Silver nanoparticles inhibit fish gill cell proliferation in protein-free culture medium

Abstract

While short-term exposures of vertebrate cells, such as from fish, can be performed in defined, serum-free media, long-term cultures generally require addition of growth factors and proteins, normally supplied with a serum supplement. However, proteins are known to alter nanoparticle properties by binding to nanoparticles. Therefore, in order to be able to study nanoparticle–cell interactions for extended periods, the rainbow trout (Oncorhynchus mykiss) gill cell line, RTgill-W1, was adapted to proliferate in a commercial, serum-free medium, InVitrus VP-6. The newly adapted cell strain was named RTgill-W1-pf (protein free). These cells proliferate at a speed similar to the RTgill-W1 cells cultured in a fully supplemented medium containing 5% fetal bovine serum. As well, they were successfully cryopreserved in liquid nitrogen and fully recovered after thawing. Yet, senescence set in after about 10 passages in InVitrus VP-6 medium, revealing that this medium cannot fully support long-term culture of the RTgill-W1 strain. The RTgill-W1-pf cell line was subsequently applied to investigate the effect of silver nanoparticles (AgNP) on cell proliferation over a period of 12 days. Indeed, cell proliferation was inhibited by 10 μM AgNP. This effect correlated with high levels of silver being associated with the cells. The new cell line, RTgill-W1-pf, can serve as a unique representation of the gill cell–environment interface, offering novel opportunities to study nanoparticle–cell interactions without serum protein interference.

• Gill
• long-term exposure
• rainbow trout
• serum-free medium
• silver nanoparticles

Acknowledgements

We thank Christopher Buser (EMEZ-Electron Microscopy ETH Zürich, Zürich, Switzerland) for TEM analysis and David Kistler (Eawag, Swiss Federal Institute of Aquatic Science and Technology, Dübendorf, Switzerland) for ICP-MS measurements. We thank Ferruccio Messi (Cell Culture Technologies, Switzerland) for the cell culture media information.

Declarations of interest

The authors declared no personal interests related to this study. The authors alone are responsible for the content and writing of the paper.

Funding information

This work was supported by the Swiss National Science Foundation in the framework of the Swiss National Research Program 64 (NRP 64) on the Opportunities and Risks of Nanomaterials (Project number: 406440-131240).

Supplementary material available online

Supplemental Table S1–S3 and Figures S1–S2.