青蒿琥酯体外分次给药对人巨细胞病毒的抗病毒效应  

Antiviral activity research of artesunate against human cytomegalovirus by fractionation dosage method in vitro

作　　者：曾爱红[1] 欧阳颖[2] 郭明明[1] 戴瑄[1] 邹德志[1] 方建培[2] 

机构地区：[1]中山大学附属第一医院急诊科,广州510080 [2]中山大学附属孙逸仙纪念医院儿科

出　　处：《中华实用儿科临床杂志》2014年第22期1703-1707,共5页Chinese Journal of Applied Clinical Pediatrics

基　　金：广东省科技计划项目（2008B030301080）

摘　　要：目的 研究青蒿琥酯（ART）体外对人巨细胞病毒（HCMV）实验室标准株及耐药株的抗病毒作用，了解分次给药法是否能增强ART抗病毒效应。方法 1.采用四甲基偶氮唑盐（MTT）法测定ART的细胞毒性，计算ART对人胚肺成纤维细胞（HEL）的最大无毒浓度（TC0）与半数中毒浓度（TC50）。2.ART抗 HCMV 作用检测：以HCMV标准株及耐药株感染HEL后，再加入含有不同浓度 ART的新鲜培养基，设更昔洛韦（GCV）对照、细胞对照和病毒对照，培养7～10 d，待病毒对照组病变达＋ ＋ ＋ ～ ＋ ＋ ＋ ＋时，采用 MTT比色法测定OD490值，计算受试药物病毒抑制率，Probit回归法计算药物对HCMV的半数抑制浓度（IC50）。3.测定ART分次给药对HCMV AD169株的抗病毒效应：实验分3组，1组：药物总量一次性给予；2组：药物总量分3次，每天给药1次；3组：药物总量分6次，每天给药3次。以GCV组作为对照，MTT法测定各组OD490值，并计算各药物浓度组的病毒抑制率，采用SPSS18.0统计软件，对各组OD值进行单因素方差分析，比较各组间差异。结果 1.ART在62.5 μmol/L以内未见明显细胞毒性作用，TC0和 TC50分别为 62.5 μmol/L和 171.7 μmol/L。2.在5 μmol/L、15 μmol/L及30 μmol/L时，ART与GCV对 HCMV AD169株生长均有明显的抑制作用，二者比较差异无统计学意义，其中GCV IC50为3.49 μmol/L，而ART IC50为2.17 μmol/L，ART治疗指数为28.8，GCV 治疗指数为716.3。另外，ART对 HCMV耐药株生长仍有明显的抑制作用，而GCV抑制作用明显下降，二者比较差异有统计学意义，其中GCV IC50为44.4 μmol/L，ART IC50为2.5 μmol/L。3.在15 μmol/L 及30 μmol/L 浓度下，ART每天 2次给药较每天 1次给药及单次给药，对病毒的抑制率有显著提高，差异有统计学意义（P 〈0.01）；而GCV同样按上述方法给药，则对病毒抑制率影响不大（P 〉0.05）。结论 1.ART浓度在 62.5μmObjective To research the antiviral activity of artesunate （ART） in vitro fighting against both standard laboratory strains and ganeiclovir（ GCV）- resistance strains of human cytomegalovirus （HCMV） and to explore whether fraetionation dosage method can obviously enhance the antiviral effect of ART. Methods 1. Cytotoxicity assay to ART was performed by the use of 3-（4,5-dimethyhhiazol-2-yl）-2,5-diphenyltetrazolium bromide （MTr） colo- rimetry. The 0% toxic concentration （ TC0 ） were determined, and median cytotoxic concentration （ TC50 ） was calculated with Probit regression method. 2. Antiviral activity assays of ART against HCMV：human embryonic lung fibroblast cells （HELs） were infected with standard laboratory strains and GCV- resistance strains of HCMV,respectively, after which virus was removed and overlays of dulbeeco＇s modified eagle medium（MEM） containing different antiviral drugs were added to the wells. All cells were cultured continuously at 37 ℃ in a 50 mL/L CO2 humidified atmosphere for 7 - 10 days and the cytopathic effect （ CPE ） was observed under a microscope. When the degree of CPE was clear （ ＋ ＋ ＋ - ＋ ＋ ＋ ＋ ）, the values of absorbency at 490 nm of all cell wells were measured by MTT colorimetry. The cell survival rate （CSR）and drug inhibitory rate （IR） for HCMV were calculated. By Probit regression method,the me- dian inhibitory concentration （ IC50 ） of 2 drugs was calculated respectively. 3. To explore whether fractionation dosage method could obviously enhance the antiviral effect of ART against HCMV, the experiment was divided into 3 groups and compared with GCV group, respectively： Group 1 ： ART antiviral compounds were added to cell layers by one dosage. Group 2 ：Total drug dosage was divided into 3 parts, and each part was added to cell layers once a day for 3 days. Group 3 ：Total antiviral compounds were divided into 6 and delivery 2 times a day. The values of absorbency at 490 nm of all cell wells were m

关 键 词：人巨细胞病毒 青蒿琥酯 抗病毒效应 

分 类 号：R285[医药卫生—中药学]