Zebrafish as a Potential Model Organism for Drug Test Against Hepatitis C Virus

Abstract

Screening and evaluating anti- hepatitis C virus (HCV) drugs in vivo is difficult worldwide, mainly because of the lack of suitable small animal models. We investigate whether zebrafish could be a model organism for HCV replication. To achieve NS5B-dependent replication an HCV sub-replicon was designed and created with two vectors, one with HCV ns5b and fluorescent rfp genes, and the other containing HCV's 5′UTR, core, 3′UTR and fluorescent gfp genes. The vectors containing sub-replicons were co-injected into zebrafish zygotes. The sub-replicon amplified in liver showing a significant expression of HCV core RNA and protein. The sub-replicon amplification caused no abnormality in development and growth of zebrafish larvae, but induced gene expression change similar to that in human hepatocytes. As the amplified core fluorescence in live zebrafish was detectable microscopically, it rendered us an advantage to select those with replicating sub-replicon for drug experiments. Ribavirin and oxymatrine, two known anti-HCV drugs, inhibited sub-replicon amplification in this model showing reduced levels of HCV core RNA and protein. Technically, this method had a good reproducibility and is easy to operate. Thus, zebrafish might be a model organism to host HCV, and this zebrafish/HCV (sub-replicon) system could be an animal model for anti-HCV drug screening and evaluation.

Zebrafish: Modeling for Herpes Simplex Virus Infections

“…A similar study used fluorescence-labeled HCV subreplicon injected at 1-8 cell-stage zebrafish larvae to study viral tropism in the liver using florescent microscopy. 24 The same study also used conventional reverse transcriptase-PCR and western blot analysis to confirm the effect of antiviral drugs in zebrafish larvae. 24 We found that creating localized wound or injury to anesthetized adult zebrafish kept in an aqueous medium containing a virus supernatant provides an opportunity to facilitate viral infection, which can be visualized or quantified using reporter viruses (Fig.…”

“…23 Ding et al, generated a zebrafish larvae model expressing NS5B that amplified the HCV replicon showing expression of HCV core RNA and protein. 24 When ribavirin and oxymatrine, two known anti-HCV drugs were used, the amplification of HCV replicon was inhibited suggesting the use of zebrafish larvae model for screening and evaluation of anti-HCV drugs. 24 In a separate study, it was shown that the HCV core protein with thioacetamide could induce HCV in the zebrafish model in a shorter period of time compared with a mouse model.…”

“…24 When ribavirin and oxymatrine, two known anti-HCV drugs were used, the amplification of HCV replicon was inhibited suggesting the use of zebrafish larvae model for screening and evaluation of anti-HCV drugs. 24 In a separate study, it was shown that the HCV core protein with thioacetamide could induce HCV in the zebrafish model in a shorter period of time compared with a mouse model. 25 A recent study by Zhao et al demonstrated that the HCV internal ribosome entry site (IRES) in the zebrafish larvae model mimics mammalian hepatic steatosis and fibrosis at gene levels and, therefore, recommends the feasibility of the zebrafish embryo model for screening anti-HCV drugs targeting HCV-IRES-mediated gene expression.…”

“…30,31,34,35 Although only few human viruses 21,[23][24][25][26][36][37][38][39] have been reported to infect zebrafish, the potential to take advantage of the zebrafish model is growing. There are several advantages of utilizing zebrafish to use microbial infections such as its small size, rapid breeding cycle, low maintenance cost offers large-scale infection analysis that can be very useful in defining infection requirements and confirming them in previously defined mammalian models 40,41 ( Table 3).…”

“…Clearly, zebrafish also provides an inexpensive whole-animal vertebrate model for screening a library of small molecules, cell-or tissue receptorspecific compounds against viral pathogens or anti-angiogenic or anti-inflammatory compounds. 24,[46][47][48][49] One study has already employed this model to confirm the antiviral effect of acyclovir using HSV-1 infected zebrafish. 21 Ideally, zebrafish in a 96-well format can be treated with antibodies 50 or compounds targeting specific cells/tissues or modified antiviral peptides with an objective to have higher efficacy.…”