| 摘要: |
| 目的 检测利福平抗黄热病毒（YFV）感染的效果并初步探索其机制。方法 用不同浓度（0.04、0.2、1、5、25、125、625 μmol/L）的利福平处理人肝癌细胞Huh-7 24 h，通过CCK-8检测利福平的细胞毒性并计算其细胞半数毒性浓度（CC50）。YFV感染Huh-7细胞的同时加入不同浓度（0.04、0.2、1、5、25 μmol/L）的利福平，检测其抗YFV感染的剂量效应，并计算IC50。YFV感染Huh-7细胞的同时加入5 μmol/L利福平，孵育不同时长（2、4、8、12、24 h），检测其抗YFV感染的时间效应。YFV感染Huh-7细胞后，在感染的不同时间段（2、4、6、8、12 h）加入25 μmol/L利福平（药物作用时间2 h，病毒感染2 h），检测其抗YFV感染最显著的起效阶段。利用病毒结合实验、内吞荧光标记实验评价利福平对YFV入侵靶细胞的影响。结果 利福平细胞毒性较弱（CC50为176.9 μmol/L），抑制YFV作用显著（IC50为1.868 μmol/L，P<0.01）；动力时间窗、结合和内吞实验表明，利福平能抑制YFV结合、入侵靶细胞（P<0.01），但不影响YFV的内吞过程。此外，利福平在感染后期的复制阶段也有一定的抑制效果（P<0.01）。结论 利福平可抑制YFV感染靶细胞，作用机制主要通过在病毒感染早期的入侵阶段阻断病毒结合靶细胞。 |
| 关键词: 利福平 黄热病毒 抗病毒药 感染 |
| DOI：10.16781/j.0258-879x.2021.01.0008 |
| 投稿时间：2020-07-22修订日期：2020-11-03 |
| 基金项目:国家科技重大专项重大传染病防治课题（2017ZX10304403-003），国家自然科学基金青年科学基金（31700147），陕西省重点研发项目（2019ZDLSF02-03）. |
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| Antiviral effect of rifampin on yellow fever virus and its preliminary mechanism |
| WU Bing-an1△,QIAN Xi-jing1△,WU Xing-an2,XU Zheng-hao1,LUO Zheng-han1,TANG Hai-lin1,ZHAO Ping1,QI Zhong-tian1* |
(1. Department of Biomedical Defense, Faculty of Naval Medicine, Naval Medical University (Second Military Medical University), Shanghai 200433, China;
2. Department of Microbiology, College of Basic Medical Sciences, Air Force Medical University, Xi'an 710032, Shannxi, China
△Co-first authors.
* Corresponding author) |
| Abstract: |
| Objective To explore the effect of rifampin on yellow fever virus (YFV) and its preliminary mechanism. Methods Indicated concentrations of rifampin (0.04, 0.2, 1, 5, 25, 125, 625 μmol/L) were incubated with human liver cancer cells Huh-7 for 24 h. The cytotoxicity of rifampin was measured by CCK-8 assay and the 50% cytotoxic concentration (CC50) was calculated. When Huh-7 cells were infected with YFV, different concentrations of rifampin (0.04, 0.2, 1, 5, 25 μmol/L) were added to detect the dose-response relationship of YFV resistance and to calculate the half inhibition concentration (IC50); rifampin (5 μmol/L) was added and incubated for different exposure time (2, 4, 8, 12, 24 h) to detect the time-effect relationship of YFV resistance. Huh-7 cells were infected with YFV for 2 h, and rifampin (25 μmol/L) was added at different time points (2, 4, 6, 8, 12 h) for 2 h incubation. The most significant anti-YFV stage was detected. The effect of rifampin on YFV invasion was evaluated by virus binding assay and cholera toxin B (CTB)/transferrin (TF) mediated endocytosis experiment. Results Rifampin had a robust anti-YFV effect (IC50=1.868 μmol/L, P<0.01) with low cytotoxicity (CC50=176.9 μmol/L). Kinetic test, binding and endocytosis experiments showed that rifampin could significantly inhibit YFV binding and invading target cells (P<0.01), but did not affect the endocytosis process of YFV. Moreover, rifampin also had inhibitory effect at late YFV replication stage (P<0.01). Conclusion Rifampin can inhibit YFV infection by blocking viral binding to the target cells at early stage of viral entry. |
| Key words: rifampin yellow fever virus antiviral agents infection |
