CN113940942A - 黄芩苷在治疗冠状病毒感染性疾病中的应用 - Google Patents
黄芩苷在治疗冠状病毒感染性疾病中的应用 Download PDFInfo
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Abstract
本发明公开了黄芩苷在治疗冠状病毒感染性疾病中的应用。通过在感染人冠状病毒HCoV‑229E的体外细胞试验和小鼠试验中验证黄芩苷的新用途,该技术方案具有新颖性,并进行了黄芩苷在治疗由HCoV‑229E病毒感染引起的肺炎的疗效试验,结果显示,黄芩苷在体外对HCoV‑229E感染细胞有一定的抑制作用;对感染HCoV‑229E病毒的肺炎小鼠肺指数及抑制率中表现出了良好的药效,能降低感染HCoV‑229E病毒的肺炎小鼠肺组织的病毒载量,升高小鼠外周血淋巴细胞比例,降低小鼠肺组织IL‑2、IL‑10、TNF‑α、IFN‑β的含量。
Description
技术领域
本发明涉及药物治疗领域,具体涉及黄芩苷在治疗冠状病毒感染性疾病 中的应用。
背景技术
冠状病毒是具有外套膜的正链单股RNA病毒。病毒包膜上存在棘突, 不同的冠状病毒的棘突差异明显,看似冠状,所以命名为“冠状病毒”。病 毒粒子外包着脂肪膜,膜表面有三种糖蛋白:刺突糖蛋白(S,SpikeProtein, 是受体结合位点、溶细胞作用和主要抗原位点);小包膜糖蛋白(E, EnvelopeProtein,较小,与包膜结合的蛋白);膜糖蛋白(M,MembraneProtein, 负责营养物质的跨膜运输、新生病毒出芽释放与病毒外包膜的形成)。少数种 类还有血凝素糖蛋白(HE蛋白,Haemaglutinin-esterase)。冠状病毒的核酸为 非节段单链(+)RNA,长27-31kd,是RNA病毒中最长的RNA核酸链,具有 正链RNA特有的重要结构特征:即RNA链5’端有甲基化“帽子”,3’端 有PolyA“尾巴”结构。这一结构与真核mRNA非常相似,也是其基因组 RNA自身可以发挥翻译模板作用的重要结构基础,而省去了RNA-DNA-RNA 的转录过程。冠状病毒的RNA和RNA之间重组率非常高,病毒极易出现变 异,抗原性发生变化的结果是导致原有疫苗失效。
冠状病毒共有七种,其中四种会引起普通的感冒症状,分别为229E、 NL63、OC43、HKU1,剩下的两种则是令人闻之丧胆的SARS(非典型肺炎) 以及MERS-CoV(中东呼吸综合征冠状病毒),这两种病毒曾入选为人类历史 上最可怕的病毒之一。2020年1月12日被世界卫生组织正式命名为“2019 新型冠状病毒”(2019-nCoV),是人类历史上确定的第7种可感染人的冠状病 毒(Human Coronavirus,HCoV)。
此外,冠状病毒基因组序列还具有同源性,可分为4个属,α,β,γ 和δ。γ和δ冠状病毒属只感染动物,α冠状病毒属包括人冠状病毒229E 和NL63,β冠状病毒属包括4个家系,HKU1和OC43属于A家系;B家 系包括SARS,与重症肺炎有关;MERS则属于β冠状病毒C家系。
人冠状病毒HCoV-229E属α群正链RNA病毒冠状病毒,是引起人类上 呼吸道感染的病原,常引起人的普通感冒。在临床上除引起呼吸道感染外还 会引起胃肠道疾病及神经系统症状,如肺炎等,研究表明,当HCoV-229E 感染肺部时,会释放促炎细胞因子,包括IL-2、IL-10、TNF-α、IFN-β,引 起肺炎。
因此迫切需要一种预防和治疗人冠状病毒药物的出现。
发明内容
黄芩苷是从双子叶唇形科植物黄芩(Scutellaria baicalensis Georgi)的干燥根中提取分离出来的一种黄酮类化合物,为黄色针状结晶。通常在稀酸条件 下比较稳定,如在2%硫酸水溶液中不会发生水解,但酸的浓度加大,温度 升高至110℃时则可水解生成葡萄糖醛酸及黄芩苷元,即黄芩素。
临床研究发现,黄芩苷具有显著的生物活性,具有抑菌、利尿、抗炎、 抗变态及解痉作用,并且具有较强的抗癌反应等生理效能,黄芩苷还能吸收 紫外线,清除氧自由基,且能抑制黑色素的生成。但目前还没有报道论述关 于黄芩苷可治疗人类冠状病毒HCoV-229E。
为了实现上述目的,本发明采用了如下技术方案:
根据本发明的一个方面,提供黄芩苷在治疗冠状病毒感染性疾病中的应 用,所述冠状病毒为人冠状病毒HCoV-229E,所述冠状病毒感染性疾病为人 冠状病毒HCoV-229E感染引起的疾病。
可选地,所述人冠状病毒HCoV-229E感染引起的疾病包括肺炎、肾炎和 肝炎。
可选地,所述人冠状病毒HCoV-229E感染引起的疾病的给药方式选用雾 化吸入给药。
可选地,所述雾化吸入给药的单次剂量为13.6mg/60kg-54.4mg/60kg。 该给药剂量是以实施例2高、中、低三个剂量雾化吸入给药2ml,并按照小 鼠系数11和人体重60kg计算而得。
可选地,在所述雾化吸入给药的制剂形式为吸入溶液,其中黄芩苷的浓 度为1.25mg/ml-5mg/ml。
可选地,所述人冠状病毒HCoV-229E感染引起的疾病通过HCoV-229E 病毒感染体外培养的人胚肺成纤维细胞MRC-5或感染BALB/c类小鼠模型构 建。
在进行黄芩苷在人冠状病毒HCoV-229E感染体外培养的人胚肺成纤维 细胞MRC-5上的应用过程中,首先进行黄芩苷对体外培养MRC-5细胞的毒 性试验,确定黄芩苷的最大无毒浓度,在最大无毒浓度范围内,将黄芩苷原 药液做2倍倍比稀释成8个浓度进行试验,其中,8个浓度按从大到小的顺 序排列分别为2.5mg/ml、1.25mg/ml、0.625mg/ml、0.31mg/ml、0.16mg/ml、 0.08mg/ml、0.04mg/ml、0.02mg/ml。验证黄芩苷在体外培养细胞MRC-5上对人类冠状病毒HCoV-229E的抑制作用。
可选地,所述黄芩苷在人冠状病毒HCoV-229E感染BALB/c类小鼠模型 中的应用。
在黄芩苷在人冠状病毒HCoV-229E感染BALB/c类小鼠模型中的应用的 试验中,将黄芩苷原药液做2倍倍比稀释共3个浓度进行试验,其中,高浓 度为5mg/ml,中浓度为2.5mg/ml,低浓度为1.25mg/ml。采用雾化方式给 小鼠给药,采用人冠状病毒HCoV-229E感染小鼠肺炎模型,检测药物黄芩 苷在小鼠肺指数及抑制率、肺组织病毒载量、肺组织炎性因子含量、外周血 淋巴细胞百分比评价受试药物的有效性。
本发明技术方案通过在感染人冠状病毒HCoV-229E的体外细胞试验, 感染人冠状病毒HCoV-229E的小鼠试验中验证黄芩苷的新用途,该技术方 案具有新颖性,并进行了黄芩苷在治疗由HCoV-229E病毒感染引起的肺炎 的疗效试验,试验结果显示,黄芩苷在体外对人冠状病毒HCoV-229E感染 细胞有一定的抑制作用;能抑制病毒HCoV-229E诱导产生的TNFα、IL 6 等细胞因子的表达,对感染HCoV-229E病毒的肺炎小鼠肺指数及抑制率中 表现出了良好的药效,能降低感染HCoV-229E病毒的肺炎小鼠肺组织的病 毒载量,升高小鼠外周血淋巴细胞比例,降低小鼠肺组织IL-2、IL-10、TNF- α、IFN-β的含量。
附图说明
图1为黄芩苷治疗感染HCoV-229E病毒的肺炎小鼠的肺指数及抑制率 的结果柱状图;
图2为黄芩苷治疗感染HCoV-229E病毒的肺炎小鼠的肺组织病毒载量 的结果柱状图;
图3为黄芩苷治疗感染HCoV-229E病毒的肺炎小鼠的外周血CD+4T淋 巴细胞百分比的结果柱状图;
图4为黄芩苷治疗感染HCoV-229E病毒的肺炎小鼠的外周血CD+8T淋 巴细胞百分比的结果柱状图;
图5为黄芩苷治疗感染HCoV-229E病毒的肺炎小鼠的外周血B淋巴细 胞百分比的结果柱状图;
图6为黄芩苷治疗感染HCoV-229E病毒的肺炎小鼠肺组织炎性因子IL-2 的结果柱状图;
图7为黄芩苷治疗感染HCoV-229E病毒的肺炎小鼠肺组织炎性因子 IL-10的结果柱状图;
图8为黄芩苷治疗感染HCoV-229E病毒的肺炎小鼠肺组织炎性因子 TNF-α的结果柱状图;
图9为黄芩苷治疗感染HCoV-229E病毒的肺炎小鼠肺组织炎性因子 IFN-β的结果柱状图。
具体实施方式
为使本发明的目的、技术方案和优点更加清楚,下面将结合附图对本发 明实施方式作进一步地详细描述。
实施例1
采用人冠状病毒HCoV-229E感染体外培养MRC-5细胞模型,通过观察 药物的TC0、TC50、IC50、TI,评价药物黄芩苷在体外对人冠状病毒 HCoV-229E的抑制作用。
一、试验材料
1.1、细胞株为人胚肺成纤维细胞MRC-5,购自北京北纳创联生物技术研究 院,本室传代,液氮保存备用。
1.2、病毒株为人冠状病毒HCoV-229E,由中国医学科学院医药生物技术研究 所提供,本实验室传代,-80℃冰箱保存备用。
1.3、受试药物为黄芩苷,浓度为5mg/ml,分子量为446,溶剂为磷酸缓冲盐 溶液(PBS)。
1.4、实验试剂见表1。其中,DMEM是一种含各种氨基酸和葡萄糖的培养基。
表1:
试剂名称 | 生产厂家 | 批号 |
DMEM培养基 | 康宁公司 | 14019014 |
PBS溶液 | 康宁公司 | 14819005 |
胰酶 | 康宁公司 | 110519010 |
胎牛血清 | 康宁公司 | 35081006 |
双抗 | Gibco公司 | 1796440 |
1.5、试验仪器见表2。
表2:
仪器名称 | 生产厂家 | 型号 |
电子分析天平 | Ohaus Corp.Brock.NJ.USA | AR1140 |
CO2培养箱 | Thermo | Thermo-371 |
生物安全柜 | Thermo | ThermoMSC1.2 |
洁净工作台 | airtech | SW-CJ-2FD |
离心机 | eppendorf | Centrifuge 5430 |
全自动细胞计数仪 | countstar | IC 1000 |
倒置显微镜 | olympus | CKX41 |
二、试验方法
2.1、原药液的制备
称取黄芩苷500mg,置于50ml容量瓶中,用PH为7的磷酸盐缓冲溶液 定容至刻度,摇匀,即得10mg/ml。为使溶解度更佳,配置好的溶液进行2 倍稀释,即原药液为5mg/ml,提供药理学实验。
将浓度为5mg/ml的黄芩苷原药液做2倍倍比稀释,分别为2.5mg/ml、 1.25mg/ml、0.625mg/ml、0.31mg/ml、0.16mg/ml、0.08mg/ml、0.04mg/ml、 0.02mg/ml,共做8个浓度加入。
2.2、黄芩苷对体外培养MRC-5细胞的毒性试验
将配制后的8个稀释度药物,加到已长成单层的MRC-5细胞培养板中, 100μL/孔,每个稀释度做4个复孔,同时设正常细胞对照。将培养板置37℃, 5%CO2培养箱中培养,每日倒置显微镜下观察细胞病变情况,连续48h,确 定细胞不出现明显病变的最低稀释倍数(最大无毒浓度TC0),并按 Reed-Muench法计算50%细胞毒性浓度(TC50),见表3和表4。
表3:
表4:
由表3、4可知,黄芩苷的最大无毒浓度TC0为0.078mg/ml,下述试验 所用浓度均在TC0浓度以下进行。
2.3、黄芩苷在体外培养细胞MRC-5上对人冠状病毒HCoV-229E的抑制作用。
取已长成单层细胞的培养板,倒掉培养液,接种HCoV-229E病毒液, 100TCID50,100μL/孔,置37℃,5%的CO2培养箱中吸附1h后,弃去病毒 液,用细胞维持液冲洗细胞面3遍后,依次加入无毒浓度以下6个稀释度的 各药液,100μL/孔,每个稀释度做3个复孔,同时设正常细胞对照和病毒对 照,置37℃,5%CO2培养箱中培养,每日倒置显微镜下观察细胞病变情况, 48-72小时后病毒对照组细胞病变为+++~~++++时记录试验结果。
其中,细胞病变按6级标准判断:
-:细胞生长正常,无病变出现;
±:细胞病变少于整个单层的10%;
+:细胞病变约占整个单层细胞的25%以下;
++:细胞病变约占整个单层细胞的50%以下;
+++:细胞病变约占整个单层细胞的75%以下;
++++:细胞病变约占整个单层细胞的75%以上。
按Reed-Muench计算50%抑制浓度(IC50)和治疗指数(TI),TI=TC50/IC50, 结果见表5、表6。
表5:
表6:
表5、6结果显示:黄芩苷对人冠状病毒HCoV-229E致体外培养MRC-5 细胞病变有一定程度的抑制作用,能抑制病毒HCoV-229E诱导产生的TNF α、IL 6等细胞因子的表达,其作用强度大小与黄芩苷浓度相关,黄芩苷浓 度越大,抑制作用越强,IC50为56ug/ml。
三、实验结论
在无毒浓度下黄芩苷在体外对HCoV-229E感染引起MRC-5细胞病变有 一定程度的抑制作用,其作用强度的大小与黄芩苷浓度相关,黄芩苷浓度越 大,抑制作用越强,治疗指数TI=2。
实施例2
采用人冠状病毒HCoV-229E感染小鼠肺炎模型,从肺指数及抑制率,肺 组织中病毒核酸表达量,肺组织中炎性因子含量,外周血中免疫细胞百分比 4个方面评价黄芩苷对人冠状病毒HCoV-229E小鼠肺炎模型的治疗作用。
一、实验材料
1.1、阳性药物为磷酸氯喹片,购自四川升和药业股份有限公司,批号为 2002114,规格为0.25g/片,口服,0.5g/60kg/天。
1.2、受试药物为黄芩苷,浓度为5mg/ml,透明澄清液体,以雾化吸入方式 给药。
1.3、实验动物为BALB/c小鼠,分别选自雌雄各51只,级别为SPF级,体 重为11±1g,合格证号分别为NO.1100112011015252(雄), NO.1100112011015253(雌),许可证号为SCXK(京)2016-0006,购自北京 维通利华实验动物技术有限公司。
1.4、病毒毒株为人冠状病毒HCoV-229E,由中国医学科学院医药生物技术 研究所提供,本实验室传代,-80℃冰箱保存备用。
1.5、实验试剂见表7
表7:
1.6、实验仪器见表8
表8:
二、实验方法
2.1、药物配制
磷酸氯喹片的配制,见表9:
受试药物的配制,见表10:
2.2、病毒传代
取已长成单层MRC-5细胞的25cm2培养瓶,弃掉培养液,用细胞维持 液冲洗细胞面3遍后,加入细胞维持液5ml,再加入HCoV-229E病毒液200μl, 置37℃5%CO2培养箱中培养72~96h,每日倒置显微镜下观察细胞病变情 况,直至80%细胞出现明显病变(CPE)后,将细胞培养瓶置于-80℃低温冰 箱冻存,病毒液反复冻融3次后,用于病毒滴度测定。
取已长成单层MRC5细胞的96孔板,弃掉培养液,用细胞维持液冲洗 细胞3遍后,按10倍倍比(10-1~10-8)稀释接种不同滴度的HCoV-229E病 毒液,共8个稀释度,100μl/孔,每个稀释度做4个复孔,同时设正常细胞 对照。96孔板置37℃、5%CO2培养箱中培养72~96h,每日倒置显微镜下 观察细胞病变情况,并记录各孔的细胞病变情况。按Reed-Muench法计算50% 细胞病变浓度(TCID50)。
2.3、小鼠模型的构建及给药
BALB/c小鼠102只,按体重随机分为正常对照组,模型对照组,氯喹 对照组,黄芩苷高、中、低剂量组。其中,高、中、低剂量分别为原药液、 1/2倍原药液、1/4倍原药液。正常对照组、模型对照组、氯喹对照组每组10 只,雌雄各半;黄芩苷高、中、低剂量组每组8只,雌雄各半。分组结束后, 除正常对照组外,其他各组小鼠乙醚轻度麻醉后,用100TCID50HCoV-229E 滴鼻感染,50μl/只,隔日1次,共感染2次。初次感染当天,氯喹对照组 以0.09g/kg/d灌胃给黄芩苷,各剂量组小鼠每组20min雾化吸入2ml药液, 正常对照组、模型对照组同等条件下给予蒸馏水灌胃,每天1次,连续4天。 末次给药后一天称量体重后,眼眶采血(抗凝),解剖取肺并称量肺重,-80℃ 保存。
2.4、小鼠肺指数及抑制率计算
小鼠称量体重后取血,解剖取肺组织并称量肺重,计算小鼠肺指数及抑 制率,具体计算公式如下:
肺指数=肺湿重(g)×100/体重(g)
2.5、小鼠肺组织中核酸检测(实时荧光定量RT-PCR法)
2.5.1、核酸裂解处理
将保存于-80℃的小鼠肺组织置于洁净的研钵中,倒入少量液氮,将肺组 织研磨成粉末,粉末置于1.5ml离心管中,加入1ml TRIzol试剂,轻弹管底, 将肺组织粉末重悬;室温水平放置离心管,孵育20min;4℃,12000rpm,离 心10min;移取上清液至新的1.5ml离心管中,加入0.2ml氯仿,用力震摇 15s,室温孵育3min至液体分层;4℃,12000rpm,离心15min;移取上清液 至新的1.5ml离心管中,加入0.5ml异丙醇,混匀,室温孵育30min;4℃,12000rpm,离心10min;弃上清,用75%乙醇1ml洗涤沉淀;4℃,7500rpm, 离心5min;吸尽上清液,短暂干燥RNA沉淀5~10min;用20μl DEPC水 溶解沉淀,-80℃低温冰箱保存。
2.5.2、核酸测定
对照品核酸处理:DEPC-H2O作为阴性对照。阳性对照品进行10、100、 1000倍梯度稀释。试剂配制:取n×18μl HCoV-229E核酸荧光PCR检测混 合液,n×1μl内部对照品,与n×1μl RT-PCR酶(n为反应管数),振荡混 匀数秒,3000rpm离心数秒。加样:取上述混合液20μl置于PCR管中,然 后将样品核酸提取液、DEPC-H2O、阳性对照品各5μl分别加入PCR管中,改进管盖,离心数秒使所有液体置于底部,立即进行PCR扩增反应。PCR 扩增:反应管仿制在定量荧光PCR仪上,循环参数设置为:45℃×10min; 95℃×15min;再按95℃×15sec→60℃×60sec,循环40次;单点荧光检测 在60℃,反应体系为25μl。
2.5.3、核酸检测计算方法,见表11。
表11:
Ct值 | 结果判断 |
UNDET或40 | 阴性 |
≦38 | 阳性 |
38~40 | 复检一次,如仍为38~40,则为阴性 |
2.6、小鼠肺组织中炎性因子检测(Elisa法)
2.6.1、样品收集及储存
组织匀浆液样本:称量50mg肺组织加入500μl生理盐水后,使用超声 细胞破碎仪匀浆组织,使用低温高速离心机4℃、10000rpm离心10分钟。 吸取上清液之后分装,保存于-80℃冰箱贮存备用,避免反复冻融。
2.6.2、样本检测
样本准备工作:样本用稀释剂2倍稀释后进行检测。检测:从已平衡至 室温的密封袋中取出微孔板,分别将不同浓度标准品,实验样本或者质控品 加入相应孔中,每孔100μl。用封板胶纸封住反应孔,室温孵育2小时。用 洗液洗板,重复操作4次。最后一次洗板结束,将板倒置,在吸水纸拍干所 有残留液体;在每个微孔内加入100μl酶标检测抗体。用封板胶纸封住反应 孔,室温孵育2小时。重复第4步洗板操作,在每个微孔内加入100μl显色底物,室温孵育30分钟。注意避光。在每个微孔内加入100μl终止液后30 分钟内,使用酶标仪测量450nm的吸光度值,计算结果。
2.7、小鼠外周血淋巴细胞比例检测(流式细胞术)
离心机4℃预冷后,小鼠摘眼球取血,向装有10ml 1×PBS的15ml离 心管中加入3滴血(约150μl),1600rpm,5min,室温离心;用移液管小 心弃去上清,每管加入1ml红细胞裂解液重悬细胞沉淀,室温裂解约5-10min 至液体从浑浊变澄清,加入10ml PBS终止裂解,2000rpm,5min,4℃离心, 弃上清(如果还有较多红细胞可重复进行步骤4)。细胞沉淀用10ml PBS重 悬,2000rpm,5min,4℃离心,弃上清,用200μl封闭液(含5%FBS的 PBS)重悬,并将细胞悬液转移至1.5ml ep管中,4℃封闭30min。避光于 封闭液中配制流式抗体如下:FITC标记抗小鼠CD3e、PE标记抗小鼠CD19, PerCP-Cy5.5标记抗小鼠CD4、APC标记抗小鼠CD8a,每一管细胞的配制体 积为:抗体各0.3μl,封闭液50μl。
细胞悬液2000rpm,5min,4℃离心,弃上清。加入流式抗体,每管50 μl,4℃避光染色30min,加入1ml PBS,2000rpm,5min,4℃离心,弃上 清。用200μl含2%FBS的PBS重悬细胞,转移至流式管中,上机检测(如 不能及时上机检测,可用PBS将4%多聚甲醛固定液稀释为2%,重悬细胞, 每管体积为200μl,4℃避光保存过夜)。
2.8、统计分析
数据采用(MEAN±SD)表示,组间差异采用T检验,P<0.05为差异具 有统计学意义。
三、试验结果
3.1、药物黄芩苷对感染HCoV-229E病毒的肺炎小鼠肺指数及抑制率的影响
表12:
注:与正常对照组比较P<0.01;与模型对照组比较P<0.01
由表12及图1可知:与正常对照组比较,模型对照组小鼠肺指数显著增 加(P<0.01);与模型对照组比较,黄芩苷高、中、低剂量组小鼠肺指数显 著降低(P<0.01),黄芩苷高、中、低剂量组抑制率分别为73.61%、44.32%、 56.17%。可见,黄芩苷高、中、低剂量均表现出了良好的药效。
3.2、黄芩苷对感染HCoV-229E病毒的肺炎小鼠肺组织病毒载量的影响
表13:
注:与正常对照组比较P<0.01;与模型组比较P<0.01
由表13、图2结果可知:与正常对照组比较,模型对照组小鼠肺组织病 毒载量显著增加(P<0.01);与模型对照组比较,黄芩苷高、中剂量组小鼠 肺组织病毒载量显著降低(P<0.01)。
3.3、黄芩苷对感染HCoV-229E病毒的肺炎小鼠外周血淋巴细胞比例的影响
表14:
注:与正常对照组比较P<0.01;与模型对照组比较P<0.05
由表14、图3、图4、图5结果可知:与正常对照组比较,模型对照组 小鼠外周血CD4+T淋巴细胞、CD8+T淋巴细胞百分比显著降低(P<0.01); 与模型对照组比较,黄芩苷高剂量组小鼠外周血CD4+T淋巴细胞、CD8+T淋 巴细胞百分比显著升高(P<0.05)。
3.4、黄芩苷对感染HCoV-229E病毒的肺炎小鼠肺组织炎性因子的影响
表15:
注:与正常对照组比较P<0.05,P<0.01;与模型对照组比较P<0.05,P<0.01
由表15、图6、图7、图8、图9结果可知:与正常对照组比较,模型对 照组小鼠肺组织IL-2、IL-10、TNF-α、IFN-β含量显著升高(P<0.05,P<0.01); 与模型对照组比较,黄芩苷高、中、低剂量组小鼠肺组织IL-2、IL-10、TNF- α含量显著降低(P<0.05,P<0.01),黄芩苷中剂量组小鼠肺组织IFN-β含 量显著降低(P<0.01)。
Claims (8)
1.黄芩苷在治疗冠状病毒感染性疾病中的应用,其特征在于,所述冠状病毒为人冠状病毒HCoV-229E,所述冠状病毒感染性疾病为人冠状病毒HCoV-229E感染引起的疾病。
2.如权利要求1所述的应用,其特征在于,所述人冠状病毒HCoV-229E感染引起的疾病包括肺炎、肾炎和肝炎。
3.如权利要求2所述的应用,其特征在于,所述人冠状病毒HCoV-229E感染引起的疾病的给药方式选用雾化吸入给药。
4.如权利要求3所述的应用,其特征在于,所述雾化吸入给药的单次剂量为13.6mg/60kg-54.4mg/60kg。
5.如权利要求3所述的应用,其特征在于,所述雾化吸入给药的制剂形式为吸入溶液,其中黄芩苷的浓度为1.25mg/ml-5mg/ml。
6.如权利要求1所述的应用,其特征在于,所述人冠状病毒HCoV-229E感染引起的疾病通过HCoV-229E病毒感染体外培养的人胚肺成纤维细胞MRC-5或感染BALB/c类小鼠模型构建。
7.如权利要求6所述的应用,其特征在于,所述黄芩苷在人冠状病毒HCoV-229E感染体外培养的人胚肺成纤维细胞MRC-5上的应用。
8.如权利要求6所述的应用,其特征在于,所述黄芩苷在人冠状病毒HCoV-229E感染BALB/c类小鼠模型中的应用。
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