Establishment and characterization of a liver cell line from black porgy, Acanthopagrus schlegelii

Abstract

In this study, a liver cell line (ASL) derived from black porgy (Acanthopagrus schlegelii) was established and characterised. After the initial primary culture, the cell line has been cultured for over 40 passages at 28 °C in Dulbecco's modified Eagle's medium (DMEM) with 15 % foetal bovine serum (FBS). Immunostaining results indicated that ASL cells contained at least two cytotypes in the initial culture stage; however, epithelioid cells dominated and replaced the other type during passage 6–20. ASL cells grow in temperature range 20–28 °C in DMEM containing 15–20 % FBS. Cytogenetic analysis showed that the modal chromosome number in ASL cells was 48. Partial amplification of the cytochrome c oxidase subunit I gene indicates that ASL originates from black porgy. ASL cells showed a transfection efficiency of approximately 15 % after transfection with the pEGFP-N3 plasmid. Cytopathic effects were observed in ASL cells after red-spotted grouper nervous necrosis virus (RGNNV) infection, and viral replication in RGNNV-infected ASL cells was confirmed using quantitative reverse transcription polymerase chain reaction, virus titre assays, western blot, and transmission electron microscopy assays. Furthermore, several antiviral genes were induced to varying degrees in RGNNV-infected ASL cells, indicating that the immune response was activated in ASL cells post RGNNV infection. In conclusion, the newly established ASL cell line is an ideal in vitro tool for studying NNV-black porgy interactions.