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. 2021 May 14;166(7):2005–2010. doi: 10.1007/s00705-021-05096-w

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© The Author(s) 2021

Open AccessThis article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 1 — Anti-HEV activity of ivermectin in Huh7-based cell culture models. (A) Effect of ivermectin treatment for 24, 48, or 72 hours on viral replication-dependent luciferase activity in the genotype 3 subgenomic Huh7-p6-Luc cell model. The untreated group served as a control (CTR) (set as 100%) (n = 12). (B) The 50% inhibitory concentration (IC₅₀) and 50% cytotoxic concentration (CC₅₀) of ivermectin in Huh7-p6-Luc cell model and Huh7 cell line were calculated using GraphPad Prism 5 software (n = 6-12). (C) The infectious Huh7-p6 cell model was treated with the indicated concentrations of ivermectin for 48 hours. The effect on viral RNA production was quantified by qRT-PCR using primers targeting the ORF2/ORF3 overlap region (n = 4-8). (D) Western blot analysis of the HEV capsid protein level in Huh7-p6 cells treated with ivermectin for 48 hours. The uninfected group (mock) served as a negative control, and the infected but untreated group served as a positive control (set as 100%) (n = 4). (E) Immunofluorescence analysis of viral ORF2-encoded capsid protein (red) in Huh7 cells treated with the indicated concentrations of ivermectin for 48 hours. Huh7 cells incubated with an anti-HEV capsid protein antibody or Huh7-p6 cells incubated with the matched IgG control antibody served as a negative control. Untreated HEV-infected Huh7 cells incubated with the anti-HEV capsid protein antibody served as a positive control. DAPI (blue) was applied to visualize nuclei (40 × oil immersion objective). (F) Huh7 cells containing the genotype 1 HEV replicon (Sar55 clone) were treated with ivermectin for 24, 48, or 72 hours, and viral-replication-dependent luciferase activity was measured (n = 10). (G) The effects of long-term treatment with 1 μM ivermectin on HEV RNA replication in the Huh7-p6 cell model were analyzed by qRT-PCR. The untreated (CTR) group served as a control (set as 100%) (n = 3-4). Western blot of the HEV ORF2-encoded capsid protein performed after 21 days of treatment. RLU, relative luciferase units. Data are presented as the mean ± SEM (*, P < 0.05; **, P < 0.01; ***, P < 0.001).