摘要
目的评价夏枯草水提物抗寨卡病毒(Zika virus,ZIKV)的活性。方法将IFNAR1-/C57BL/6小鼠分为空白组、模型组、布喹那组(10 mg·kg^-1)、夏枯草水提物高剂量组(900 mg·kg^-1)及低剂量组(450 mg·kg^-1),连续灌胃给药12 d,每天1次。第1次灌胃给药30 min后,一次性腹腔注射寨卡病毒,制备寨卡病毒感染模型,空白组给予同体积生理盐水。分别于给药第3、5天检测小鼠血清中病毒载量,并观察小鼠12 d的生存情况。体外实验中,采用四甲基偶氮唑盐比色法(MTT)检测夏枯草水提物对Vero和HeLa细胞的毒性;通过观察细胞病变(CPE)现象,检测夏枯草水提物对寨卡病毒引起的细胞病变的影响;应用Real time PCR和Western Blot检测夏枯草水提物对病毒核酸复制和蛋白表达的影响;通过不同给药模式实验研究夏枯草水提物可能的抗病毒机制。结果与空白组比较,模型组小鼠平均生存时间明显降低(P<0.01),第3、5天血清中病毒RNA拷贝数升高(P<0.001);与模型组比较,高、低剂量组小鼠平均生存时间提高(P<0.01),并能明显降低小鼠第3天、第5天血清中病毒RNA拷贝数(P<0.001,P<0.01,P<0.05);与模型组比较,布喹那组小鼠存活率和病毒RNA拷贝数变化均不明显(P>0.05)。夏枯草水提物体外抗寨卡病毒的半数有效浓度(EC50)=(51.32±2.13)μg·mL^-1,治疗指数(SI)=48.71。与空白组比较,模型组能明显检测到病毒核酸复制和蛋白表达升高(P<0.001);与模型组比较,布喹那组能明显抑制病毒核酸复制和蛋白表达(P<0.001);不同浓度的夏枯草水提物组均能不同程度上抑制病毒核酸复制和蛋白表达(P<0.001)。不同给药模式实验结果显示,预混给药效果优于治疗给药和同时给药。结论夏枯草水提物可提高寨卡病毒感染小鼠的生存率,并可明显降低小鼠血清中的寨卡病毒RNA水平。体外实验结果显示,夏枯草水提物能明显减少寨卡病毒引起的细胞病变、抑制寨卡病毒核酸复制和蛋白表达,并可能通过直接杀灭寨卡病毒或者抑制病毒识别和进入宿主细胞发挥抗病毒作用。
Objective To evaluate the activity of Prunella vulgaris aqueous extract against Zika virus(ZIKV).Methods IFNAR1-/C57BL/6 mice were divided into blank group,model group,brequinar group(10 mg·kg^-1),Prunella vulgaris aqueous extract high-dose group(900 mg·kg^-1)and low-dose group(450 mg·kg^-1).Drugs were administered by intragastric administration once a day for 12 days.Thirty minutes after the first intragastric administration,Zika virus was injected intraperitoneally to prepare the Zika virus infection model.The blank group mice were given the same volume of saline.The viral load in the serum of mice was detected on the 3rd and 5th days of administration,and the survival of the mice was observed for 12 days.In the in vitro experiments,the MTT colorimetric assay method was used to detect the toxicity of Prunella vulgaris aqueous extract to Vero and HeLa cells.By observing the CPE,the effect of Prunella vulgaris aqueous extract on cytopathy caused by Zika virus was detected.Real time PCR and Western Blot were used to detect the effect of Prunella vulgaris aqueous extract on replication of viral nucleic acid and protein expression.The possible antiviral mechanism of Prunella vulgaris aqueous extract was studied through different modes of administration.Results Compared with the blank group,the mean survival time of mice in the model group was significantly reduced(P<0.01),the viral RNA copy number in blood increased on days 3 and 5(P<0.001).Compared with the model group,the mean survival time of mice in the high and low dose groups were improved(P<0.01),and it significantly reduced viral RNA copy number in blood of mice on day 3 and day 5(P<0.001,P<0.01,P<0.05).Compared with the model group,the survival rate and viral RNA copy number of mice in the brequinar group were not significantly changed(P>0.05).The EC50 of Prunella vulgaris aqueous extract against Zika virus is(51.32±2.13)μg·mL^-1,SI=48.71.Compared with the blank group,the viral RNA replication and increased protein expression in the model group could be clearly detected(P<0.001).Compared with the model group,the brequinar group can significantly inhibit viral RNA replication and protein expression(P<0.001).Different concentration of Prunella vulgaris aqueous extract groups can inhibit viral RNA replication and protein expression to varying degrees(P<0.001).Experimental results of different administration modes showed that the effect of premix administration was better than that of treatment administration and simultaneous administration.Conclusion Prunella vulgaris aqueous extract can improve the survival rate of mice infected with Zika virus,and significantly reduce the level of viral RNA in the serum of mice.In vitro experiments results showed that Prunella vulgaris aqueous extract can significantly reduce the cytopathic effect caused by Zika virus,inhibit the nucleic acid replication and protein expression of ZIKV,and may play an antiviral role by directly killing the Zika virus or inhibiting virus recognition and entering the host cell.
作者
李兆新
许江涛
林雪妹
吕东勇
刘小虹
李耿
LI Zhaoxin;XU Jiangtao;LIN Xuemei;LYU Dongyong;LIU Xiaohong;LI Geng(Laboratorial Animal Center,Guangzhou University of Chinese Medicine,Guangzhou 510000 Guangdong,China;Chinese Medicine College,Guangzhou University of Chinese Medicine,Guangzhou 510006 Guangdong,China;First Affiliated Hospital,Guangzhou University of Chinese Medicine,Guangzhou 510000 Guangdong,China)
出处
《中药新药与临床药理》
CAS
CSCD
北大核心
2020年第12期1408-1415,共8页
Traditional Chinese Drug Research and Clinical Pharmacology
基金
广东省重点领域研发计划“岭南中医药现代化”重点专项(192019082450200013)
国家自然科学基金项目(81973549,81803813)
广东省教育厅科技计划项目(2019KYTD102,2018KCXTD007)。
关键词
夏枯草水提物
寨卡病毒
体内外实验
抗病毒活性
Prunella vulgaris aqueous extract
Zika virus
in vitro and in vivo experiments
antiviral activity
