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作 者:张燕[1,2,3] 鲜雨琦 高磊琼 沈犁 华子瑜[1,2,3] ZHANG Yan;XIAN Yuqi;GAO Leiqiong;SHEN Li;HUA Ziyu(Institute of Pediatrics,Key Laboratory of Child Development and Disorders of Ministry of Education,China International Science and Technology Cooperation Base of Child Development and Critical Disorders,Chongqing Key Laboratory of Pediatrics,Children's Hospital of Chongqing Medical University,Chongqing,400014,China)
机构地区:[1]重庆医科大学附属儿童医院儿科研究所,儿童发育疾病研究教育部重点实验室 [2]儿童发育重大疾病国家国际科技合作基地 [3]儿科学重庆市重点实验室,重庆400014
出 处:《第三军医大学学报》2018年第14期1271-1278,共8页Journal of Third Military Medical University
基 金:国家自然科学基金面上项目(81370777);重庆市卫生计生委中医药科技项目(ZY201702069)
摘 要:目的利用沙眼衣原体(Chlamydia trachomatis,Ct)菌株Ct-L2/Pomp A-GFP,快速筛查黄连素、雷公藤红素、黄芩苷、三羟黄酮体外抑制Ct作用。方法通过CCK-8检测药物细胞毒性作用,得出50%细胞毒性药物浓度(50%cytotoxic concentration,CC50);通过菌株Ct-L2/Pomp A-GFP快速筛查四种中药体外抑制Ct的效果,得出50%抑制率药物浓度(50%inhibitory concentrations,IC50);使用治疗指数(therapeutic index,TI)综合评估药物安全性和有效性。选择安全药物浓度(≥90%细胞存活率),通过活细胞实时观察、免疫荧光染色和末端包涵体形成单位测定,再次确认药物的细胞毒性和体外抑制Ct作用。结果黄连素、雷公藤红素、黄芩苷、三羟黄酮的CC50(mean±SD)分别为(97.0±2.2)、(2.0±0.08)、(557.1±29.1)、(205.8±9.1);IC50(mean±SD)分别为(2.73±0.09)、(0.60±0.02)、(260.0±10.2)、(60.9±3.8);TI分别为35.5、3.3、2.1、3.4,提示黄连素更安全有效。黄连素、雷公藤红素、黄芩苷、三羟黄酮安全药物浓度分别为10.0、0.2、80.0、10.0μmol/L,结果显示黄连素具有明显的Ct抑制作用(P<0.000 1),其他3种药物无明显抑制Ct作用。结论黄连素可能成为抑制Ct安全有效的药物。Objective To compare the inhibitory effects of the effective components derived from 4 traditional Chinese drugs, namely berberine (BER), celastrol (CEL), baicalin, and baicalein, on Chlamydia trachomatis (Ct) CtL2/PompA-GFP strain in vitro. Methods CCK-8 assay was used to determine the 50% cytotoxic concentration (CC50) of the 4 agents in human cervical cancer HeLa 229 cells according to the dose-cytotoxicity curve. The inhibitory effect of each agent on Ct was estimated by observing the decrease in GFP intensity, and the 50% inhibitory concentration (IC50) of each agent was calculated based on the dose-inhibition curve. The therapeutic index (TI) was used to assess both the safety and efficacy of the 4 agents. Using the safe drug concentration (defined as a ≥90% cell viability in HeLa 229 cells), the safety and efficacy of the agents for inhibiting Ct were further assessed by observing the live cells, immunofluorescence assay (IFA) and endpoint inclusion-forming unit assay (IFUs). Results The CC50s of BER, CEL, baicalin and baicalein were 97.0±2.2, 2.0±0.08, 557.1±29.1, and 205.8±9.1, respectively and their IC50s were 2.73±0.09, 0.60±0.02, 260.0±10.2, and 60.9±3.8, respectively. The TIs of BER, CEL, baicalin and baicalein were 35.5, 3.3, 2.1, and 3.4, respectively, showing a better safety and a stronger efficacy of BER among the 4 agents. The safe concentrations of BER, CEL, baicalin and baicalein in HeLa 229 cells were 10, 0.2, 80, and 10 μmol/L, respectively. The results showed that BER could markedly inhibit Ct (P〈0.0001), while the other 3 agent did not produce obvious inhibitory effect on Ct. Conclusion BER can obviously inhibit Ct in vitro and may serve as a potent anti-chlamydia drug with a good safety profile.
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