剂学杂志,2011,17(2):152-155.
[14]李玲,吴佳敏,欧阳建军,等.抗流感病毒性肺炎的有效中药复方筛选及机制研究[J].中国免疫学杂志,2018,34(8):1168-1173.
[15]郭姗姗,高英杰,马雪萍,等.一叶抗流感胶囊抑制甲型H1N1流感病毒感染的体内外研究[J].中国实验方剂学杂志,2014,20(18):123-127.
[16]尹香琳,张婧瑶,刘卫东,等. 1-磷酸鞘氨醇受体2介导PI3K/Akt/e NOS通路抑制甲型流感病毒诱导的病毒性肺炎[J].中国病理生理杂志,2018,34(11):2062-2067.
[17]Garten R J,Davis C T,Russell C A,et al. Antigenic and genetic characteristics of swine-origin 2009 A(H1N1)influenza viruses circulating in humans[J]. Science,2009,325(5937):197-201.
[18]陈创荣,刘建兴,李耿,等.岗藿抗感汤对小鼠H1N1流感病毒肺炎的影响[J].中华中医药杂志,2015,30(9):3318-3321.
[19]刘蓉,武志强,何敏,等.桂枝挥发油与桂皮醛对病毒性肺炎小鼠细胞因子及T细胞亚群的影响[J].中国实验方剂学杂志,2015,21(18):139-143.
[20]马月霞.常用中药注射液不同配伍抗小鼠甲型H1N1流感重症肺炎的疗效研究[D].北京:北京中医药大学,2014.
[21]孙兰,段书敏,周军,等.热毒宁注射液体外抑制甲型H1N1流感病毒的研究[J].现代药物与临床,2014,29(8):848-851.
[22]孙兰,刘艾林,王振中,等.热毒宁注射液及其组分对流感病毒神经氨酸酶的抑制作用研究[J].现代药物与临床,2014,29(1):27-31.
[23]张艳丽,顾立刚,曾郁敏,等.毒热平注射液体外抗甲型流感病毒的作用研究[J].辽宁中医药大学学报,2008,10(3):6-8.
[24]陈美娟,葛李,肖顺汉,等.银黄注射液体内外抗流感病毒的实验研究[J].时珍国医国药,2007,18(3):591-592.
[25]卢芳,冯镇凯,杨晓旭,等.基于免疫调节作用的淫羊藿各拆分组分筛选研究[J].中华中医药学刊,2018,36(6):1331-1335.
[26]路宇仁,陈昳冰,崔元璐,等.淫羊藿苷药理作用研究进展[J].中国实验方剂学杂志,2018,24(17):209-220.
[27]饶鸿宇,陈滔彬,何彦,等.南药巴戟天化学成分与药理研究进展[J].中南药学,2018,16(11):1567-1574.
[28]费辛,蔡宇波,曹兰芳,等.喘可治注射液对大鼠抗炎和免疫调节机制的研究[J].药学服务与研究,2011,11(5):333-336.
[29]毛鑫,姚荣妹,高英杰,等.抗病毒口服液治疗甲型H1N1流感病毒感染药效及机制探讨[J].中国药物警戒,2018,15(9):518-522.
[30]Nurmi K,Kareinen I,Virkanen J,et al. Hemin and cobalt protoporphyrin inhibit NLRP3 inflammasome activation by enhancing autophagy:a novel mechanism of inflammasome regulation[J]. J Innate Immun,2017,9(1):65-82.
[31]GAO L N,CUI Y L,WANG Q S,et al. Amelioration of Danhong injection on the lipopolysaccharide-stimulated systemic acute inflammatory reaction via multi-target strategy[J]. J Ethnopharmacol,2013,149(3):772-782.
[32]于艳,贾天柱,才谦.茅苍术及其麸炒品对胃溃疡大鼠抗炎作用的比较研究[J].中国中药杂志,2016,41(4):705-710.
[33]刘颖,时宇静,时瀚,等.银翘解毒软胶囊对流感病毒感染小鼠肺炎模型肺组织病毒载量及M1蛋白表达的影响[J].药学学报,2011,46(6):650-655.
[2]LU C J,JIANG Z T,FAN X M,et al. Ametabonomic approach to the effect evaluation of treatment in patients infected with influenza A(H1N1)[J]. Talanta,2012,100(20):51-56.
[3]李立,杜雅薇,寇爽,等.金银花、连翘对甲型H1N1流感免疫调节通路影响的生物信息学分析[J].中国实验方剂学杂志,2017,23(10):201-204.
[4]王燕. H1N1流感病毒性肺炎临床诊治启示[J].临床肺科杂志,2018,23(11):2128-2129.
[5]陈琪,吴莹,齐晓宇,等.中医成药治疗甲型H1N1流感研究进展[J].现代生物医学进展,2016,16(19):3793-3796.
[6]王振中,鮑琳琳,孙兰,等.热毒宁注射液抗甲型H1N1流感病毒作用机制研究[J].中草药,2014,45(1):90-93.
[7]朱泽豪,廖柳,严夏.基于贝叶斯框架的中药注射液联合西药治疗小儿支气管哮喘的Meta分析[J].中成药,2018,40(8):1889-1894.
[8]吴文先,高雅,刘霞.喘可治注射液不同给药途径治疗婴幼儿毛细支气管炎临床疗效比较[J].中医临床研究,2016,8(25):113-114.
[9]郑红英,杨春莲,何丽林.喘可治注射液联合多索茶碱治疗慢性阻塞性肺疾病急性发作期的疗效观察[J].现代药物与临床,2018,33(4):809-812.
[10]肇静娴,曾耀英,王青,等.喘可治注射液对人外周血单个核细胞Th1/Th2细胞因子谱的影响[J].中国免疫学杂志,2006,22(4):356-359.
[11]李承刚.纳米材料PAMAM G3及甲型H1N1流感病毒导致小鼠急性肺损伤的致病机理研究[D].北京:北京协和医学院,2010.
[12]高晶晶,李鑫,乔瑞洁,等.百日咳小鼠呼吸道感染模型的建立与应用[J].微生物学免疫学进展,2018,46(5):28-35.
[13]郭姗姗,刘颖,高英杰,等.小儿肺热咳喘口服液对甲型H1N1流感病毒感染小鼠的影响[J].中国实验方剂学杂志,2011,17(2):152-155.
[14]李玲,吴佳敏,欧阳建军,等.抗流感病毒性肺炎的有效中药复方筛选及机制研究[J].中国免疫学杂志,2018,34(8):1168-1173.
[15]郭姗姗,高英杰,马雪萍,等.一叶抗流感胶囊抑制甲型H1N1流感病毒感染的体内外研究[J].中国实验方剂学杂志,2014,20(18):123-127.
[16]尹香琳,张婧瑶,刘卫东,等. 1-磷酸鞘氨醇受体2介导PI3K/Akt/e NOS通路抑制甲型流感病毒诱导的病毒性肺炎[J].中国病理生理杂志,2018,34(11):2062-2067.
[17]Garten R J,Davis C T,Russell C A,et al. Antigenic and genetic characteristics of swine-origin 2009 A(H1N1)influenza viruses circulating in humans[J]. Science,2009,325(5937):197-201.
[18]陈创荣,刘建兴,李耿,等.岗藿抗感汤对小鼠H1N1流感病毒肺炎的影响[J].中华中医药杂志,2015,30(9):3318-3321.
[19]刘蓉,武志强,何敏,等.桂枝挥发油与桂皮醛对病毒性肺炎小鼠细胞因子及T细胞亚群的影响[J].中国实验方剂学杂志,2015,21(18):139-143.
[20]马月霞.常用中药注射液不同配伍抗小鼠甲型H1N1流感重症肺炎的疗效研究[D].北京:北京中医药大学,2014.
[21]孙兰,段书敏,周军,等.热毒宁注射液体外抑制甲型H1N1流感病毒的研究[J].现代药物与临床,2014,29(8):848-851.
[22]孙兰,刘艾林,王振中,等.热毒宁注射液及其组分对流感病毒神经氨酸酶的抑制作用研究[J].现代药物与临床,2014,29(1):27-31.
[23]张艳丽,顾立刚,曾郁敏,等.毒热平注射液体外抗甲型流感病毒的作用研究[J].辽宁中医药大学学报,2008,10(3):6-8.
[24]陈美娟,葛李,肖顺汉,等.银黄注射液体内外抗流感病毒的实验研究[J].时珍国医国药,2007,18(3):591-592.
[25]卢芳,冯镇凯,杨晓旭,等.基于免疫调节作用的淫羊藿各拆分组分筛选研究[J].中华中医药学刊,2018,36(6):1331-1335.
[26]路宇仁,陈昳冰,崔元璐,等.淫羊藿苷药理作用研究进展[J].中国实验方剂学杂志,2018,24(17):209-220.
[27]饶鸿宇,陈滔彬,何彦,等.南药巴戟天化学成分与药理研究进展[J].中南药学,2018,16(11):1567-1574.
[28]费辛,蔡宇波,曹兰芳,等.喘可治注射液对大鼠抗炎和免疫调节机制的研究[J].药学服务与研究,2011,11(5):333-336.
[29]毛鑫,姚荣妹,高英杰,等.抗病毒口服液治疗甲型H1N1流感病毒感染药效及机制探讨[J].中国药物警戒,2018,15(9):518-522.
[30]Nurmi K,Kareinen I,Virkanen J,et al. Hemin and cobalt protoporphyrin inhibit NLRP3 inflammasome activation by enhancing autophagy:a novel mechanism of inflammasome regulation[J]. J Innate Immun,2017,9(1):65-82.
[31]GAO L N,CUI Y L,WANG Q S,et al. Amelioration of Danhong injection on the lipopolysaccharide-stimulated systemic acute inflammatory reaction via multi-target strategy[J]. J Ethnopharmacol,2013,149(3):772-782.
[32]于艳,贾天柱,才谦.茅苍术及其麸炒品对胃溃疡大鼠抗炎作用的比较研究[J].中国中药杂志,2016,41(4):705-710.
[33]刘颖,时宇静,时瀚,等.银翘解毒软胶囊对流感病毒感染小鼠肺炎模型肺组织病毒载量及M1蛋白表达的影响[J].药学学报,2011,46(6):650-655.