Original scientific paper

https://doi.org/10.20471/acc.2025.64.01.10

Optimization of HCV Cell Culture System for Evaluation of Antiviral Drugs against HCV Genotypes 2a and 4a

Naiera M. Helmy ; Microbial Biotechnology Department, Biotechnology Research Institute, National Research Center, Giza, Egypt
Reem El-Shenawy ; Microbial Biotechnology Department, Biotechnology Research Institute, National Research Center, Giza, Egypt
Tawfeek H. Abdelhafez ; Microbial Biotechnology Department, Biotechnology Research Institute, National Research Center, Giza, Egypt
Ashraf A. Tabll ; Microbial Biotechnology Department, Biotechnology Research Institute, National Research Center, Giza, Egypt; Egypt Center for Research and Regenerative Medicine (ECRRM), Cairo, Egypt
Ana Petrović ; Josip Juraj Strossmayer University of Osijek, Faculty of Dental Medicine and Health in Osijek, Osijek, Croatia
Elizabeta Horvatić ; Josip Juraj Strossmayer University of Osijek, Faculty of Dental Medicine and Health in Osijek, Osijek, Croatia
Martina Smolić ; Josip Juraj Strossmayer University of Osijek, Faculty of Dental Medicine and Health in Osijek, Osijek, Croatia
Dominik Gjoni ; Josip Juraj Strossmayer University of Osijek, Faculty of Dental Medicine and Health in Osijek, Osijek, Croatia
Robert Smolić ; Josip Juraj Strossmayer University of Osijek, Faculty of Dental Medicine and Health in Osijek, Osijek, Croatia *

* Corresponding author.

Abstract

Evaluation of hepatitis C virus (HCV) infection in cell culture is important for development
of new antiviral agents and for understanding the viral life cycle, as well as the correlation among
the genotypes spreading worldwide. Development of HCV cell culture systems offers great advantage by
enabling growth of the virus in cell culture, allowing for discovery of potential therapeutics and vaccines.
The aim of this study was to introduce the steps of HCV infection in the cell culture system as a determinant
for the competence of more potent antiviral compounds. Two plasmids, JFH1 (genotype 2a) full
length and 4a (ED43/C-NS2/NS5A) were subjected to in vitro transcription for HCV RNA synthesis
followed by transfection of Huh7.5 with the resulting RNA from each genotype processed to generate
the virus. Later, Huh7.5 cells infected with the replicated virus resulted in the generated HCV infected
cells. Immunostaining of the infected cells showed fluorescence of green color, indicating positive infection
of both genotypes 2a and 4a. In addition, real-time polymerase chain reaction was performed to
measure viral load from extracted cells previously infected with the RNAs of genotypes 2 and 4, which
resulted in 32,500 and 107,375 copies/mL, respectively. In conclusion, introduction of HCV cell culture
system is a convenient method to evaluate active antiviral drugs against HCV genotypes 2a and 4a using
both qualitative and quantitative techniques in distinguishing the effectiveness of antiviral agents.

Keywords

Hepatitis C virus; Infection; Huh7.5; JFH1; Genotype 4

Hrčak ID:

335721

URI

https://hrcak.srce.hr/335721

Publication date:

31.3.2025.

Article data in other languages: croatian

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